Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211

Contents lists available at ScienceDirect

Journal of Pharmaceutical and Biomedical Analysis

journal homepage: www.elsevier.com/locate/jpba

Review

Medicinal plants in the treatment of women’s disorders:

Analytical strategies to assure quality, safety and efficacy
Milena Masullo, Paola Montoro, Angela Mari, Cosimo Pizza, Sonia Piacente ∗
Dipartimento di Farmacia, Università degli Studi di Salerno, Via Giovanni Paolo II n. 132, 84084 Fisciano (SA), Italy

a r t i c l e i n f o a b s t r a c t

Article history: During last decades an increasing number of herbal products specifically targeting women’s disorders
Received 12 January 2015 has appeared in the worldwide marketplace. This growth highlights the need for a critical evaluation of
Received in revised form 17 March 2015 quality, safety and efficacy of these products.
Accepted 19 March 2015
Analytical techniques applied to the quality control of the main medicinal plants used for women health
Available online 28 March 2015
(relief of menopause and menstrual related symptoms) have been reviewed. Thanks to the innovation
in analytical technology, identification and detection of secondary metabolites dramatically improved.
Keywords:
In particular, hyphenated techniques have proved to be the most suitable for the rapid identification of
Women’s disorders
Herbal products
compounds in plant matrix. Moreover, taking into account that differences in sample quality are not only
Quality control found in the main compounds or in the chemical markers but also in the low-concentration compounds,
Analytical strategies fingerprint analysis might be a simple way for identification and quality control of herbal products con-
Statistical analysis taining a large number of low amounts of unknown compounds. Furthermore in several papers the
informations obtained from the analysis of a plant have been processed by statistical elaborations.
Medicinal plants here discussed are classified on the basis of the chemical markers used for their quality
control.
© 2015 Elsevier B.V. All rights reserved.

Contents

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 190
2. Analytical methods for plants containing tannins used in the treatment of women’s disorders . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 191
2.1. Alchemilla vulgaris . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 191
2.2. Oenothera biennis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 191
2.3. Potentilla erecta . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 191
3. Analytical methods for plants containing anthocyans used in the treatment of women’s disorders . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 193
3.1. Rubus idaeus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 193
3.2. Vitis vinifera . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 193
4. Analytical methods for plants containing iridoids used in the treatment of women’s disorders . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 194
4.1. Valeriana officinalis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 194
4.2. Verbena officinalis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 194
4.3. Vitex agnus castus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 194
5. Analytical methods for plants containing flavonoids used in the treatment of women’s disorders . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 194
5.1. Capsella bursa-pastoris . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 195

Abbreviations: CE, capillary electrophoresis; DAD, diode array detector; ELSD, evaporative light scattering; ESI, electrospray ionization; FLD, fluorescence detector; GC–MS,
gas chromatography–mass spectrometry; HPLC, high performance liquid chromatography; HPTLC, high performance thin layer chromatography; HR-MS, high resolution
mass spectrometry; HSCC, high-speed countercurrent chromatography; HS-SPME, headspace-solid phase microextraction; IR, infrared spectroscopy; IT, ion trap; LC–MS,
liquid chromatography mass spectrometry; MALDI, matrix assisted laser desorbition ionization; MS, mass spectrometry; NIR, near-infrared diffuse reflectance spectroscopy;
PAD, photodiode array detection; PCA, principal component analysis; PLS-DA, partial least squares discriminant analysis; TLC, thin layer chromatography; ToF, time of flight;
UPLC, ultra-performance liquid chromatography.
∗ Corresponding author. Tel.: +39 089969763; fax: +39 089969602.
E-mail address: piacente@unisa.it (S. Piacente).

http://dx.doi.org/10.1016/j.jpba.2015.03.020
0731-7085/© 2015 Elsevier B.V. All rights reserved.
190 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211

5.2. Carthamus tinctorius . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 195

5.3. Glycine max . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 195
5.4. Medicago sativa . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 195
5.5. Paeonia spp. (Paeoniaceae) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 196
5.5.1. Paeonia lactiflora . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 196
5.5.2. Paeonia officinalis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 196
5.6. Passiflora edulis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 196
5.7. Polygonum spp. (Poligonaceae) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 196
5.7.1. Polygonum cuspidatum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 196
5.7.2. Polygonum hydropiper . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 197
5.7.3. Polygonum aviculare . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 197
5.8. Rubia cordifolia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 197
5.9. Trifolium pratense . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 197
6. Analytical methods for plants containing phenolic acids used in the treatment of women’s disorders . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 197
6.1. Angelica sinensis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 197
6.2. Artemisia spp. (Asteraceae) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 198
6.2.1. Artemisia abrotanum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 198
6.2.2. Artemisia capillaris . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 198
6.2.3. Artemisia frigida . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 198
6.2.4. Artemisia vulgaris . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 198
6.3. Chamaemelum nobile . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 199
6.4. Curcuma spp. (Zingiberaceae) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 199
6.4.1. Curcuma longa . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 199
6.4.2. Curcuma xanthorrhiza . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 199
6.4.3. Curcuma zeodaria . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 199
6.5. Echinacea purpurea . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 199
6.6. Humulus lupulus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 200
6.7. Magnolia officinalis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 200
6.8. Pimenta dioica . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 200
6.9. Piper methysticum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 200
7. Analytical methods for plants containing terpenes used in the treatment of women’s disorders . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 201
7.1. Alisma orientalis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 201
7.2. Calendula officinalis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 201
7.3. Caulophyllum thalictroides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 202
7.4. Cimicifuga racemosa . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 202
7.5. Crocus sativus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 202
7.6. Cyperus rotundus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 203
7.7. Foeniculum vulgare . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 203
7.8. Panax ginseng . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 203
7.9. Pelargonium graveolens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 203
7.10. Salvia spp. (Lamiaceae) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 204
7.10.1. Salvia sclarea . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 204
7.10.2. Salvia miltiorrhiza . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 204
7.11. Tanacetum partenium . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 204
8. Analytical methods for plants containing steroids used in the treatment of women’s disorders . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 205
8.1. Dioscorea spp. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 205
9. Concluding remarks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 205
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 205

1. Introduction raw materials can also be influenced by human adulterations due

to dishonesty or unscrupulous operators. Errors could be acciden-
During last decades an increasing number of herbal products tal botanical substitution (misidentification of plant species) or
specifically targeting women in menopause and with menstrual intentional botanical substitution (deliberate exchange with other,
affections has appeared in the worldwide marketplace. This growth sometimes more toxic, plant species). The variability in the content
highlights the need for a critical evaluation of quality, safety and and concentrations of constituents of plant material, together with
efficacy of these products. the range of extraction techniques and processing steps used by dif-
Ensuring that plant-based products are of suitable quality is ferent manufacturers, results in marked variability in the quality of
important for several reasons. Herbs are natural products and, for commercially available herbal products [1]. Thus quality control of
this reason, they do not have a consistent, standardized composi- herbal products is needed to ensure their consistency, safety, and
tion [1]. efficacy.
Plants contain numerous chemical constituents and if we ana- The current approaches to the quality control of herbal products
lyze different parts of the plant (e.g. roots, leaves), we certainly are either compound-oriented or pattern-oriented [2], the former
find a different qualitative and quantitative profile of constituents. targeting specific components with known chemical structures,
The reason of this variability is that the content and concentration the latter targeting all detectable components. Regarding this lat-
of constituents can be influenced by several factors including cli- ter approach, fingerprint analysis is often accepted by regulatory
mate, growing conditions, time of harvesting, and post-harvesting authorities as a tool to identify herbal formulations and to assess
factors, such as storage conditions (e.g. light, temperature, humid- their quality. A fingerprint is a characteristic profile or pattern
ity) and processing (e.g. extraction and drying). The quality of plant which chemically represents the sample [3].
 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
In the first approach, selection of chemical markers is crucial for
the quality control of herbal products. Ideally, chemical markers
should be components that contribute to the therapeutic effects of
a medicinal plant. Considering that only a small number of chemi-
cal compounds were shown to have clear pharmacological actions,
and a large number of plants are not studied for their bioactive
metabolites, other chemical components can be used as markers.
The European Medicines Agency (EMEA) [4] defines chemical mark-
ers as chemically defined constituents or groups of constituents of
a medicinal plant which are of interest for quality control purposes
regardless of whether they possess any therapeutic activity. The
quantity of a chemical marker can be an indicator of the quality of
a herbal medicine. This point is very important, because the chem-
ical marker approach for quality control is realized by means of
both qualitative and quantitative analyses. The analysis of chem-
ical markers requires specific analytical methods for qualitative
analysis and validated, accurate, precise, and robust methods for
quantitative analysis.
The purpose of this review is to assess the evidence for qual-
ity of the main medicinal plants used for women health (relief of
menopause and menstrual related symptoms) with a specific focus
on their phytochemical composition. Medicinal plants used with
this purpose are classified on the basis of the chemical markers,
not always corresponding to the active constituents, used for the
quality control (Table 1).
2. Analytical methods for plants containing tannins used in
the treatment of women’s disorders
Tannins are a heterogeneous group of high molecular weight
polyphenolic compounds with the capacity to form reversible and
irreversible complexes with proteins, polysaccharides, alkaloids,
nucleic acids and minerals. On the basis of their structural char-
acteristics it is possible to divide them into four major groups:
gallotannins, ellagitannins, complex tannins, and condensed tan-
nins [5].
(1) Gallotannins are tannins in which galloyl units or their meta-
depsidic derivatives are linked to diverse polyol-, catechin-, or
triterpenoid units.
(2) Ellagitannins are tannins in which at least two galloyl units are
C–C coupled to each other.
(3) Complex tannins are tannins in which a catechin unit is glyco-
sidically linked to a gallotannin or an ellagitannin unit.
(4) Condensed tannins are all oligomeric and polymeric proantho-
cyanidins formed by linkage of C-4 of one catechin with C-8 or
C-6 of the next monomeric catechin.
In traditional medicine, the tannin-containing plant extracts are
used as astringents, against diarrhea, as diuretics, against stomach
and duodenal tumors, and as antiinflammatory, antiseptic, antioxi-
dant and haemostatic pharmaceuticals [6]. Anti-inflammatory and
haemostatic activity make this compounds, and principally proan-
thocyanidins, largely used to treat menstrual affection. The activity
of tannins for relieving menopausal symptoms is also known,
and a patent related to the use of proanthocyanidins in relieving
menopausal and perimenopausal symptoms was published [7].
2.1. Alchemilla vulgaris
Alchemilla vulgaris L. (Rosaceae), also known as lady’s mantles
for its ornamental leaves, is a wild plant used for women’s disorders
and reported for the presence of flavonoids and tannins [8]. A. vul-
garis belongs to a number of herbal drugs included in The European
Pharmacopoeia (Ph. Eur.) analyzed only by their tannin content. The
191
tannin content, determined using the non-specific Folin–Ciocalteu
method, an old colorimetric method involving redox chemistry,
is calculated relative to a pyrogallol standard and expressed as
percentage of pyrogallol. Thus, Moller et al. developed a method
based on a reversed-phase gradient HPLC system coupled to DAD,
fluorescence, electrochemical and MS detectors. The HPLC system
developed with UV detection at 250 nm provided characteristic fin-
gerprints of the herbal drugs. Methanolysis of A. vulgaris extracts
generated methyl gallate and ellagic acid, which were analyzed
by HPLC [9]. Successively, a new method based on HPLC coupled
with four kinds of detectors, DAD, FLD, electrochemical ampero-
metric and MS assembled by a micro-splitter valve was developed
for the characterization and differentiation of A. vulgaris extracts
adding 2,5-dihydroxybenzoic acid to the sample solution in order
to eliminate the drift interference of retention time [10].
2.2. Oenothera biennis
Evening primrose (Oenothera biennis L., Onagraceae) is a native
North American traditional medicinal plant. All the parts of this
plant have been widely used as a traditional remedy for several
disorders. It has been receiving a lot of attention all over the world
for its seed oil (up to 25%) rich in polyunsaturated fatty acids mainly
linoleic acid (LA, 60–80%) and ␥-linolenic acid (GLA, 8–14%). These
omega-6 fatty acids are now becoming increasingly popular as
oral and topical remedies for neurodermatitis. Their mechanism
of action is based on the concept that the disease is caused by an
underlying metabolic disorder of long-chained essential fatty acids
which are, in turn, precursors of the highly active prostaglandins
E1 and E2. Evening primrose oil has been tested in several studies
to support its use for breast pain, premenstrual symptoms (PMS),
eczema, cirrhosis, rheumatoid arthritis, menopause [11]. In partic-
ular, it is reported to relieve the discomforts of PMS, menopause,
menstruation, endometriosis and fibrocystic breasts by interfering
with the production of inflammatory prostaglandins released dur-
ing menstruation. Many PMS sufferers are found to have unusually
low levels of GLA, which is why food supplements containing this
fatty acid might help so much. In women with fibrocystic breasts,
essential fatty acids can minimize breast inflammation and pro-
mote the absorption of iodine, a mineral that can be present in
abnormally low levels in women with this condition. In menopause,
it is widely reported that evening primrose oil reduces hot flushes
and increases feelings of well being. The oil of O. biennis must be
subjected to rigorous analyses as part of any quality control pro-
gram. GC is generally applied for the determination of fatty acid
contents (mainly the content of ␥-linolenic acid is dosed), but more
sophisticated methodology may be necessary for the structural
analysis of triacylglycerols, for example. Methods for analysis of
evening primrose oil were reviewed in 1999 [12]. The analysis of
triacilglycerols is generally used for standardization of the oil from
the plant [13].
Although most of the queries for quality control of this species
are focused on the oil, the plant contains as further bioactive com-
pounds ellagitannins, that can have a role as anti-inflammatory
principles relieving menstrual symptoms. Leaves and roots of O.
biennis were investigated for the presence of hexameric and hep-
tameric ellagitannins, by using HPLC coupled with DAD and HR-MS
[14]. A previous analytical approach was carried out with the aim
of determining low molecular weight phenolics with antioxidant
properties from the seeds of the plant [15].
2.3. Potentilla erecta
The rhizome of Potentilla erecta (L.) Raeuschel (Rosaceae),
known with the trivial name of tormentil, is a medicinal and
food source used as nutritional supplement for its effects against
192 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211

Table 1
Medicinal plants discussed in the present review with relative chemical markers, analytical techniques and references.

Plants Chemical markers Analytical techniques References

Alchemilla vulgaris Flavonoids, tannins HPLC-DAD, HPLC-FLD, HPLC-ADC, LC–MS [9,10]

Alisma orientalis Terpenes HPLC-DAD-ESI-MS/MS, UPLC/Q-TOF-MS [206,207]
Angelica sinensis Phenolic acids and phthalides HPLC-UV, HPLC-PAD, HPLC-PAD-MS, [132,133,135,136]
HPLC-PAD-API/MS, UHPLC-QTOF-MS/MS,
HPLC-PAD
Artemisia abrotanum Essential oils, sesquiterpene GC–MS, LC–MS [137,138]
Artemisia capillaris Phenolic, chlorogenic acids, sesquiterpenoids LC–UV, LC-ELSD, LC–MS, UPLC-DAD [140–142]
Artemisia frigida Essential oil GC–MS [143]
Artemisia vulgaris Flavonoids, phenolic acids, HPLC-DAD, HPLC-ESI-QTOF-MS, HPLC-DAD [3,146]
Calendula officinalis Triterpenoid HPLC, HPTLC [208–210]
Capsella bursa-pastoris Flavonoids, caffeoyl acids HPLC-DAD-MS [61]
Carthamus tinctorius Quinochalcones, flavonoids HPLC-DAD [66–68]
Caulophyllum thalictroides Alkaloids, steroidal glycosides triterpene TLC, HPLC, GC, UPLC [211–215]
glycoside
Chamaemelum nobile Spiroethers and coumarins, flavonoids, HPLC-PAD-MS, UHPLC-UV-MS, GC–MS [147,148]
phenolic acids
Cimicifuga racemosa Triterpene glycosides HPLC-UV, HPLC-ELSD, HPLC-PAD, LC–MS, [204,215–218]
SIM-LC–MS, APCI-LC–MS, LC-TIS-MS,
HPLC-TOF-ESI-MS, HPLC-PDA
Crocus sativus Monoterpenes UV–vis spectrophotometry, NIRS, GC–MS, HPLC, [224–231]
LC, FTNIR, MIR, multi-element stable isotope
analysis, SCARs
Curcuma longa Phenolic compounds TLC, HPTLC, NIR, microemulsion electrokinetic [152–166]
chromatography, CE, SFC, DART, LC-ESI-MS/MS
and LC-ESI-(QqQ)-MS, HPLC-DAD and
HPLC-ESI-MS
Curcuma xanthorrhiza Phenolic compounds HPLC–DAD, HPLC-DAD-ESI-MSn [167,168]
Curcuma zeodaria Phenolic compounds HPLC, UPLC-UV-MS [166,169,170]
Cyperus rotundus Sesquiterpenes GC–MS, HPTLC, HPLC-UV, LC-ESI-MS/MS, [233–238]
UHPLC-QTOF-MS,
Dioscorea alata Steroidal saponin UHPLC-ELSD, UHPLC–MS, FT-IR [274,277]
Dioscorea opposita Steroidal saponin UHPLC-ELSD, UHPLC-MS, [274]
Dioscorea villosa Steroidal saponin HSCC-ELSD, HPLC-ELSD, UHPLC-ELSD, [274–276],
UHPLC–MS,
Echinacea purpurea Alkamides, phenylpropanoids, TLC with video Densitometry, HPLC–DAD, [175–179,46,180,181]
polysaccharides, volatile oils, flavonoids LC-ESI-MS, HPLC-ELSD, NMR,
Foenicum vulgare Essential oil GC, GC–MS, Raman spectroscopy, ESI-MS, HPTLC [241–247]
Glycine max Isoflavones HPLC-UV, LC–MS, UPLC, UFLC, NMR [71–79]
Humulus lupulus Prenylchalcones, prenylflavonoids, HPLC-UV, HPLC–DAD, HPLC-ECD, capillary [184–189]
prenylphloroglucinols electrophoresis, HPTLC, HPLC–MS, HPLC–MS2 ,
NMR
Magnolia officinalis Phenolics TLC, HPLC, UPLC-DAD-TOF-MS, NMR [190,191]
Medicago sativa Isoflavones HPLC-UV, HPLC-DAD, HPLC-FLD, LC–MS, [83–90]
GC-IT/MS
Oenothera biennis Fatty acids, tannins GC–MS, HPLC-DAD, HPLC-HRMS [12–15]
Paeonia lactiflora Monoterpenes, phenolic compounds, UPLC-PDA-QTOF-MS, UPLC-QTOF-MS [94–96]
flavonoids
Paeonia officinalis Alkaloids, tannins, saponins, glycosides, TLC, HPLC [98,99]
carbohydrates, flavonoids, terpenes, steroids
Panax ginseng Triterpenes TLC, HPTLC, HPLC-UV, HPLC-DAD, HPLC-ELSD, [250–253]
HPLC-CAD, HPLC-PAD, LC-APCI-MS, LC-ESI-MS
UPLC-QToF-MS, NMR
Passiflora edulis Phenolic compounds, thiols, terpenes, fatty TLC, HPLC, HPLC–MS, GC–MS [32,101–104]
acid esters, alcohols
Pelargonium graveolens Essential oil GC–MS, Vibrational spectroscopy [254,256]
Pimenta dioica Essential oils, phenolic compounds GC, GC–MS, HPLC [193–196]
Piper methysticum Lactones GC, HPLC, DNA based analysis, NIR, [199–203]
Polygonum cuspidatum Quinones, stilbenes HPTLC, HPLC, UPLC-PDA, HPLC-DAD-FICL [106–108]
Polygonum hydropiper Flavonoids 2D-TLC, LC–MS [110,111]
Polygonum aviculare Flavonoids HPLC-DAD-MSn , UHPLC-DAD [112,114]
Potentilla erecta Flavonoids, triterpenoids, organic, phenolic LC–MS, MALDI-TOF-MS [16]
acids, tannins
Rubia cordifolia Anthraquinones HPLC [117]
Rubus idaeus Flavonoids, anthocyans HPLC–DAD, HPLC-ESI-MS, HPLC–MS-MS, [22–33]
MALDI-TOF-MS, MS, GC–MS, HPLC–MS
Salvia sclarea Essential oil GC, GC–MS, TLC [258–261]
Salvia miltiorrhizae Phenolic compounds, diterpenes LC–MS/MS, HPLC-DAD, TLC, [264–266],
Tanacetum parthenium Monoterpenes, sesquiterpene lactones, HPLC-UV, NMR [268–273]
flavonoid glycosides
Trifolium pratense Isoflavones IR, HPLC, HPTLC [123–126]
Valeriana officinalis Sesquiterpenoids, iridoids HPTLC, HPLC-DAD, GC–MS, MS, NMR [44–48]
Verbena officinalis Iridoids ATR-IR, NIR, HPLC, HPLC-DAD-ESI-MS, [49–52]
HPLC-DAD
Vitex agnus-castus Flavonoids HPLC, LC–MS, HR-MS, GC/MS [53,55–58]
Vitis vinifera Flavonoids, anthocyans LC–MS, NMR [35–39]
 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
inflammation, gastrointestinal disorders and pre-menstrual symp-
toms (PMS) [16]. Most of the positive effects can be attributed to
the high amounts of polyphenols in all the plant parts [17]. The
chemical composition of P. erecta is mostly based on the pres-
ence of different flavonoids, triterpenoids, organic and phenolic
carboxylic acids, most of which isolated from the roots and rhi-
zomes. In addiction, P. erecta is considered a tannin-rich plant [16],
for the high concentration of proanthocyanidin oligomers in its
underground parts. In a recent paper, a strategy based on direct flow
injection-ESI-IT-MS has been used to profile proanthocyanidins
(PAs) occurring in this species. To achieve deeper structural infor-
mation and to focus the analysis on PAs with high polimerization
degree (DP), MALDI-ToF-MS, was used. Finally, LC–MS2 analyses
were executed by using a diol stationary phase to detect PAs [16].
3. Analytical methods for plants containing anthocyans
used in the treatment of women’s disorders
Anthocyans (ACNs), i.e. anthocyanins and anthocyanidins,
belong to the group of plant constituents, collectively known as
flavonoids, which occur in the western diet at relatively high
concentrations. ACNs show the ability to scavenge reactive oxy-
gen species and display a variety of pharmacological properties
which make them potential anti-inflammatory agents. The molec-
ular basis for their pharmacological activity includes the regulation
of different mechanisms mainly involved in: (1) suppression of
the inflammatory response through targeting phospholipase A2,
PI3K/Akt, and NF-␬B pathways, (2) protection from cardiovascular
disease, (3) growth/differentiation control and tumor suppression,
(4) reduction of diabetes incidence through modulation of insulin
sensitivity and glucose utilization, (5) neuroprotection through
amelioration of oxidative stress and A␤ deposition, and (6) hep-
atoprotective activity through interference with TNF-␣ and TGF-␤
in the liver. The estrogen-like activity of anthocyans could be uti-
lized in cancer and hormone-replacement therapy. The molecular
mechanisms of protective and therapeutic activity of anthocyans in
various pathological conditions, which may not be attributed solely
to their antioxidant activity but also to direct blockage of signaling
pathways [18]. Related to these important evidences, is the action
on women disorders [19].
3.1. Rubus idaeus
Raspberry (Rubus ideaus L., Rosaceae) flavonoids and mainly
anthocyans have significant antioxidant activity and several
biological activities in moderate and chronic disorders, and
women’s disorders. Differences in anthocyanin composition of
juices obtained from different berry fruits create the possibility of
detecting the adulterations of expensive raspberry and black cur-
rant juices with cheap strawberry and red currant juices on the
basis of anthocyanin analysis [20]. With similar purposes the anal-
ysis of selected organic acids was proposed too [21].
In 1999 flavonoids and anthocyans from R. ideaus and other
selected berries were analyzed by HPLC-ESI-MS. For the identifi-
cation of aglycons, DAD was also used [22]. A paper on the analysis
of anthocyanins was published in 2002 [23]. Anthocyanins from
red raspberries were extracted from the fruit by homogeniza-
tion in acidified methanol. The methanolic extract was centrifuged
and the supernatant analyzed by reversed-phase HPLC. The elu-
ent was monitored at 371 and 520 nm before being introduced
into a single quadrupole mass spectrometer through an atmo-
sphere pressure chemical ionization probe operating in positive
ion mode. This method allowed the identification of eight antho-
cyanins. A method based on tandem mass spectrometry (MS/MS)
coupled with HPLC was described in 2005 [24]. Scan for the
193
precursor ions of commonly found anthocyanidins (cyanidin, del-
phinidin, malvidin, pelargonidin, petunidin, and peonidin) using
LC/MS/MS on a triple quadrupole instrument allowed the specific
detection of each anthocyanin. Further characterization of each
anthocyanin was performed using MS/MS product-ion analysis,
common-neutral-loss analysis, and selected reaction monitoring
(SRM). In 2008 a method for the analysis of anthocyanins from
raspberries by MALDI-ToF-MS was reported [25]. Furthermore,
in 2008 a method based on Direct Introduction MS was pro-
posed for the analysis of polyphenols in berries, including Rubus
[26]. In last years other papers have been published applying
and improving these methods of analysis [27–29]. Also extraction
of phytochemicals from raspberry, principally assisted by ultra-
sound [30] and by microwave [31], has been reported. Investigation
of the fruit flavor by using SPME coupled with stereoselective
GC–MS was reported in 2010 [32]. Authors have used HS-SPME
and GC–MS to determine the enantiomeric ratios of chiral fla-
vor and fragrance indicators in the raw materials and in the
products.
Other studies involve the analysis of ellagitannins [33]. The use
of gradient RP-HPLC with DAD and MSn detection for the analysis
of ellagitannins, ellagic acid conjugates and quercetin conjugates
in raspberries is described. MSn is a particularly powerful tool for
the analysis of trace levels of natural products in the extracts as
interpretation of fragmentation patterns.
3.2. Vitis vinifera
Grape seed (Vitis vinifera L., Vitaceae) proanthocyanidins and
flavonoids are reported to be effective in improving the physi-
cal and psychological symptoms of menopause increasing muscle
mass and reducing blood pressure in middle-aged women [34].
Quality control of V. vinifera products involves principally wine,
the principal product obtained from V. vinifera berries, however in
this context we will evaluate only the improvement in analysis of
grapes or functional foods derived from grapes. In 2008 Cavaliere
and coworkers described a method for the analysis of polyphenols
in grape berries by Rapid Resolution LC–MS [35].
A simple and precise LC–MS method was developed for the
quantification of anthocyanidins in fifteen grape juice samples,
four grape berries and four grape skins [35]. An improved LC–MS
method was published in 2012 by Xu and coworkers: under opti-
mized conditions, five major anthocyanidins including delphinidin,
cyanidin, petunidin, peonidin and malvidin in the hydrolyzed grape
extracts were detected [36].
The application of MS for the analysis of berries polyphenols
has recently been reviewed [37]. MS has been shown to play a
very important role in the research of polyphenols in grape and
wine and in the quality control of products. The soft ionization of
LC–MS makes these techniques suitable to study the structures of
polyphenols and anthocyanins in grape extract and to characterize
polyphenolic derivatives. The coupling of the several MS techniques
presented is shown to be highly effective in the structural charac-
terization of the large number of low and high molecular weight
polyphenols in grape and can be highly effective in the study of
grape metabolomics.
NMR is a technique largely applied to this matrix. Most of these
techniques are able to discriminate adulteration, mainly to identify
the grape fruits’ extract from extracts obtained from other berries;
in fact differences in anthocyanin composition of juices obtained
from different berry fruits create the possibility of detecting the
adulterations [38].
Two headspace-based methodologies have been proposed to
characterize the aroma of grape juice: static headspace (SHS) and
HS-SPME [39].
194 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
4. Analytical methods for plants containing iridoids used in
the treatment of women’s disorders
Iridoids represent a large group of cyclopentapyrane monoter-
penoids that occur widespread in nature, mainly in Dicotyledonous
plant families like Apocynaceae, Scrophulariaceae, Diervillaceae,
Lamiaceae, Loganiaceae and Rubiaceae. Recently, more extensive
studies revealed that iridoids exhibit a wide range of bioactivities,
such as neuroprotective, antinflammatory and immunomodulator,
hepatoprotective and cardioprotective. Iridoids are marker com-
pounds of different species with a specific traditional use against
menstrual symptomatology and menopausal disorders [40].
4.1. Valeriana officinalis
Valeriana officinalis L. (Valerianaceae), valerian, is a bushy plant
whose roots and rhizomes have been used since the 11th cen-
tury for their tranquilizing, menstruation, and sedative effects.
Its antispasmodic activity on smooth muscles has been demon-
strated both in vivo and in vitro [41]. Currently, valerian is
prescribed for reducing pain, cyclic cramps, anxiety, and stress.
It is also used to treat the cramps associated with dysmenor-
rhea; in 2011 a clinical trial has been conducted in this regard
[41]. Phytochemical researches led to the discovery of a number
of sesquiterpenoids and iridoids among which germacrane-type
sesquiterpenoids, volvalerenals A–E and volvalerenic acids A–C,
together with valtrate, isovaleroxyhydroxydihydrovaltrate, 1,5-
dihydroxy-3,8-epoxyvalechlorine, valeriotriate B, jatamanvaltrate
B, jatamanvaltrate C, valerenic acid, and acetoxyvalerenic acid
[42,43].
Analysis of V. officinalis was run principally on essential oils and
iridoids by GC methods. In 2002 V. officinalis was analyzed with
other sedative plants in a sedative herbal preparation by GC–MS
[44]. In 2005 electronic nose was applied with the aim to dis-
criminate several valerian varieties [45]. To overcome the major
limitations of the current methods used for the analysis of tinctures,
a new approach based on NMR spectroscopy and MS was tested
with different tinctures. Diffusion-edited 1 H NMR (1D-DOSY) and
1 H NMR with suppression of the EtOH and water signals were
applied for the first time to the direct analysis of commercial herbal
tinctures derived from Echinacea purpurea, Hypericum perforatum,
Ginkgo biloba, and V. officinalis. The direct injection of the tinctures
in the MS detector to obtain the corresponding metabolic profiles
was also performed. Using both NMR and MS methods it was possi-
ble to obtain metabolic fingerprints which allowed to differentiate
tinctures prepared with different plants [46]. In a recent paper the
application of HPTLC to the analysis of the extracts of V. officinalis
was explored [47]. A strategy based on multi-wavelength chro-
matography fingerprinting of herbal materials, using HPLC with
a UV–vis diode array detector has been applied to the analysis
of V. officinalis. The enhanced fingerprints were constructed by
compiling into a single data vector the chromatograms from four
wavelengths (226, 254, 280 and 326 nm) [48].
4.2. Verbena officinalis
Verbena officinalis L. (Verbenaceae) is a medicinal plant
traditionally used because of its diuretic, expectorant and anti-
rheumatic effects. Moreover, anti-inflammatory, analgesic and
antioxidant activities have been reported for this plant [49].
Analgesic and anti-inflammatory activity justify its use to relieve
menstrual symptoms. Main secondary metabolites present in
the plant are iridoids, phenylpropanoids, flavonoids, triterpenes
and monoterpenes, with verbenalin (iridoid) and verbascoside
(phenylpropanoid) as the major constituents [49]. The European
Pharmacopeia defines verbenalin as the quality determining
compound with a minimum content of 1.5%. For this reason,
quality control of V. officinalis often is run with respect of this
marker compound. Several analytical techniques like HPLC, HPTLC,
GC and micellar electrokinetic capillary chromatography for
qualitative or quantitative analysis of the chemical constituents
in V. officinalis has been reported [49]. Among these reports, Bilia
and coworkers developed an HPLC-DAD-ESI-MS method for the
analysis of the constituents of aqueous preparations of verbena
and lemon verbena and the evaluation of their antioxidant activity
[50]. Successively, simultaneous determination by HPLC of four
bioactive compounds in Verbena officinalis L. has been reported by
Liu and coworkers [51]. A recent paper proposes a method based on
Attenuated-Total-Reflectance IR (ATR-IR) and NIR in hyphenation
with multivariate analysis to quantify verbenalin and verbascoside
in V. officinalis. In addition a HPLC method as a reference was estab-
lished and validated and compared with the spectroscopic method
[49]. Identification of V. officinalis, as described above, is performed
based on morphological and phytochemical analyses, which unfor-
tunately are not reliable enough to distinguish V. officinalis from
other relevant species of the genus Verbena. Thus, the most impor-
tant adulterations could remain undetected. In 2009 a method
was proposed based on comparison of ITS (Internal Transcribed
Spacers) sequences and molecular markers (RAPD) [52].
4.3. Vitex agnus castus
Vitex agnus-castus L. (Verbenaceae), chasteberry, is one of the
most popular and effective herbs used for the prevention and
treatment of pre-menstrual syndrome (PMS) [53] and hyperpro-
lactinemia because of its hormone-like effects [54]. The chemical
composition of the unpolar fraction is characterized by the
presence of diterpenoids, essential oils and ketosteroids, while
flavonoids are described in polar extracts [53–55]. Among these
latter, casticin, a methoxylated flavone, is considered by European
Pharmacopoeia (Ph. Eur.) the reference standard for standardiza-
tion of dry extracts of the species. An analytical HPLC method for the
quantitative determination of casticin in V. agnus-castus fruits has
already been developed [56]. Recently several studies have been
published reporting the use of LC–MS techniques for the quality
control of the polar fractions of V. agnus-castus [55] fruits and a
comparison in the chemical composition of different part of plants
was performed by HR-MS [53].
Many analytical studies are reported about V. agnus-castus
essential oils, most of them realized by GC–MS techniques [57,58].
5. Analytical methods for plants containing flavonoids used
in the treatment of women’s disorders
Flavonoids are polyphenolic compounds ubiquitous in nature.
More than 4000 flavonoids have been recognized, many of which
occur in vegetables, fruits and beverages like tea, coffee and fruit
drinks. Flavonoids occur as aglycones, glycosides and methylated
derivatives. Small amount of aglycones are frequently present and
occasionally represent a considerably important proportion of the
total flavonoid compounds in the plant. Flavonoids have gained
much attention because of their broad biological and pharmacolog-
ical activities including antimicrobial, cytotoxic, anti-inflammatory
as well as cancer preventing activities and principally the capacity
to be powerful antioxidants which can protect the human body
from free radicals and reactive oxygen species. Antiinflammatory
and antioxidant activity make these compounds largely used to
treat menstrual affection. Among flavonoids, isoflavonoids con-
stitute a characteristic and very important subclass of flavonoids.
Their structures are based on the 3-phenylcromen skeleton that
is chemically derived from the 2-phenylchromen skeleton by an
 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
aryl-migration mechanism. Structurally, isoflavonoids can be clas-
sified according to the oxidation of the C15 skeleton, their
complexity and the internal formation of the heterocyclic rings.
Due to their ubiquitous distribution in food and the claimed benefi-
cial health effects of foods containing isoflavones, their distribution
in foods and their healthy properties have been reviewed [59].
Isoflavones have been proposed to have estrogenic activity and play
a putative role in the control of menopause disorders.
5.1. Capsella bursa-pastoris
Capsella bursa-pastoris (L.) Medik. (Brassicaceae), also known as
shepherd’s purse, is a wild herb with high nutritional value that
can be eaten raw or cooked. It is reported in the list of the plants
used for women’s disorders. Investigation of different extracts of its
aerial parts led to the identification of phenolic compounds quan-
tified by HPLC-DAD, organic acids, and amino acids detected by
HPLC-UV, and free fatty acids and sterols analyzed by GC-IT-MS.
The plant material was found rich in kaempferol-3-O-rutinoside,
quinic acid, arginine, palmitic acid, and ␤-sitosterol [60]. Karioti
and coworkers [61] developed an HPLC-DAD-MS method to ana-
lyze the “Olivis” preparation, a blend of four herbal drugs, namely
Crataegus oxyacantha L., Olea europea L., Capsella bursa-pastoris L.
and Fumaria officinalis L. The lack of the marker constituents of
some of the declared plant species (i.e. C. bursa-pastoris) and the
presence of banned adulterants, responsible for the strong antihy-
pertensive effect of the supplement prompted to analyze this herb.
The analyses proved the presence of indole alkaloids belonging to
the group of Rauwolfia sp., such as ajmaline, reserpine and yohim-
bine. Seven flavonoids and two caffeoyl acid derivatives were found
in C. bursa-pastoris [61].
5.2. Carthamus tinctorius
Carthamus tinctorius L. or safflower, commonly called Honghua
in China, is an annual or biennial herbal plant belonging to the
family Compositae. Safflower is considered to promote blood
circulation, to remove blood stasis, promote menstruation and
alleviate pain. In the aspect of clinical practice, safflower is
mainly applied for blood-stasis syndrome with dysmenorrhea,
amenorrhea, post-partum abdominal pain and mass [62]. Many
chemical constituents such as quinochalcones, flavonoids, alka-
loids, polyacetylenes, alkane-diol, fatty acids, steroids, lignans, have
been isolated from safflower. Among them, quinochalcones and
flavonoids are considered as the biologically active constituents of
safflower. The quinocalchone hydroxysafflor yellow A is considered
as one of the standard compounds to evaluate the quality of crude
drug safflower and related preparations containing safflower; also
kaempferide is chosen as standard to evaluate the quality of saf-
flower. Several reports regard the quality control of Chinese herbal
preparation like “Danhong”, “Shu-Jin-Zhi-Tong”, “Xuebijing” with
C. tinctorius as constituent [63–65].
However, the two marker compounds might be insufficient to
fully illustrate the quality of safflower since 104 compounds have
been isolated and identified [62]. So, Li developed an HPLC-DAD
method for the simultaneous determination of four marker com-
pounds, kaempferol-3-O-rutinoside, safflomin A, safflomin B and
bidenoside C, in the extract of the flowers of C. tinctorius [66].
Fan and coworkers [67] developed an HPLC-DAD method to eval-
uate the quality of 46 batches of C. tinctorius from different areas
through a simultaneous quantitation of 10 components. Significant
variations were found in the content of these compounds in these
tinctures. A fingerprinting method has been developed to describe
the chemical constituents and to control the quality of many Chi-
nese Material Medicas. HPLC fingerprinting has been applied to
evaluate the quality of safflower from different producing areas,
195
indicating similar chemical profiles for safflower from various habi-
tats [68].
5.3. Glycine max
Soy (Glycine max (L.) Merrill, Fabaceae), a legume originating
from Asia, is widely distributed throughout the world, for preparing
food products. More recently its consumption is linked, among dif-
ferent properties, to the reduction of menopausal symptoms. This
finding has resulted in the development and commercialization of
many functional foods and food supplements based on soy ingredi-
ents. Soy extracts contain a mixture of isoflavones belonging to the
group of phytoestrogens. The major isoflavones in soybeans include
daidzein, glycitein, genistein, their glycosides, glycoside malonates
and glycoside acetates, in which the predominant isoflavone forms
in soybeans and non-fermented soy products are the glycoside mal-
onates, 6 -O-malonylgenistin and 6 -O-malonyldaidzin [69]. Stud-
ies have shown that they play an important role in reducing climac-
teric symptoms in menopausal and postmenopausal women [70].
A recent review reports the sample preparation and anal-
ysis for the quantification of isoflavones in soybeans and soy
foods. Modern techniques including ultrasound-assisted extrac-
tion, pressurized liquid extraction, supercritical fluid extraction and
microwave-assisted extraction, and analysis by HPLC are reported
[71]. In the quality control of soy the amount of isoflavones,
both aglycones and glycosides, is usually determined by means
of reversed-phase HPLC-UV [72]. Although the HPLC method
s have some advantages when they are applied to the analysis
of isoflavones in terms of specificity, sensitivity, and straightfor-
ward operation, they require a relatively long period of time,
normally from 20 min to 65 min. So, the use of high-throughput
liquid chromatography technologies has been reported in the last
decade for isoflavone analysis, reducing the chromatographic time
to less than 10 min. These techniques allow the use of short
columns, packed with 3 ␮m particles, supporting elevated pres-
sures, thus reducing analysis time, solvent consumption, and,
consequently, environmental impact [73]. Among these studies,
Apers and coworkers developed an HPLC method using two linked
monolithic silica-based reversed-phase C18 columns. This method
for determination of isoflavones in soy extracts needs less than
25 min. Among the high-throughput methods reported in the
literature for isoflavones analysis, UPLC and ultra-fast liquid chro-
matography (UFLC) are cited for their determination in soybeans
cultivars, soy bits, soymilk, texturized soy protein, and soy-based
nutritional supplements [73].
Several research groups have analyzed isoflavones from differ-
ent types of soy seeds by LC–MS [74–76]. Caligiani and coworkers
performed the quali-quantitative determination of isoflavones in
soybean extracts by 1 H NMR [77]. The complexity of natural
soy isoflavones makes the rigorous standardization difficult to
achieve, and most chromatographic methods only quantify the
main isoflavone forms as total isoflavone content [78]. Garcia
and coworkers optimized methods for conventional and perfusion
RP-HPLC to characterize 26 commercial soybean products. Charac-
terization of soybean products was carried out on the basis of their
protein profiles obtained by both chromatographic methods [79].
5.4. Medicago sativa
Alfalfa (Medicago sativa L., Fabaceae) is the main Medicago
species widely grown throughout the world, predominantly as a
source of high quality forage for livestock, renewable energy pro-
duction, phytoremediation and as a source of phytochemicals [80].
It is also used as a human food ingredient, consumed as sprouts in
salads, sandwiches or soups, as leaf protein concentrates or as food
supplements [81]. Despite this use, alfalfa have pharmacological
196 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
activities, being used in some human health disfunctions, and
among these anemia, endometriosis, osteoporosis and menopausal
symptoms [71].
The plant contains many important constituents including
saponins, sterols, coumarins, flavonoids, phenolics, vitamins, pro-
teins, minerals, and other nutrients [82]. As above reported for
Glycine max, to isoflavones present in M. sativa are ascribable some
of the pharmacological activity of alfalfa. Murphy and coworkers
analyzed isoflavones in retail and institutional foods (alfalfa and
soy) by HPLC [83]. Further HPLC [84–86]], ESI-MS [84,85], and CE
[87] methods were employed to analyze flavonoid content in M.
sativa. More recently, two glycosides (daidzin and genistin) and six
aglycones (daidzein, glycitein, genistein, formononetin, prunetin
and biochanin A) were determined by HPLC-DAD in three differ-
ent extracts (aqueous, hydroalcoholic and alcoholic) of M. sativa
[81]. Abo Markeb quantified 5- and 7-hydroxyflavone in the M.
sativa samples by HPLC-FLD [88]. A study compared the isoflavone
production of the callus cell suspension cultures of M. sativa to
the original plants. The extracts were analyzed by LC–MS for
their isoflavones, mainly formononetin, biochanin A, daidzein, and
genistein [89].
In the sprouts of M. sativa and G. max, phenolic compounds,
sterols and triterpenes were determined by HPLC-DAD, organic
acids by HPLC-UV and fatty acids and volatile compounds by GC-
IT/MS [80]. The metabolic profiling of triterpene saponins in M.
sativa was also investigated using HPLC-ESI-MS [90].
5.5. Paeonia spp. (Paeoniaceae)
5.5.1. Paeonia lactiflora
Paeonia lactiflora Pallas, also named Chinese Paeony, is a Chi-
nese herb. A decoction of its root has been used to treat painful or
inflammatory disorders in traditional Chinese medicine.
Different compounds have been isolated from this plant. These
include monoterpenoid glucosides, flavonoids, tannins, stilbenoids,
triterpenoids and steroids, and phenols. Biological activities
include antioxidant, antitumor, antipathogenic, immune-system-
modulation activities, cardiovascular-system-protective activities
and central-nervous-system activities [91].
A water/ethanol extract of Radix Paeoniae is known as total gly-
cosides of paeony (TGP), of which paeoniflorin is the major active
component. Preclinical studies show that TGP/paeoniflorin is able
to diminish pain, joint swelling, synovial hypertrophy, and the
severity of bone erosion and cartilage degradation in experimental
arthritis [92].
P. lactiflora was clinically tested on Polycystic ovary syndrome
(PCOS), a prevalent, complex endocrine disorder characterized
by polycystic ovaries, chronic anovulation and hyperandrogenism
leading to symptoms of irregular menstrual cycles, hirsutism, acne
and infertility [93].
Recently an advanced method has been proposed for the qual-
ity control of this species. In this study, an UPLC- PDA-QToF-MS
based chemical profiling was established for rapid global quality
evaluation of Radix Peoniae. By virtue of the high resolution, high
speed of UPLC and the accurate mass measurement of ToF-MS, a
total of 40 components including 29 monoterpene glycosides, 8
galloyl glucosides and 3 phenolic compounds were simultaneously
separated within 12 min, identified through the matching of for-
mulas with those of published components in a library, and were
further elucidated by adjusted lower energy collision-induced dis-
sociation (CID). The established method was successfully applied
to rapidly and globally compare the quality of Radix Paeonia alba
and Radix Paeoniae rubra, two post-harvesting handled products of
Radix Paeoniae. Thus, UPLC-PDA-QToF-MS based chemical profil-
ing resulted a powerful approach for the global quality evaluation
of Radix Paeoniae [94].
A similar method, integrated with Multivariate Data Analysis
was published in 2013 [95].
An interesting new method, performed directly on the herbal
material, was proposed in 2015 [96]. To assess the inherent quality
of different grades and of different tissues in roots of P. lac-
tiflora, laser microdissection coupled with UPLC-QToF-MS was
applied. The results show that the quantity of the main components
decreased with increase in root diameter from 0.3 cm to 0.7 cm.
Above 0.7 cm of diameter, quantity and diversity of these compo-
nents increased proportionally with the increase in root diameter.
The tissue-specific study indicated that the high content of paeoni-
florin and albiflorin are mainly distributed in the cork and cortex.
According to the results of this study, roots of P. lactiflora greater
than 1.7 cm in diameter can be considered of better quality for
medicinal use than smaller.
5.5.2. Paeonia officinalis
Paeonia officinalis L. is native to South Eastern Europe but it
has been widely introduced elsewhere as a garden plant in many
varieties. Dried and powdered roots of P. officinalis are used as a
medicine in both Indian and Chinese system of medicines. The roots
are cleansed carefully in cold water with a brush and allowed to
remain in water for a short period of time, then they are spread out
on trays in the sun. Phytochemical screening revealed that the roots
of P. officinalis contain alkaloids, tannins, saponins, glycosides, car-
bohydrates, flavonoids, terpenes, steroids and proteins [97]. Peony
root has been used medicinally for over 2000 years and it gained
a reputation as a treatment for epilepsy and to promote menstru-
ation. The root is harvested in the autumn from plants that are at
least two years old and is dried for later use. Quality control of this
species is reported by TLC [98], and by HPLC fingerprint [99].
5.6. Passiflora edulis
Passiflora edulis Sims (passion fruit) belongs to the family Pas-
sifloraceae. P. edulis extracts of the leaves have been used for
centuries as sedatives by native Brazilians, and also for women’s
disorders. A drink from the flower is used to treat asthma, bronchi-
tis, and whooping cough [100]. The leaves of P. edulis, traditionally
used in American countries to treat both anxiety and nervousness
by folk medicine, are rich in polyphenols, which have been reported
as natural antioxidant. Several methods for the quality control of
Passiflora species are reported in literature.
HPLC and HPLC coupled to MS were also approached with the
aim to quantify polar compounds in extracts of P. edulis [102,101].
A method based on GC–MS for the assessment of quality focused
on flavor has been reported [32]. Chemical characterizations can
provide authentication of samples, detection of adulterations, and
differentiation between closely related species. Different methods
based on modern planar chromatography techniques are reported
for P. edulis [102–104].
5.7. Polygonum spp. (Poligonaceae)
5.7.1. Polygonum cuspidatum
Polygonum cuspidatum Sieb. et Zucc., a traditional, popular Chi-
nese medicinal herb, is widely distributed in Southern China and
Japan. The root of Polygonum cuspidatum has been used in the treat-
ment of inflammation, female disorders, infection, jaundice, skin
burns and hyperlipemia diseases [105].
Currently, over 67 compounds from the root of this plant
have been isolated and identified; they are quinones, stilbenes,
flavonoids, coumarins, lignans and others. At present, emodin and
polydatin are used as the marker compounds to characterize the
quality of this plant in the Pharmacopoeia of the People’s Republic
of China [105].
 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
Emodin, resveratrol, and polydatin are the main active compo-
nents of the rhizome. In 2005 a simple densitometric HPTLC method
for quantification of these compounds was reported [106]. The
same compounds, with the addition of physcion, were determined
in 2010 by HPLC [107]. A rapid and accurate UPLC-PDA method was
established for simultaneous detection of 5 compounds including
polydatin, resveratrol, emodin-8-glucoside, emodin, and physcion
[108].
5.7.2. Polygonum hydropiper
Flavones and flavonoid glycosides, such as quercetin galac-
tosides, a sesquiterpene acid, viscosumic acid, oxymethylan-
thraquinones and polygonic acid were identified in all the parts of
Polygonum hydropiper. Plant by its own or mixed with other herbs is
used in the treatment of diarrhea, dyspepsia, itching skin, excessive
menstrual bleeding, hemorrhoids and other diseases [109].
2D-TLC was applied to the quality control of P. hydropiper. Micro
two-dimensional separations were performed on polar bonded sta-
tionary phases of the type cyanopropyl-silica using non-aqueous
eluents as the first direction eluents and aqueous eluents as the sec-
ond direction eluents. The chromatographic process was performed
in micro scale using 5 × 5 cm plates, small volume of eluents and
10 mL of plant extracts to obtain satisfying separation [110]. Anal-
ysis of flavonoids by means of LC–MS was also reported for this
species and proposed for quality control [111].
5.7.3. Polygonum aviculare
Polygonum aviculare L., also known as knotgrass, is an annual
herbaceous plant commonly found in all the continents. P. avicu-
lare L. is widely used as an herbal remedy and has its monograph in
the European Pharmacopoeia. Infusions prepared from the herb of
P. aviculare have been traditionally used in the treatment of upper
respiratory disorders as well as externally as a remedy for skin
affections and female diseases [112]. Previous studies have shown
that extracts from P. aviculare possess anti-inflammatory activity
probably due to flavonoids [113]. There have been several studies
focused on the flavonoid composition of P. aviculare [112,114].
A study by HPLC-DAD-MSn addressed to the flavonoid con-
stituents of knotgrass aerial parts showed that, apart from
well-defined compounds, it contains a series of flavonol glu-
curonides which could not be unequivocally identified without
isolation [112]. It has also been shown that flavonol glucuronides
are dominating constituents in P. aviculare [112].
UHPLC-DAD coupled with ion trap or time of flight mass
detectors together with the analysis of acidic hydrolysis products
allowed a comprehensive determination of flavonoid composi-
tion. Among dominating compounds, the occurrence of myricetin,
kaempferol, isorhamnetin and kaempferide glucuronides was
reported. The developed method can be used as a suitable tool for
a more insightful, metabolome-based standardization of flavonoid
rich fractions of P. avicularis [112].
5.8. Rubia cordifolia
Different classes of compounds were isolated from Rubia
cordifolia L. (Rubiaceae) such as anthraquinones, naphtho-
quinones, bicyclic hexapeptides, terpenes and carbohydrates.
Anti-proliferative and antioxidant activities are reported for R.
cordifolia L., used in Chinese Traditional Medicine for relieving
the symptoms due to endometriosis, such as menstrual abdom-
inal pain, post-menstrual abdominal pain, lumbosacral pain, and
bearing-down pain and distention in inferior belly [115].
Very few report on analytical approaches for this plant are
available in literature. In 2007 a paper focused on the analysis of
anthraquinones was published by Mishchenko et al. [116]. In 2014
a rapid, simple and specific RP-HPLC method has been developed
197
for the quantitative determination of alizarin in the methanolic
extracts of roots and aerial parts of R. cordifolia [117]. In a research
work published in 2010 an attempt was made to establish sys-
tems of standardization of herbal supplements based on R. cordifolia
[118].
5.9. Trifolium pratense
Plants from the genus Trifolium (Fabaceae) have been used in
traditional medicine by many cultures. In Turkish folk medicine,
for example, some Trifolium species are used for their expecto-
rant, analgesic, antiseptic properties and also to treat rheumatic
aches. Some species are also grown as pasture crops for animals
in the Mediterranean. The high quercetin concentration and soy-
asaponin occurrence make the seeds of some Trifolium species
a potential source of health beneficial phytochemicals to be use
in human nutrition. However, T. pratense L. (red clover) has also
gained popularity for the treatment of menopausal symptoms
[119].
In 1965, Schultz showed that biochanin A and formononetin
occur as glycosides in red clover [119]. More recently, isoflavones,
their glycosides, their malonate glycosides and their acetyl
glycosides were determined in red clover extracts using chromato-
graphic and spectrometric methods [119–122].
Quality control of red clover was reviewed in 2004; this
review highlights practical considerations of value to basic sci-
ence and clinical investigators engaged in the study of botanical
supplements. Lessons and examples are drawn from the authors’
experience in designing and developing a red clover standardized
extract for evaluation in Phase I and Phase II clinical trials [123].
Wu and coworkers [124] developed a method for the qual-
ity control of Red Flower Oil preparation, a mixture of several
plant essential oils, among which T. pretense, used in Traditional
Chinese Medicine. Fourier transform IR (FT-IR) was applied and
two-dimensional correlation IR spectroscopy (2D IR) was used to
enhance the resolution. More recently, HPLC [125] and HPTLC [126]
methods were developed for the analysis of T. pretense. Finally, red
clover was included in a classification study, with the aim to clas-
sify some markers of common herbs used in Western medicine
according to the Biopharmaceutical Classification System (BCS)
[127].
6. Analytical methods for plants containing phenolic acids
used in the treatment of women’s disorders
The term “phenolic acids” describes phenols that possess one
carboxylic acid functionality. The naturally occurring phenolic acids
contain two distinguishing constitutive carbon frameworks: the
hydroxycinnamic and hydroxybenzoic structures. Although the
basic skeleton remains the same, the numbers and positions of the
hydroxyl groups on the aromatic ring create the variety. Phenolic
compounds in many plants are polymerized into larger molecules
such as hydrolizable tannins. Moreover, phenolic acids may arise in
food plants as glycosides or esters with other natural compounds
such as sterols, alcohols, glycosides and hydroxyfatty acids. Sev-
eral biological activities are reported for phenolic compounds and
among these activity against menstrual disorders, and activity on
the vascular and uterine smooth muscles.
6.1. Angelica sinensis
The dried root of Angelica sinensis (Oliv.) Diels (Umbelliferae),
known as Danggui in China, is commonly used in Traditional Chi-
nese Medicine (TCM). This herb is used to treat menstrual disorders,
amenorrhea, dysmenorrheal and is in common use in dietary sup-
plements available in China, USA and Europe as health benefits food
198 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
products for women [128]. As for the menstrual cycle and treat-
ment of menopausal symptoms caused by the hormonal changes,
it can produce favorable effects, so it is also known as “female gin-
seng” in Europe [129]. Danggui cultivated in Gansu Province, China,
is considered to be an authentic high-quality herb on the basis of
thousands of years of traditional experience. Several other substi-
tute herbs are also used in clinical applications in other districts
and countries, such as Angelica acutiloba in Japan, Levisticum offic-
inale Koch in Europe, and Angelica gigas in Taiwan. The roots of L.
officinale are called European Danggui and are listed in the German
Pharmacopeia [130].
Over 70 compounds have been separated and identified from
A. sinensis, including those from essential oils (mainly includ-
ing monomeric phthalides), phthalide dimers, coumarins, organic
acids and their esters, polysaccharides, polyacetylenes, vitamins,
amino acids, and others [131]. Recent phytochemical and pharma-
cological studies reveal that the major bioactive components of A.
sinensis are phenolic acids and phthalides [132].
Z-Ligustilide (3-butylidene-4,5-dihydroisobenzofuranone) is
one of the main active components of A. sinensis, which exhibits
significant effects on improving blood fluidity and strong antiox-
idant activity, inhibits the contractile function of the vascular
and uterine smooth muscles and the proliferation of the vascu-
lar smooth muscle cells [133]. Therefore, Z-ligustilide has been
chosen as a ‘marker compound’ to assess the quality of Radix Angel-
icae sinensis and its products [134], together with ferulic acid, also
used to study the pharmacokinetics of A. sinensis [131]. Various
analytical methods have been reported to analyze the chemical
constituents of A. sinensis [133]. Among these methods, HPLC-
UV, HPLC-PAD and HPLC-PAD-MS have been mostly employed to
determine phthalides, ferulic acid, coniferyl ferulate, falcarindiol,
␣-linolenic acid and linoleic acid in A. sinensis. However, using
HPLC-PAD or HPLC-MS to identify chemical constituents, it is still
necessary to directly compare UV spectra or mass spectra and chro-
matographic retention times with standards. Wang and coworkers
[133] developed an HPLC-PAD-API/MS method for analysing the
chemical constituents of A. sinensis. ESI and APCI spectra, in both
positive ion (PI) and negative ion (NI) modes, provided very useful
information concerning the molecular weights of detected com-
pounds, allowing the detection of 15 constituents. Many analytical
methods including LC and GC have been previously developed
for the quality evaluation of DG [132]. However, Z-ligustilide is
a volatile and unstable compound, decomposing rapidly at high
temperature to form other phthalides through oxidation, isomer-
ization, dimerization [133], and therefore GC techniques should
not be suitable for analyzing the thermolabile components such
as coniferyl ferulate and (Z)-ligustilide. Fingerprinting methods
to assess the consistency of A. sinensis dietary supplements were
developed using both HPLC-DAD and flow-injection mass spec-
trometric (FIMS). The components responsible for the chemical
differences were pinpointed by the loadings plots of PCA [135].
Recently, Bai developed an HPLC-DAD combined with UHPLC-
QToF-MS/MS (ultra high performance liquid chromatography
photodiode array detector quadrupole time-of-flight mass spec-
trometry) method for simultaneously determining ten bioactive
components belonging to phenolic acids, alkyl phthalides, hydrox-
ylphthalides and phthalide dimers to quantitatively evaluate the
effect of seven drying methods on the quality of A. sinensis [132].
Cinnamic acid, ferulic acids and derivatives of quercetin were also
analyzed by HPLC-PDA [136].
6.2. Artemisia spp. (Asteraceae)
The genus Artemisia is grown worldwide and includes several
well-known medicinal herbs, and among these A. abrotanum, A.
capillaris, A. frigida and A. vulgaris.
6.2.1. Artemisia abrotanum
Analytical methods are reported for the essential oil and
artemisinin content of A. abrotanum L., by GC–MS and LC–MS
[137,138]. A sample of leaf oil of A. abrotanum, collected in Cuba,
was studied by GC and GC–MS, leading to the identification of fifty-
seven compounds of which trans-sabinyl acetate (33.4%) was the
major component [138].
6.2.2. Artemisia capillaris
A. capillaris Thunb. has been widely used in East Asia
for the treatment of circulatory disorders, such as dysmen-
orrhea [139]. It is reported to contain phenolic compounds,
chlorogenic acid analogs [140], and sesquiterpenoid deriva-
tives (arteannuin B, artemisitene, artemisinin, dihydroartemisinic
acid and artemisinic acid) [141]. Avula developed LC-UV,
LC-ELSD and LC–MS analytical methods for the quantita-
tive determination of sesquiterpenoids from various species
of Artemisia samples, among which A. capillaris [141]. Zhao
reported the presence in A. capillaris of nine chlorogenic acid
analogs (chlorogenic acid, cryptochlorogenic acid, neochlorogenic
acid, 3,5-dicaffeoyl-quinic acid, 4,5-dicaffeoylquinic acid, 3,4-
dicaffeoylquinic acid, chlorogenic acid methyl ester, cryptochloro-
genic acid methylester, neochlorogenic acid methylester) by a
LC–MS technique [140]. Chlorogenic acid, 3,5-di-O-caffeoylquinic
acid, 4,5-di-O-caffeoylquinic acid, jaceosidin, and eupatilin were
detected in A. capillaris by UPLC-DAD analysis and comparison
with A. annua was performed by multivariate analytical methods
[142].
6.2.3. Artemisia frigida
GC–MS analysis was performed on the essential oil isolated from
the aerial parts of A. frigida Willd., showing an high content of 1,8-
cineole and camphor [143].
6.2.4. Artemisia vulgaris
Artemisia vulgaris L., commonly known as mugwort, is a
shrub of temperate zones of Europe, Asia, North Africa, and
North America. A. vulgaris is reported as uterine stimulant.
Phytochemical analysis of A. vulgaris showed that the major chem-
ical constituents were eudesmanolides (sesquiterpene lactones),
essential oils (such as cineole, wormwood oil, thujone), triter-
penes, coumarin, flavonoids [144]. The presence of flavonoids
eriodictyol and apigenin in A. vulgaris with their weak estro-
genic activity, may account for its folkloric use to treat menstrual
disorders, as an emmenagogue and uterine relaxant. Further-
more, relaxing activities in other smooth muscles, the ileum
and the trachea, are reported. Although not directly tested on
the uterus, the smooth muscle relaxing effect could explain
why it has been reported to be an uterine antispasmodic
[145].
Melguizo-Melguizo et al. [146] reported the presence of 22 com-
pounds, of which 15 were phenolic compounds, mainly chlorogenic
acid derivatives or flavonoids, using an HPLC coupled ESI-QToF-MS.
In this report the most abundant compound was found to be 3,5-O-
dicaffeoylquinic acid. Also protocatechuic acid and quinic acid were
present in high amounts while flavonoids showed a significantly
lower concentration.
Alaerts and coworkers obtained fingerprints by a HPLC-DAD
method aimed to the distinction, identification and quality control
of four different Artemisia species, i.e. A. vulgaris, A. absinthium, A.
annua and A. capillaris samples. The lowest similarity between the
fingerprints of the four species occurred at 214 nm. The distinction
of the four species of Artemisia was visualized by PCA in score plots
[3].
 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
6.3. Chamaemelum nobile
Adulteration of commercial chamomile products is one of
the most significant drawbacks in the promotion of herbal
chamomile products. Roman chamomile (Anthemis nobilis, syn.
Chamaemelum nobile L. (All.), Asteraceae) is known to contain
several classes of biologically active lipophilic and hydrophilic
molecules including essential oils, coumarins and several polyphe-
nols (primarily the flavonoids). The main constituents of the Roman
chamomile oil have been reported to be primarily angelate, tiglate
and butyrate esters [147]. In addition, the Roman chamomile
oils often contain monoterpene and sesquiterepene deriva-
tives. Among these biologically important molecules, spiroethers
and coumarins are of interest due to their unique biological
activity profiles. Of the two spiroether isomers, the cis form
was reported to exhibit more potent antispasmodic and anti-
inflammatory activity than the trans form and herniarin is reported
to possess spasmolytic activity [148]. Ma and coworkers devel-
oped an HPLC-PAD-MS method to validate and quantitatively
determine cis-en-yn-dicycloether and trans-en-yn-dicycloether
as well as a major coumarin compound, hernarin [148]. The
use of HPLC-PAD-MS allowed the isolation and identification of
other constituents such as umbelliferone, phytol, luteolin-7-O-␤-
glucoside, (Z)-2-␤-glucopyranosyloxyl-4-methoxycinnamic acid,
(E)-2-␤-glucopyranosyloxyl-4-methoxycinnamic acid, apigenin-7-
O-␤-glucoside, apigenin-7-(6 -O-acetyl)-glucoside, umbelliferone
7-O-␤-glucoside, and 4,5-dicaffeoylquinic acid. Flowers contained
large amount of spiroethers with higher contents of trans-
spiroether than the cis form in both flowers [148]. Avula and
coworkers used a method based on high separation efficiency
UHPLC with UV and QToF detection for the quantification of
phenolic compounds (cis-GMCA), chlorogenic acid, trans-GMCA,
quercetagetin-7-O-␤-glucopyranoside, luteolin-7-O-␤-glucoside,
apigetrin, chamaemeloside apigenin-7-(6 -O-acetyl)-glucoside,
apigenin, and one poly-acetylene (tonghaosu) in C. nobile [149].
Roman chamomile samples confirmed the presence of chama-
maeloside and apigenin as major compounds. PLS-DA was used to
discriminate between commercial chamomile samples.
An analytical method based on the GC–MS analysis was devel-
oped by Wang [147] and applied to the analysis of non-polar
compounds in various chamomile samples.
6.4. Curcuma spp. (Zingiberaceae)
6.4.1. Curcuma longa
Curcuma longa L., also known as turmeric, is a plant native of
Southern Asia and is cultivated extensively throughout the warmer
parts of the world. The drug is represented by the rhizome and
tuberous root. Among its several activities, it is most commonly
used in postpartum recovery, and is reported to treat excessive
vaginal discharges and menstrual disorders [145]. Turmeric is a
spice most subjected to adulteration since it is frequently traded
in ground form. Turmeric powder is adulterated with rhizomes of
cheaply available Curcuma species especially with those containing
the coloring pigment curcumin such as Curcuma zedoaria (Christm.)
Rosc. or ‘yellow shoti’ syn. Curcuma xanthorrhiza Roxb. ‘Manjakua’
and Curcuma malabarica Vel. leading to toxicity and poor quality of
the product [150].
Characteristic constituents responsible for the therapeutic
effect of C. longa are the curcuminoids, namely curcumin,
desmethoxycurcumin and bisdesmethoxycurcumin [151]. The
three curcuminoids are the basis for the quality control of C. longa
and derived preparations.
Quali-quantitative analysis of curcuminoids by different meth-
ods including TLC [152], HPTLC [153,154], NIR spectroscopy [155],
microemulsion electrokinetic chromatography [156], CE [157]
199
supercritical fluid chromatography [158], and direct analysis in real
time (DART), an ion source technique [159] has been reported.
Methods based on LC-ESI-MS2 [160–162] and LC coupled with
a hybrid triple quadrupole linear ion trap [163] were employed.
HPLC-DAD and HPLC-ESI-MS methods were also developed to ana-
lyze the tinctures of turmeric [164]. Since by HPLC is difficult to
produce complete separation of the three curcuminoids (special
stationary phase is needed) and the analysis is time consuming
(10 min at least), UPLC methods for simultaneous quantification
of the three curcuminoids were developed [165,166].
6.4.2. Curcuma xanthorrhiza
Curcuma xanthorrhiza Roxb. is one of the most commonly used
ingredients in Indo-Malaysian traditional medicines. The known
curcuminoids (curcumin, demethoxycurcumin and bisdemethoxy-
curcumin) of C. xanthorrhiza were quantified by HPLC [167], and
later Ruslay and coworkers developed an HPLC-DAD and HPLC-
DAD-ESI-MSn to identify in the rhizome of this species the three
compounds [168].
6.4.3. Curcuma zeodaria
Curcuminoids of C. zeodaria Rosc. have been analyzed by HPLC
[169,170] and by UPLC-UV-MS methods [166].
GC–MS methods were developed for qualitative and/or quanti-
tative determination of volatiles of C. longa, C. xanthorrizha, and C.
zeodaria [171,172].
6.5. Echinacea purpurea
Echinacea purpurea (L.) Moench (Asteraceae), also known as the
purple coneflower, is an herbal medicine that has been used for
centuries, for the treatment of common cold, coughs, bronchitis,
upper respiratory infections, and some inflammatory conditions.
Recently papers reporting a use in circulation disorders, feminine
endocrine disorders and menstrual symptoms have been published
[173,174]. The three species currently used are Echinacea angus-
tifolia DC, E. pallida (Nutt.) Nutt. and E. purpurea [175]. Due to
their close taxonomic alliance, it is difficult to distinguish between
them and incidences of incorrectly labeled commercial products
have been reported. The main chemical constituents include alka-
mides, phenylpropanoids, polysaccharides and volatile oils, as well
as minor constituents such as flavonoids. The chemical compo-
sition determined using LC is traditionally used to establish the
quality of plant material and preparations, as well as to iden-
tify the species [175]. Different analytical methods were applied
to this topic, and in particular in 2003 HPLC coupled with UV
photodiode-array detection and LC-ESI-MS were developed for the
simultaneous analysis of caffeic acid derivates and alkamides in
the roots and extracts of E. purpurea. The method was proposed for
the quality control of plant material and extracts [176]. An accurate
analysis focused on caffeic acid derivatives was proposed in 2004 by
Li et al. The method was applied to 16 different commercial prepa-
rations and proposed for quality control [177]. HPLC coupled with
mass spectrometry was applied successively to the simultaneous
analysis of caffeic acid derivatives and alkamides [178]. E. purpurea
was distinguished from E. angustifolia by means of HPLC coupled
to ELSD, and the method was applied to raw material and extracts
[179]. NMR was proposed as an alternative quality control method:
Diffusion-edited 1 H NMR (1D-DOSY) and 1 H NMR with suppression
of the EtOH and water signals were applied for the first time to the
direct analysis of commercial herbal tinctures derived from E. pur-
purea [46]. UPLC was explored for the same purpose in 2011 [180],
and in 2013 commercial products of E. purpurea were analyzed for
their content in phenylpropanoid by TLC with video Densitometry
[181]. Finally, a chemometric approach was proposed to process
200 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
data obtained from hyperspectral imaging of roots and leaves of
raw material of E. purpurea [182].
6.6. Humulus lupulus
Humulus lupulus L. (Cannabaceae) is well-known throughout
the world as the raw material in the brewing industry. The female
inflorescences (hop cones or “hops”) are widely used to preserve
beer and to give it a characteristic aroma and flavor. In addition hop
cones have long been used for medicinal purposes. In particular,
hop preparations were mainly recommended for the treatment
of sleeping disorders, as a mild sedative, and for the activation of
gastric function as bitter stomachic [183]. In addition, hop extracts
reduce hot flushes in menopausal women, and a placebo-controlled
study on the use of a standardized (8-prenylnaringenin) hop extract
revealed that the daily administration in menopausal women
decreased the incidence of hot flushes and other discomforts
associated to estrogen deficiency (sweating, insomnia, heart palpi-
tation, irritability). Moreover vaginal dryness in postmenopausal
women was significantly reduced by the topical application of a gel
containing hyaluronic acid, liposomes, vitamin E and hop extract
[183].
Hop cones are characterized by a unique and complex pool
of secondary metabolites, comprising both prenylflavonoids and
prenylphloroglucinols. Three classes of compounds are of par-
ticular relevance in relation to bitterness intensity, sensorial
properties and health benefits: prenylchalcones (xanthohumol,
desmethylxanthohumol), prenylflavanones (isoxanthohumol, 6-
prenylnaringenin, 8-prenylnaringenin) and prenylphloroglucinols,
also known as bitter acids or hop acids [183]. The well-known
8-prenylnaringenin has been shown to be one of the most
potent phytoestrogens currently known. Therefore, xanthohu-
mol, isoxanthohumol, 6-prenylnaringenin, 8-prenylnaringenin are
appropriate active constituents for the chemical standardization of
hop dietary supplements to be used by menopausal women, except
the chalcone desmethylxanthohumol which is unstable and readily
cyclizes to form 6-prenylnaringenin and 8-prenylnaringenin [184].
Several analytical methods to quantify these compounds in var-
ious matrices, such as hop extracts, hop products, beer, dietary
supplements, human urine, serum, rat plasma, tissues have been
developed [185]. As regards hop extracts, methods based on HPLC-
UV, HPLC coupled ECD, CE and HPTLC were applied for the analysis
of prenylated flavonoids [185]. Prenylflavonoids and hop bitter
acids in hop extracts have also been analyzed using HPLC–MS [186].
Magalhães developed a methodology based on the sample
purification by adsorption of phenolic compounds from the matrix
to polyvinylpolypyrrolidone (PVPP) and subsequent desorption of
the adsorbed polyphenols followed by the analysis of the extract
by HPLC-DAD and HPLC-ESI-MS2 [184].
Recently, Prencipe and coworkers developed an analytical
method for the metabolite fingerprinting of bioactive compounds
in hop, together with a simple extraction procedure. Different
extraction techniques, including maceration, heat reflux extraction
(HRE), ultrasound-assisted extraction (UAE) and microwave-
assisted extraction (MAE) were compared in order to obtain a
high yield of the target analytes. The analysis of hop constituents,
including prenylflavonoids and prenylphloroglucinols (bitter acids)
was carried out by means of HPLC-UV/DAD, HPLC-ESI-MS and MS2
[187].
A broad spectrum of preparation and analytical liquid chro-
matography techniques coupled with mass spectroscopy (MS)
have been applied to the analysis of various proanthocyanidins
in hops [188]. Fingerprints of commercial cultivars of H. lupu-
lus have been also obtained coupling 2D NMR datasets with PCA
[189].
6.7. Magnolia officinalis
Magnoliae officinalis Cortex (Houpo) is the dried stem bark,
root bark or branch bark of Magnolia officinalis Rehd. et Wils
(Magnoliaceae). It has been used as a TCM for more than 2000
years for the treatment of epigastric stuffiness, vomiting and diar-
rhea, abdominal distention and constipation, cough and dyspnea
[190]. Neolignans, sesquiterpenes, sesquiterpene neolignans, and
phenylpropanoids have been identified in M. officinalis. Oligomeric
neolignans present in the plant are linked through the aromatic
rings. The quality control of this herbal medicine is currently based
on the assay of two active biphenolic compounds, namely mag-
nolol (M) and honokiol (H) by TLC or HPLC. Advanced methods for
the analysis of M. officinalis cortex were proposed in last years.
An UPLC-DAD-ToF-MS fingerprinting method was developed for
the quality control and source discrimination of Cortex Magnoliae
officinalis produced in Zhejiang Province (Wen-Hou-Po). Data were
statistically evaluated using similarity analysis, hierarchical cluster
analysis (HCA) and discriminant analysis [191].
In 2012 a method based on 1 H NMR coupled with chemometric
analysis was proposed to identify the metabolites contributing to
the differences between the samples and to discriminate different
medicinal parts and geographical origins of these samples. The cor-
relation between the data from 1 H NMR and HPLC was performed
with the mixOmics software based on an unsupervised method.
Honokiol and magnolol were the main compounds responsible for
the discrimination of samples from different batches, thus prov-
ing that the choice of these two compounds as markers for quality
assessment by HPLC is relevant. The two sources of Magnoliae offic-
inalis Cortex recorded in the Chinese Pharmacopoeia, M. officinalis
and M. officinalis var. biloba, could be differentiated by 1 H NMR data,
but the pattern recognition analysis by PLS-DA was unsuccessful
in discriminating samples from various geographical origins. The
combination of 1 H NMR that gives a comprehensive profile of the
metabolites and HPLC that targets two biomarkers is an efficient
mean for the quality control of M. officinalis Cortex [190].
6.8. Pimenta dioica
Pimenta dioica (Merr.) L., syn. Pimenta officinalis (Berg) L. (Myr-
taceae) is widely distributed in West Indies, Mexico, and South
America and traditionally known as allspice, pimenta, pimento,
clove pepper and Jamaica pepper. The plant has been cultivated
in Egypt, where it is known as “fulful afrangi”. It is traditionally
used as a spice and condiment and as a remedy against menstrual
disorders, while it is industrially used for tanning purposes and
as flavoring and perfuming agent in soaps, tonics, as well as for
appetizing medicines. The plant shows high content and biological
diversity of the active constituents, phenolic acids, flavonoids, cat-
echins, galloylglucosides, phenylpropanoids, diterpenes and lupeol
[192].
Analytical approaches to this species, involve both essential oils
and phenolic compounds. For the first aspect, in 2013 Amma and
coworkers studied extensively essential oils by using a compara-
tive approach based on GC and GC–MS [193]. In 2006 analysis of
the essential oils was approached by multidimensional GC [194].
Polyphenolic compounds were studied and analyzed in a recent
paper that deals with the separation of 27 known compounds and
two new polyphenolic glucosides from the berries of Pimenta dioica
[195]. Standardization of preparations obtained from P. dioica was
gained both based on essential oil or salycilic acid [196].
6.9. Piper methysticum
Kava (Piper methysticum G. Forster) is the name of a plant and
drink traditionally prepared by macerating its roots in cool water
 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
or coconut water [197]. It has been used for many centuries in
the South Pacific and Hawaii for social ceremonies, relaxation,
medicine, and a multitude of other purposes [197]. More recently,
standardized kava extracts, containing 30% active constituents,
have been used globally as an anxiolytic [197]. Additionally, a
tight inverse correlation between high rates of kava consump-
tion and low incidences of cancer for populations in the South
Pacific has been reported. Some evidence points to the efficacy
of black cohosh, exercise, and possibly kava kava (P. methys-
ticum) in the treatment of menopausal symptoms [198]. In the
last two decades various procedures concerning the separation
and detection of kavalactones have been routinely carried out
by GC (without previous derivatization of kavalactones) [199]
and HPLC [200] but most of them are not validated or only
partially validated. Recently, analyses by supercritical fluid chro-
matography and micellar electrokinetic chromatography have also
been reported. Both GC and HPLC can be used for the anal-
ysis of kavalactones with some advantages and disadvantages
for each method. Using GC analysis, methysticin and yangonin,
which are two of the major components, are generally not sepa-
rated. In addition, the high temperature caused the decomposition
of methysticin. Concerning HPLC analyses, the reversed-phase is
generally better because highly reproducible with a very low
detection limit for all compounds even if the quantitative anal-
ysis of the kavalactones by LC needs to be carried out in the
absence of light to prevent the cis/trans isomerization of yango-
nin. Recently analysis of Kavalactone has been reviewed by Bilia
et al. [201].
DNA based analysis was applied for the correct identification of
plant species and subspecies. This was object of a patent in 2007
[202]. NIR spectroscopy coupled with chemiometric methods was
applied to the quality control of kava kava, and in particular classi-
fication by PLS. The measurements were reproducible and showed
repeatability on par with the HPLC method [203].
7. Analytical methods for plants containing terpenes used
in the treatment of women’s disorders
Terpenoids are a class of natural products widespread in nature,
mainly in plants as constituents of essential oils. Their building
block is the hydrocarbon isoprene, CH2 C(CH3 ) CH CH2 . Terpene
hydrocarbons therefore have molecular formula (C5 H8 )n and they
are classified according to the number of isoprene units, as hemiter-
penoids, consisting of a single isoprene unit, monoterpenoids,
consisting of two isoprene units, sesquiterpenes, diterpenes, triter-
penes and tetraterpenoids with three, four, six and eight isoprene
units, respectively. Most of the terpenoids have multicyclic struc-
tures that differ from one another by their functional groups and
basic carbon skeletons. These types of natural lipids can be found
in every class of living things, and therefore considered as the
largest group of natural products. Several medicinal plants for
women’s healthcare contain terpenes which are responsible to
induce menstruation or abortion, to reduce menstrual bleeding
and postpartum hemorrhage and to alleviate menstrual diseases
[145].
The detection of triterpene glycosides using UV is well known
for its insensitivity because of the weak chromophoric function-
ality of triterpene glycosides in the 200–210 nm region. LC–MS or
LC–MS2 methods may work well for the identification and quantifi-
cation of the wide range of triterpene glycosides [204]. However,
such equipment is expensive and may not be readily available.
Thus, the development of an alternative HPLC method with a reli-
able and reproducible detection is desirable. The inexpensive ELSD
may be suitable for the routine analysis of plant matrix containing
triterpenes [204].
201
7.1. Alisma orientalis
Alisma orientalis Sam. belongs to Alismataceae family and its
dried tuber is known in traditional Chinese medicine (TCM) as
Rhizoma Alismatis or Zexie. Terpenes, including protostane triter-
penoids, kaurane diterpenes, and guaiane sesquiterpenes, are the
major chemical constituents of this plant [205]. To date, all pro-
tostane triterpenes occur only in Alisma plants, and they are
considered to be chemotaxonomic markers of the genus. Alisolide,
alisol O and alisol P, alisol A, alisol B, alisol B 23-acetate, 25-O-
methylalisol A, 25-anhydroalisol A 24-acetate, 25-anhydroalisol A,
alisol E 23-acetate, as well as 13␤,17␤-epoxyalisol A were isolated
from the rhizome of A. orientalis.
A. orientalis is a component, together other five herbal medicine
(Paeonia lactiflora, Angelica sinensis, Ligusticum chuanxiong, Poria
cocos, Atractylodis macrocephalae) of Danggui-Shaoyao-San (DSS),
a famous traditional Chinese medicine formula, used as a classical
gynecological remedy in China for centuries. Chen and cowork-
ers developed an HPLC-DAD-ESI-MS2 method for the qualitative
and quantitative analysis of the major constituents of this herbal
product [206].
Chemical profiles of A. orientalis have been investigated by TLC,
HPCE-UV, HPLC-UV, HPLC-ELSD, and HPLC-IT-MS [207]. In all these
reports only three compounds, namely alisol A, alisol A 24-actetate,
and alisol B 23-actetate were isolated and investigated. Liu devel-
oped an UPLC/Q-ToF-MS method for the analysis of protostane
triterpenoids in A. orientalis. 20 components were simultaneously
separated within 7 min, and identified either through comparing
the retention time (Rt) and CID fragmentation behaviors with those
of the reference standards, or matching empirical molecular for-
mulae with those of published compounds. Moreover, Liu reported
firstly the CID fragmentation pathway of protostane triterpenoids
[207].
7.2. Calendula officinalis
Calendula officinalis L. (Asteraceae) is an annual herb, native
of the Mediterranean region, also known as marigold. Its flow-
ers have been widely used in traditional medicine and also used
as female diseases. The major constituents of C. officinalis include
steroids, terpenoids, triterpenoids (isolated in both free and ester
form), flavonoids, phenolic acids and carotenes [208]. Faradiol,
rutin, caffeic acid and chlorogenic acid have all been isolated from
C. officinalis and have shown biological activities. The most potent
anti-inflammatory effects of C. officinalis have been attributed to
the faradiol monoesters, compounds belonging to the triterpenoid
family [208]. Neukirch et al. [209] quantified simultaneously eight
pentacyclic terpenoids using reversed-phase HPLC of 10 varieties
of C. officinalis, showing that the variety Calypso Orange Florensis
produces the largest amounts of the bioactive terpenoids, in partic-
ular of faradiol laurate which is present in the whole flowers of this
variety at levels which are two-fold higher than those previously
determined in the specialized ray florets. Loescher and coworkers
[208] compared HPTLC and HPLC for qualitative and quantitative
analysis of the major constituents of C. officinalis and to investigate
the effect of different extraction techniques on the composition of C.
officinalis extracts from different parts of the plant. They observed
that HPTLC analysis is able to provide useful qualitative data for
the quick determination of key standards in samples, with easy
comparisons to be made between different extracts. Quantitatively,
however, HPTLC data showed high variability which may be due
in part to limitations in the sensitivity of the software. Authors
claimed that HPLC is a more robust method and is preferred for
quantitative analysis.
Several reports on the essential oil composition of C. offici-
nalis showed as the quantitative amount of the monoterpenes,
202 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
sesquiterpenes and sesquiterpene alcohols, varies from country to
country [210].
7.3. Caulophyllum thalictroides
Caulophyllum thalictroides (L.) Michx (Berberidaceae) is a peren-
nial plant that grows in the United States and Canada. This plant
is called blue cohosh and is well-known as a traditional women’s
herb to ease childbirth and to treat uterine inflammation among
Native Americans [211,212]. Traditionally the roots and rhizomes
of C. thalictroides are used for the treatment of menstrual difficul-
ties and to induce uterine contractions. Extracts of the underground
parts of C. thalictroides are used as an herbal dietary supple-
ment for regulation of the menstrual cycle and women’s diseases
and as an antispasmodic [211,212]. Alkaloids (caulophyllumines
A, caulophyllumines B, and magnoflorine), steroidal glycosides
(caulosaponin and caulophyllosaponin), and triterpene glycosides
have been isolated and identified. Saponins in blue cohosh are con-
sidered to be responsible for the uterine stimulant effects together
with teratogenic alkaloids [211,212]. As this regards, there is con-
siderable concern about the safety of blue cohosh with reports of
new born babies having heart attacks or strokes after the mother
consumed blue cohosh to induce labor. Analytical methods have
been reported for the analysis of alkaloids alone or for alkaloids and
saponins. Alkaloid levels were firstly determined by TLC densito-
metric and HPLC methods in the extract of C. thalictroides [213] and
also by GC in dietary supplements that contain blue cohosh [212].
Successively, HPLC methods were developed for the quantitative
analysis of alkaloids and saponins from C. thalictroides roots [214]
and from extracts of blue cohosh roots and dietary supplements
[215]. More recently, analytical methods including HPLC, UPLC and
HPTLC have been reported for the determination of major alkaloids
and triterpene saponins in the roots of C. thalictroides and in dietary
supplements claiming to contain blue cohosh [211].
7.4. Cimicifuga racemosa
Cimicifuga racemosa (L.) Nutt., or Actaea racemosa (Ranuncu-
laceae) commonly known as black cohosh, is native to North
America. The roots and rhizomes have been used traditionally
by Native Americans for gynecological conditions [214]. Various
preparations of this plant have also been used for the treatment
of menopausal disorders in Germany for over 50 years, and are
presently available in USA as dietary supplements. A number of
clinical studies have been published in support of the use of black
cohosh as alternative treatment of menopausal symptoms [214].
Chemically, C. racemosa extracts contain triterpene glycosides,
such as actein, 26-deoxyactein, and phenolic compounds, such as
ferulic acid and isoferulic acid, flavonoids, such as formononetin
[214]. Currently, the standardization of black cohosh preparations
is based on the content of triterpene glycosides, calculated as
26-deoxyactein. In general, RP-HPLC with UV detection is used
for the analysis of C. racemosa samples, but triterpene glyco-
sides of C. racemosa have a weak UV absorbance. Ganzera et al.
reported an HPLC-ELSD method for the determination of actein,
26-deoxyactein, and cimiracemoside A [216] while Li developed
an HPLC method with ELSD and PAD for the quantitative anal-
ysis of 16 constituents of C. racemosa [214]. Avula et al. [217]
developed an HPLC-ELSD method for the analysis of terpenoids in
different C. racemosa samples. Because of the increase in botan-
ical trade between the USA and China in recent years, Chinese
species of Actaea, including ‘shengma’, have become an adulterant
in US-made black cohosh products. Moreover, due to overlap of the
distribution area and similarity of leaf appearance of some species
with black cohosh, some American species of Actaea might also
be misidentified as black cohosh. Canadian Health has found that
several cases of liver toxicity were associated with people who took
black cohosh products adulterated with related herbal species. For
this reason, Jiang et al. [218] evaluated the authencity and described
the phytochemical profile of 11 C. racemosa products by HPLC-DAD
and selected ion monitoring (SIM)-LC–MS.
LC-Atmospheric pressure chemical ionization (APCI)-MS for
triterpene glycosides [219], an LC-turbo ion spray (TIS)-MS method
to examine the LC–MS chromatography or “fingerprint profile” of C.
racemosa samples and derived products [220] have been reported.
Ma et al. studied the metabolite profiling of C. racemosa extracts
by HPLC-ESI-ToF-MS technique and principal component analysis
identifying 15 chemical markers [221]. Moreover, the phytochem-
ical fingerprints of fifteen species of Cimicifuga were established
using HPLC-PDA and LC–MS techniques. Polyphenols and triter-
pene glycosides made the black cohosh clearly distinguishable from
most other species of Cimicifuga [222].
7.5. Crocus sativus
Saffron, which is obtained from the dried red stigmas of Cro-
cus sativus L. (Iridaceae), is the most expensive spice in the world.
It has broad use in the food industry as an additive for coloring
and flavoring foods. It is also employed as a drug in traditional
medicine and in Persian traditional medicine saffron is used to
treat menstrual disorders [223]. Saffron has also been found to be
effective in relieving symptoms of premenstrual symptoms (PMS),
dysmenorrhea and irregular menstruation [223]. The typical color,
taste, aroma and flavor of saffron are determined by the following
compounds: crocins (glycosylated apocarotenoids with glucose,
gentiobiose, neapolitanose or triglucoses as the saccharidic moi-
eties) are responsible for the strong coloring capacity, picrocrocin
(the glucosylated monoterpene precursor of safranal) confers the
bitter taste and safranal (a monoterpene aldehyde derived from
the chemical or enzymatic dehydration of picrocrocin during saf-
fron handling, drying and storage) gives rise to its characteristic
odor and aroma.
The quality of saffron and its commercial value are determined
by specifications described within the ISO/TS-3632 standard [224]
that establishes the spectrophotometric quantification of picro-
crocin and safranal in aqueous saffron extracts by absorbance
measurements at 440, 257 and 330 nm. Unfortunately, this method
presents some disadvantages because safranal is just barely water
soluble and also exhibits adsorption in the same (320–340 nm)
range as cis-crocin isomers [225].
Several works concerning different qualitative aspects of saf-
fron are present in literature; different analytical techniques have
been applied, primarily UV–vis spectrophotometry and NIRS, to
determine the characteristic chemical compounds [224]. Mass
spectrometry combined with GC, HPLC and LC analysis have been
focused on the identification of volatile molecules, coloring pig-
ments or taste compounds [224–226].
Studies reported the investigation of saffron as performed by
MIR (Mid-Infrared spectroscopy) [227], and by multi-element
stable isotope analysis [228]. The recourse to DNA fingerprinting
for food authentication is reported for routine quality con-
trol. A method based on Sequence-Characterized Amplified
Regions (SCARs) was applied to 24 different food products
containing different amounts of saffron in order to detect adul-
teration/contamination with seven common bulking agents [229].
Also NMR combined with PCA was used to discriminate between
Iranian saffron and commercial samples by analyzing methanol
extracts [230], between authentic Greek saffron and four typical
plant-derived materials utilized as bulking agents in saffron, i.e.,
Crocus sativus stamens, safflower, turmeric, and gardenia [231]
and between Italian Protected Designation of Origin (PDO) saffron
from L’Aquila, S. Gimignano and Sardinia and commercial saffron
 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
samples available on the Italian market [224]. Due to its high mar-
ket value, perceived value, demanding production, and premium
price, attempts have been made to adulterate saffron with various
substances with similar color and morphology to increase the
volume and weight of commercial lots. The most frequently incor-
porated materials are Carthamus tinctorius, Calendula officinalis
and Arnica montana flowers, Bixa orellana seeds, Hemerocallis sp.
petals, Curcuma longa rhizomes and Crocus vernus stigmas [229].
7.6. Cyperus rotundus
Cyperus rotundus L. (purple nut sedge), belonging to the fam-
ily Cyperaceae, is a perennial herb, used in Chinese Traditional
Medicine for the treatment of primary dysmenorrhea [232]. Sev-
eral reports have stated the presence of sesquiterpenes, such as
valencene and nootkatone, furochromones, sterols, flavonoids, and
triterpenes in the rhizomes of this plant [233]. GC–MS methods
have been developed to analyze essential oil composition of C.
rotundus [234,235]. Priya Rani developed HPTLC and HPLC methods
with UV detection to identify three sesquiterpenoids solavetivone,
aristolone and nootkatone in the acetone extract of C. rotundus
[236]. Kumar and coworkers studied the hydroalcoholic fraction of
C. rotundus by LC-ESI-MS/MS, identifying the presence of polyphe-
nols, flavonoids and sesquiterpenes [233].
Jaiswal et al. [237] obtained a metabolite profiling and deter-
mined the content of (+)-nootkatone in rhizome of C. rotundus. Laser
dissected tissues, namely, the cortex, hypodermal fiber bundles,
endodermis, amphivasal vascular bundles, and whole rhizomes
were analyzed by UHPLC-QToF-MS. GC–MS analysis was used
for profiling essential oil constituents and quantitation of (+)-
nootkatone. The quantitative determination of ferulic acid in C.
rotundus by HPLC was also reported [238].
7.7. Foeniculum vulgare
Fennel (Foeniculum vulgare Mill., Ombrelliferae) is an aromatic
perennial herb with a deep thick taproot. Different varieties are
cultivated as a spice or vegetable, for an essential oil used to flavor
foods, and in some countries, for medicinal purposes. Few extracts
of F. vulgare and isolated compounds have been evaluated for sev-
eral activities, namely, antiaging, anti-inflammatory, antimicrobial
and antispasmodic [239].
It seems that fennel can be effective in reducing the severity of
dysmenorrhea, but it has an unpleasant taste in view of most of the
volunteers [240].
Most of analytical literature on fennel regards the analysis of
essential oil, markers for quality control of the species. Most of
the analyses were performed by GC and mono and bi-dimensional
GC–MS [241,242].
In 2005 Raman spectroscopy was proposed as an alternative to
GC for the anlaysis of fennel essential oil [243]. Cross-sections of
fennel seeds were investigated by the use of Raman spectroscopy
and Raman mapping to localize the essential oil and to analyze its
chemical composition directly in the plant. Furthermore the practi-
cability of a home-built mobile transportable Raman spectrometer
to perform on-site measurements was successfully tested.
An interesting paper, focused on a metabolomics approach for
quality control of herbs, introduced fennel in the analyses. The
direct infusion of the samples in ESI-MS and the analyses in both
positive and negative ion mode led to a clear differentiation of the
different samples. To verify if the same approach could be effective
also for mixtures of plant extracts, five different commercial dietary
supplements were analyzed by ESI-MS. The data were evaluated
by multivariate data analysis and the obtained results suggested
that the method allows a satisfactory and rapid characterization of
complex mixtures of commercial dietary supplements [244].
203
A simple HPTLC method has been developed for the simultane-
ous quantification of umbelliferone, psoralen, and eugenol in the
fruit of Foeniculam vulgare. The technique enables rapid and sen-
sitive simultaneous analysis in different samples. The method was
validated for precision, repeatability, and accuracy in accordance
with ICH guidelines [245]. Adulteration issues on this species are
reported in literature, and generally for the essential oil the method
with better results in the identification of adulterations is isotopic
ratio MS [246]. Anethole, an active compound of F. vulgare, was also
analyzed in different samples of F. vulgare [247].
7.8. Panax ginseng
Panax ginseng L. roots (Araliaceae) have been widely used in
Chinese traditional medicine since ancient times owing to their
stimulating and tonic properties. Ginseng roots have been used
to treat various diseases, including cancer and cardiovascular dis-
eases, in East Asian countries. Considering these effects, ginseng
may hold value in treating postmenopausal women. A previous
randomized controlled trial (RCT) reported that ginseng extract
alleviated some menopausal symptoms, such as depression, and
also improved general health and well-being [248]. The pharma-
cological activities of ginseng or its crude extracts are based on the
presence of a mixture of triterpenic saponins referred to as gin-
senosides. Ginsenosides are classified into three groups according
to the type of aglycones, i.e., dammarane, ocotillol and oleanane
triterpenes. Furthermore, the dammarane type to which most gin-
senosides belong can be generally classified as protopanaxadiol
(PPD; ginsenosides Rb1 (Rb1), Rc, Rd, Rg3, Rh2, etc.) or protopanax-
atriol (PPT; Rg1, Re, Rg2, Rh1, etc.). Although the sugar chains in the
PPD-type group are attached to C-3 or C-20, the sugar chains in the
PPT-type group are linked to a hydroxyl moiety at C-6 or C-20. In
addition, the two types can be further differentiated based on the
types of sugar chains and the aliphatic chain at C-17 [249].
Methods based on TLC, HPTLC, HPLC coupled with many differ-
ent kinds of detectors, including UV, DAD, ELSD, charged aerosol
detector (CAD), pulsed amperometric (PAD) detectors have been
applied [250] to the analysis of P. ginseng roots. Among these,
ELSD analyses produce good chromatographic parameters, includ-
ing high precision and accuracy, and the limit of detection (LOD)
for ginsenosides has been determined at approximately 100 ng.
CAD was developed as an alternative to ELSD to detect poor
UV-responsive analytes. Using an HPLC-CAD system, Wang et al.
quantified ginsenosides in P. ginseng founding that CAD produced
improved chromatographic parameters, including sensitivity, lin-
earity and reproducibility, over UV and ELSD [251]. MS techniques
have been widely used in the analysis of P. ginseng root [252]. Atmo-
spheric pressure chemical ionization (APCI) and ESI are typically
used in ginseng analysis, with ESI being more common. MALDI has
been used for the mass spectrometric imaging of ginsenoside local-
ization in P. ginseng root [250]. Quadrupole (Q), triple quadrupole
(QqQ), ion trap (IT), ToF and Fourier transform ion cyclotron
resonance (FT-ICR) are used as mass analyzers for ginseng anal-
ysis [250]. Recently, UPLC coupled with quadrupole time-of-flight
mass spectroscopy (UPLC-QToF-MS) has been applied as a power-
ful analytical tool for rapid analysis of the complex components
or metabolites and chemical transformations in ginseng-related
products [249]. Metabolite profiling and fingerprint analysis were
obtained by 1 H NMR spectroscopy to identify potential biomarkers
capable of distinguishing different ginseng species, varieties, and
commercial products [253].
7.9. Pelargonium graveolens
Pelargonium graveolens L. Her.ex Ait (known also as Geranium
graveolens) (Geraniaceae) is used traditionally for the treatment of
204 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
hyperglycaemia in multiple folk medicine systems. Geranium spp.
are also used as astringent, diuretic, antidiabetic, antispasmodic
for stomach troubles and menstrual symptoms, and as gargle for
throatand tonsils. FDA (US Food and Drug Administration) clas-
sified geranium oils as GRAS (Generally Recognised As Safe) for
food use [254]. Hence, geraniums are promoted as important aro-
matic plants and flavoring agents in perfumery, cosmetic, food and
pharmaceutical industries [255]. Quality control of this species is
focused mainly on essential oils, analyzed by GC–MS, but also by
vibrational spectroscopy [254]. The standardization of geranium
oil along with its bioactivity, toxicity and adulterations have been
reported in a review published in 2002 [256].
7.10. Salvia spp. (Lamiaceae)
7.10.1. Salvia sclarea
Salvia sclarea L. (clary sage) is one of the most popular Salvia
species in Turkey and many Countries. Clary sage seed has approx-
imately 29% oil content and this oil contains >50% of ␣-linolenic
acid. This plant, occurring in the Mediterranean basin and Iran, is
one of the most important aromatic plants cultivated worldwide
as a source of essential oil. The essential oil or extracts of the aerial
parts of S. sclarea have a broad spectrum of effects including anal-
gesic, antiinflammatory, antioxidant, antifungal, and antibacterial
[257].
In 2008 clary sage essential oils obtained by hydro-distillation of
dried aerial parts were analyzed by GC and GC–MS [258]. Fifty com-
ponents were characterized in cultivated plants with linalyl acetate
(35.9%), germacrene D (13.3%), linalool (12.8%) and sclareol (9.27%)
as dominating constituents, while 45 constituents were identified
for wild plants with linalyl acetate (34.0%), linalool (18.5%), germa-
crene D (10.0%) and sclareol (8.7%) as the major constituents. In
2011 Yalcin and coworkers published a study evaluating the effect
of ␥-irradiation on bioactivity, fatty acid composition and volatile
compounds of clary sage seed based on the fact that ␥-irradiation
is widely applied in the preservation of spice quality [259].
TLC was in addition proposed for a rapid fingerprint of Salvia
sclarea [260]. Most of the adulterations of S. sclarea are valuable by
essential oil composition and bioactivity [261].
7.10.2. Salvia miltiorrhiza
Radix Salvia miltiorrhizae Bunge, a plant native in China, is listed
in the Chinese Pharmacopoeia with the name of Dan-Shen and used
in Chinese folk medicine for the treatment of different pathologies.
Two classes of major bioactive components in Dan-shen, including
water soluble phenolic acids and lipophilic diterpenoid quinones,
have effectiveness in treating coronary heart disease, heart-stroke,
cerebrovascular diseases, menstrual disorders, miscarriage, hep-
atitis and insomnia. The study of S. miltiorrhiza components is very
complex, therefore selective and efficient analytical methods are
required to simultaneously determine their structures for further
studies of the pharmacological effects and to control the quality of
the preparations. Currently, there are many methods for the deter-
mination of phenolic compounds and diterpenes among which
LC–MS2 was shown to be a powerful method for separation and
identification of individual molecules in complex samples [262].
Cryptotanshinone, tanshinone I and anshinone IIA are impor-
tant active constituents in S. miltiorrhiza Bunge. Cryptotanshinone
is usually used against inflammation, tanshinone I for therapy of
angina pectoris and anshinone IIA for improving blood circulation
[263].
Separation and determination of the constituents of this medic-
inal plant was gained mainly by HPLC–MS. A simple and sensitive
HPLC-DAD method was established and validated to simulta-
neously quantify 7 major saponins, i.e., notoginsenoside R1,
ginsenoside Re, Rg1, Rb1, Rh1, Rg2 and Rd, in “Danshen Dropping
Pills” (DSDP), the best sold traditional Chinese medicine. The
method involved the solid-phase extraction (SPE) and chromato-
graphic separation on a reversed-phase Agilent Zorbax SB-C18
column [264]. In a paper published in 2009 the chemical charac-
teristics of S. miltiorrhiza collected from different locations in China
were revealed, and salvianolic acid B, rosmarinic acid, cryptotan-
shinone, and tanshinones I and IIA were optimized as markers for
the quality control of S. miltiorrhiza. The comparison was done
on the basis of the simultaneous quantitative determination of
13 hydrophilic and lipophilic compounds, by UPLC and hierarchi-
cal analysis. The entire chromatographic pattern showed that S.
miltiorrhiza was significantly different from its adulterant Salvia
przewalskii [265]. A paper reporting TLC analysis fingerprint was
also published in 2006 [266].
7.11. Tanacetum partenium
Feverfew (Tanacetum parthenium L.) (Asteraceae) is a medici-
nal plant traditionally used for the treatment of fevers, migraine
headaches, rheumatoid arthritis, stomach aches, toothaches, insect
bites, infertility, and problems with menstruation and labor dur-
ing childbirth. The feverfew herb has a long history of use in
traditional and folk medicine, especially among Greek and early
European herbalists. The plant contains a large number of natural
products, but the active principles probably include one or more
sesquiterpene lactones, including parthenolide. Other potentially
active constituents include flavonoid glycosides and pinenes [267]
The chemistry of feverfew is now well defined. The most impor-
tant biologically active principles are sesquiterpene lactones, the
principal one being parthenolide. Parthenolide is found in the
superficial leaf glands (0.2–0.5%), but not in the stems, and com-
prises up to 85% of the total sesquiterpene content [267]. More than
30 sesquiterpene lactones have been identified in feverfew. In gen-
eral, there are 5 different types of sesquiterpene lactones, which
may be classified by chemical ring structures. Feverfew contains
eudesmanolides, germacranolides, and guaianolides. Parthenolide
is a germacranolide [267].
This plant is marketed in the United States in a variety of forms
and compositions. Although its therapeutic efficacy is still uncer-
tain, the sesquiterpene lactone parthenolide is the constituent
recommended to be measured for quality control of feverfew
preparations.
Quality control of T. parthenium was firstly proposed in 1992
[268]. Three physicochemical methods (HPLC, NMR spectroscopy,
and HPLC of a derivative) have been used to measure parthenolide
in authenticated Tanacetum parthenium (feverfew) and in several
commercial products. Similar results were obtained for all three
physicochemical assays and also for the bioassay. Authenticated T.
parthenium grown in the UK contained a high level of parthenolide
in leaves, flowering tops and seeds but a low level in stalks and
roots [268].
In 2000 a validated liquid chromatographic method was devel-
oped and used to estimate parthenolide in a number of U.S.
feverfew market products formulated as capsules, tablets, or crude
powder [269]. In the same year, efficacy and safety of this plant and
its preparations was reviewed [270].
A NMR spectroscopic and pattern recognition analytical
approach to investigate composition and variability of different
samples of T. partenium was proposed in 2002. 1 H NMR spec-
troscopy and PCA was used to discriminate between batches
of 14 commercially available feverfew samples based on multi-
component metabolite profiles [271]. Improvement was gained
only in 2013, when a paper describing the analysis by a spectropho-
tometric method of the different sesquiterpene lactones in the
plant was published [272]. Standardization of formulation based
on T. parthenium was developed in 2009. A spray-dried extract was
 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
standardized in the content of the sesquiterpene lactone partheno-
lide. The total flavonoid and parthenolide contents in the
spray-dried extract were 1.31% and 0.76% (wt/wt). The spray-dried
allowed its tableting by direct compression. Tablet properties were
in accordance with the proposed specifications [273].
8. Analytical methods for plants containing steroids used
in the treatment of women’s disorders
Steroids (Greek, stereos = solids), widely distributed in the ani-
mal and plant kingdom, present a tetracyclic system arranged in
the form of a perhydrocyclopentanophenanthrene. They include
great variations in structure and encompass compounds of vital
importance to life, such as cholesterol, the bile acids, sex hormones,
vitamin D, corticoid hormones, cardiac aglycones, antibiotics, and
insect molting hormones.
8.1. Dioscorea spp.
Yam is the common name for rhizomes of plants from the genus
Dioscorea (Dioscoreaceae) which are widely distributed all over the
world. Over 50 steroidal saponins have been identified from various
Dioscorea species. Steroidal saponins, furostanol glycosides such as
methyl parvifloside and protodeltonin and spirostanol glycosides
like deltonin and glucosidodeltonin, are reported to be the major
physiologically active constituents in yam [274]. The most well-
known species of this genus is Dioscorea villosa L., also called wild
yam. The rhizomes and roots of this plant are used for their phytoe-
strogenic properties in the treatment of menstrual and menopause
complaints and are claimed to improve women’s health [274].
Analytical methods for the determination of steroidal com-
pounds using HSCC-ELSD and HPLC-ELSD have been reported for
Dioscorea villosa and Dioscorea spp [275,276]. A UHPLC method
with shorter retention times and good resolution and sensitivity
for quantitative determination of 11 steroidal saponins from the
rhizomes and roots of Dioscorea villosa, D. alata and D. opposite was
developed by Avula and coworkers [274]. Detection of the saponins
was achieved by the use of an ELS detector. A highly sensitive
UHPLC–MS to identify and confirm the structure of compounds in
Dioscorea samples and dietary supplements that claimed to contain
D. villosa was developed. FT-IR spectroscopy combined with mul-
tivariate analysis was used to predict dioscin content from African
yam tubers (D. alata L.) [277].
9. Concluding remarks
As highlighted, several strategies have been applied for the
quality control of medicinal plants employed in the treatment
of women’s disorders. Thanks to the innovation in analytical
technology, identification and detection of secondary metabolites
dramatically improved. In particular, hyphenated techniques
have proved to be the most suitable for the rapid identification
of compounds in plant matrix. Advanced methods have been
proposed for the quality control of these species in order to obtain
specificity, sensitivity, and straightforward operation, reducing
the analysis time. Often, powerful analytical techniques such as
HPLC-DAD-MS, UPLC-MS, HPLC–MS/MS, NMR were employed to
obtain a metabolic fingerprint. Medicinal plants here discussed
were classified, on the basis of the chemical markers used for the
quality control, in plants containing tannins, antocyans, iridoids,
flavonoids, phenolic acids, terpenes and steroids. This classification
allows to select the most suitable analytical technique on the basis
of the chemical composition of the medicinal plant. Qualitative
profiles of the herbal products have been discussed, taking in
consideration that differences in sample quality is not only found
205
in the main compounds or in the chemical markers but also in
the low-concentration compounds, and so fingerprint analysis
might be an interesting way for identification and quality control
of herbal products, containing a large number of low amounts of
unknown compounds. In many cases, quantitative analysis of the
selected medicinal plants showed that the content in chemical
markers can be influenced by several factors including climate,
growing conditions, time of harvesting, and post-harvesting fac-
tors, such as storage conditions and processing and varies not only
in different parts of the plants but also from country to country. In
several papers information obtained from the analysis of a plant
was processed by statistical elaborations. The most widely used
approach is multivariate data analysis, in PCA, a method that allows
the differentiation among groups of samples, providing valuable
information on the origin and/or among technological treatments.
The analytical strategies here reported can be used during the
standardization process with the aim to assure quality, safety
and efficacy to herbal products. In several cases the discussed
analytical approaches allow to explore the eventual adulteration
and substitution with similar common herbs, often characterized
by different bioactivity profiles and lower commercial value.
References
[1] V. Schulz, R. Hänsel, V.E. Tyler, Rational Phytotherapy A Physicians’ Guide to
Herbal Medicine, 4th ed., Springer, Berlin, 2000.
[2] D.K.W. Mok, F.T. Chau, Chemical information of Chinese medicine: a challenge
to chemist, Chemom. Intell. Lab. Syst. 82 (2006) 210–217.
[3] G. Alaerts, S. Pieters, H. Logie, J. Van Erps, M. Merino-Arévalo, B. Dejaegher, J.
Smeyers-Verbeke, Y. Vander Heyden, Exploration and classification of chro-
matographic fingerprints as additional tool for identification and quality
control of several Artemisia species, J. Pharm. Biomed. Anal. 95 (2014) 34–46.
[4] The European Medicines Agency: Reflection paper on markers used for
quantitative and qualitative analysis of herbal medicinal products and
traditional herbal medicinal products. http://www.emea.europa.eu/pdfs/
human/hmpc/25362907en.pdf
[5] P. Schofield, D.M. Mbugua, A.N. Pell, Analysis of condensed tannins: a review,
Anim. Feed Sci. Technol. 91 (2001) 21–40.
[6] L. Giovannelli, G. Carderni, C. Cecchini, S. Silvi, C. Orpianesi, A. Cresci, Red
wine polyphenols influence carcinogenesis, intestinal microflora, oxidative
damage and gene expression profiles of colonic mucosa in F344 rats, Mutat.
Res. 591 (2005) 237–246.
[7] P. Rohdewald, Method and compositions for relieving menopausal and per-
imenopausal symptoms U.S. Pat. Appl. Publ. (2007), US 20,070,269,541, A1
20071122.
[8] L.M. D’Agostino, I. Dini, E. Ramundo, F. Senatore, Flavonoid glycosides of
Alchemilla vulgaris, Phytother. Res. 12 (1998) S162–S163.
[9] C. Moller, S.H. Hansen, C. Cornett, Characterisation of tannin containing herbal
drugs by HPLC, Phytochem. Anal. 20 (2009) 231–239.
[10] Y. He, Q. Wu, S.H. Hansen, C. Cornett, C. Moller, P. Lai, Differentiation of tannin-
containing herbal drugs by HPLC fingerprints, Pharmazie 68 (2013) 155–159.
[11] N. Sekeroglu, Evening primrose: Oenothera biennis L, in: A.S. Awaad, V.K.
Singh, J.N. Govil (Eds.), Recent Progress in Medicinal Plants, vol. 28, Studium
Press LLC, Houston, 2010, pp. 143–158.
[12] J. de la Cruz, Antioxidant potential of evening primrose oil administration in
hyperlipemic rabbits, Life Sci. 65 (1999) 543–555, 65.
[13] P.R. Redden, Y.S. Huang, X. Lin, D.F. Horrobin, Separation and quantification
of the triacylglycerols in evening primrose and borage oils by reversed-
phase high-performance liquid chromatography, J. Chromatogr. A 694 (1995)
381–389.
[14] M. Karonen, J. Parker, A. Agrawal, J.P. Salminen, First evidence of hexameric
and heptameric ellagitannins in plants detected by liquid chromatog-
raphy/electrospray ionization mass spectrometry, Rapid Commun. Mass
Spectrom. 24 (2010) 3151–3156.
[15] M. Wettasinghe, F. Shahidi, R. Amarowicz, Identification and quantification
of low molecular weight phenolic antioxidants in seeds of evening primrose
(Oenothera biennis L.), J. Agric. Food Chem. 50 (2002) 1267–1271.
[16] A. Mari, D. Eletto, C. Pizza, P. Montoro, S. Piacente, Integrated mass spectrom-
etry approach to profile proanthocyanidins occurring in food supplements:
analysis of Potentilla erecta L. rhizomes, Food Chem. 141 (2013) 4171–4178.
[17] A. Bazylko, M. Tomczyk, A. Flazinska, A. Legas, Chemical fingerprint of Poten-
tilla species by using HPTLC method, JPC – J. Planar. Chromat. 24 (2011)
441–444.
[18] R. Domitrovic, The molecular basis for the pharmacological activity of antho-
cyans, Curr. Med. Chem. 29 (2011) 454–469.
[19] P.J. Curtis, P.A. Kroon, W.J. Hollands, R. Walls, G. Jenkins, C.D. Kay, A. Cassidy,
Cardiovascular disease risk biomarkers and liver and kidney function are not
altered in postmenopausal women after ingesting an elderberry extract rich
in anthocyanins for 12 weeks, J. Nutr. 139 (2009) 2266–2271.
206 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
[20] A. Stoj, A. Malik, Z. Targonski, Comparative analysis of anthocyanin composi-
tion of juices obtained from selected species of berry fruits, Pol. J. Food Nutr.
Sci. 15 (2006) 401–407.
[21] A. Stoj, Z. Targonski, Use of content analysis of selected organic acids for the
detection of berry juice adulterations, Pol. J. Food Nutr. Sci. 15 (2006) 41–47.
[22] S. Hakkinen, S. Auriola, High-performance liquid chromatography with elec-
trospray ionization mass spectrometry and diode array ultraviolet detection
in the identification of flavonol aglycons and glycosides in berries, J. Chro-
matogr. A 829 (1998) 91–100.
[23] W. Mullen, M.E.J. Lean, A. Crozier, Rapid characterization of anthocyanins in
red raspberry fruit by high-performance liquid chromatography coupled to
single quadrupole mass spectrometry, J. Chromatogr. A 966 (2002) 63–70.
[24] Q. Tian, M.M. Giusti, G.D. Stoner, S.J. Schwartz, Screening for anthocyanins
using high-performance liquid chromatography coupled to electrospray ion-
ization tandem mass spectrometry with precursor-ion analysis, product-ion
analysis, common-neutral-loss analysis, and selected reaction monitoring, J.
Chromatogr. A 1091 (2005) 72–82.
[25] D.C. Grant, R.J. Helleur, Rapid screening of anthocyanins in berry samples
by surfactant-mediated matrix-assisted laser desorption/ionization time-
of-flight mass spectrometry, Rapid Commun. Mass Spectrom. 22 (2008)
156–164.
[26] G. McDougall, I. Martinussen, D. Stewart, Towards fruitful metabolomics:
High throughput analyses of polyphenol composition in berries using direct
infusion mass spectrometry, J. Chromatogr. B: Analyt. Technol. Biomed. Life
Sci. 871 (2008) 362–369.
[27] I. Gulcin, F. Topal, R. Cakmakci, M. Bilsel, A.C. Goren, U. Erdogan, Pomologi-
cal features, nutritional quality, polyphenol content analysis, and antioxidant
properties of domesticated and 3 wild ecotype forms of raspberries (Rubus
idaeus L.), J. Food Sci. 76 (2011) C585–C593.
[28] C.M. Bradish, P. Perkins-Veazie, G.E. Fernandez, G. Xie, W. Jia, Comparison
of flavonoid composition of red raspberries (Rubus idaeus L.) grown in the
southern United States, J. Agric. Food Chem. 60 (2012) 5779–5786.
[29] R.C. Pimpao, T. Dew, P.B. Oliveira, G. Williamson, R.B. Ferreira, C.N. Santos,
Analysis of phenolic compounds in Portuguese wild and commercial berries
after multienzyme hydrolysis, J. Agric. Food Chem. 61 (2013) 4053–4062.
[30] F. Chen, Y. Sun, G. Zhao, X. Liao, X. Hu, J. Wu, Z. Wang, Optimiza-
tion of ultrasound-assisted extraction of anthocyanins in red raspberries
and identification of anthocyanins in extract using high-performance liq-
uid chromatography–mass spectrometry, Ultrason Sonochem. 14 (2007)
767–778.
[31] Y. Sun, X. Liao, Z. Wang, X. Hu, F. Chen, Optimization of microwave-assisted
extraction of anthocyanins in red raspberries and identification of antho-
cyanin of extracts using high-performance liquid chromatography–mass
spectrometry, Eur. Food Res. Technol. 225 (2007) 511–523.
[32] U. Ravid, M. Elkabetz, C. Zamir, K. Cohen, O. Larkov, R. Aly, Authenticity
assessment of natural fruit flavour compounds in foods and beverages by
auto-HS-SPME stereoselective GC–MS, Flavour Frag. J. 25 (2010) 20–27.
[33] W. Mullen, T. Yokota, M.E.J. Lean, A. Crozier, Analysis of ellagitannins and
conjugates of ellagic acid and quercetin in raspberry fruits by LC–MSn , Phy-
tochemistry 64 (2003) 617–624.
[34] M. Terauchi, N. Horiguchi, A. Kajiyama, M. Akiyoshi, Y. Owa, K. Kato, T. Kubota,
Effects of grape seed proanthocyanidin extract on menopausal symptoms,
body composition, and cardiovascular parameters in middle-aged women: a
randomized, double-blind, Menopause 21 (2014) 990–996.
[35] C. Cavaliere, P. Foglia, R. Gubbiotti, P. Sacchetti, R. Samperi, A. Lagana, Rapid-
resolution liquid chromatography/mass spectrometry for determination and
quantitation of polyphenols in grape berries, Rapid Commun. Mass Spectrom.
22 (2008) 3089–3099.
[36] Y. Xu, J.E. Simon, M.G. Ferruzzi, L. Ho, G.M. Pasinetti, Q. Wu, Quantification
of anthocyanidins in the grapes and grape juice products with acid assisted
hydrolysis using LC/MS, J. Funct. Foods 4 (2012) 710–717.
[37] R. Flamini, Recent applications of mass spectrometry in the study of grape
and wine polyphenols, ISRN Spectrosc 2013 (2013), Article ID 813563.
[38] C.W.P. Grandizoli, F.R. Campos, F. Simonelli, A. Barison, Grape juice quality
control by means of 1 H NMR spectroscopy and chemometric analyses, Quim.
Nova 37 (2014) 1227–1232.
[39] M.C. Gonzalez-Mas, L.M. Garcia-Riano, C. Alfaro, J.L. Rambla, A. Padilla, A.
Gutierrez, Headspace-based techniques to identify the principal volatile com-
pounds in red grape cultivars, Int. J. Food Sci. Nutr. 44 (2009) 510–518.
[40] R. Tundis, M.R. Loizzo, F. Menichini, G.A. Statti, F. Menichini, Biological and
pharmacological activities of iridoids: recent developments, Mini-Rev. Med.
Chem. 8 (2008) 399–420.
[41] P. Mirabi, M. Dolatian, F. Mojab, H.A. Majd, Effects of valerian on the severity
and systemic manifestations of dysmenorrhea, Int. J. Gynaecol. Obstet. 115
(2011) 285–288.
[42] Z.Z. Han, X.P. Zu, J.X. Wang, H.L. Li, B.Y. Chen, Q.X. Liu, X.Q. Hu, Z.H. Yan,
W.D. Zhang, Neomerane-type sesquiterpenoids from Valeriana officinalis var.
latifolia, Tetrahedron 70 (2014) 962–966.
[43] P.C. Wang, J.M. Hu, X.H. Ran, Z.Q. Cheng, H.Z. Jiang, Y.Q. Liu, J. Zhou, Y.X. Zhao,
Iridoids and sesquiterpenoids from the roots of Valeriana officinalis, J. Nat.
Prod. 72 (2009) 1682–1685.
[44] M.M. Shabana, N.M. El Fiki, I.A. Shehata, M. Shoukry, Quality control of a
sedative herbal drug, Bull. Fac. Pharm. (Cairo Univ.) 40 (2002) 215–221.
[45] R. Baby, M. Cabezas, E. Castro, R. Filip, N.E. Walsoe de Reca, Quality control of
medicinal plants with an electronic nose, Sens. Actuators B: Chem. 106 (2005)
24–28.
[46] M. Politi, M. Zloh, M.E. Pintado, P.M.L. Castro, M. Heinrich, J.M. Prieto, Direct
metabolic fingerprinting of commercial herbal tinctures by nuclear magnetic
resonance spectroscopy and mass spectrometry, Phytochem. Anal. 20 (2009)
328–334.
[47] B. Meier, D. Spriano, Modern HPTLC – a perfect tool for quality control of
herbals and their preparations, J. AOAC Int. 93 (2010) 1399–1409.
[48] J.R. Lucio-Gutiérrez, J. Coello, S. Maspoch, Enhanced chromatographic finger-
printing of herb materials by multi-wavelength selection and chemometrics,
Anal. Chim. Acta 710 (2012) 40–49.
[49] S.A. Schoenbichler, L.K.H. Bittner, J.D. Pallua, M. Popp, G. Abel, G.K. Bonn, C.W.
Huck, Simultaneous quantification of verbenalin and verbascoside in Verbena
officinalis by ATR-IR and NIR spectroscopy, J. Pharm. Biomed. Anal. 84 (2013)
97–102.
[50] A. Bilia, M. Giomi, M. Innocenti, S. Gallori, F. Vincieri, HPLC-DAD-ESI-MS analy-
sis of the constituents of aqueous preparations of verbena and lemon verbena
and evaluation of the antioxidant activity, J. Pharm. Biomed. Anal. 46 (2008)
463–470.
[51] Z. Liu, Z. Xu, H. Zhou, G. Cao, X.D. Cong, Y. Zhang, B.C. Cai, Simultaneous
determination of four bioactive compounds in Verbena officinalis L. by using
high-performance liquid chromatography, Phcog. Mag. 8 (2012) 162.
[52] J. Ruzicka, B. Lukas, L. Merza, I. Goehler, G. Abel, M. Popp, J. Novak, Identifica-
tion of Verbena officinalis based on ITS sequence analysis and RAPD-derived
molecular markers, Planta Med. 75 (2009) 1271–1276.
[53] A. Mari, P. Montoro, G. D’Urso, M. Macchia, C. Pizza, S. Piacente, Metabolic
profiling of Vitex agnus castus leaves, fruits and sprouts: Analysis by
LC/ESI/(QqQ)MS and (HR) LC/ESI/(Orbitrap)/MSn , J. Pharm. Biomed. Anal. 102
(2015) 215–221.
[54] A. Kuruüzüm-Uz, K. Ströch, L.O. Demirezer, A. Zeeck, Glucosides from Vitex
agnus-castus, Phytochemistry 63 (2003) 959–964.
[55] A. Mari, P. Montoro, C. Pizza, S. Piacente, Liquid chromatography tandem
mass spectrometry determination of chemical markers and principal com-
ponent analysis of Vitex agnus-castus L. fruits (Verbenaceae) and derived food
supplements, J. Pharm. Biomed. Anal. 70 (2012) 224–230.
[56] E. Hoberg, B. Meier, O. Sticher, Quantitative high performance liquid chro-
matographic analysis of diterpenoids in Agni-casti fructus, Planta Med. 66
(2000) 352–355.
[57] A.A. Hamid, L.A. Usman, S.A. Adebayo, M.F. Zubair, S.E. Elaigwu, Chemical
constituents of leaf essential oil of north-central Nigerian grown Vitex agnus-
castus L, Adv. Environ. Res. 4 (2010) 250–253.
[58] C. Proestos, D. Sereli, M. Komaitis, Determination of phenolic compounds in
aromatic plants by RP-HPLC and GC–MS, Food Chem. 95 (2006) 44–52.
[59] F.R. Marin, J.A. Perez-Alvarez, C. Soler-Rivas, Isoflavones as functional food
components, Stud. Nat. Prod. Chem. 32 (2005) 1177–1207.
[60] C. Grosso, J. Vinholes, L.R. Silva, P. Guedes de Pinho, R.F. Goncalves, P. Valentao,
A.K. Jager, P.B. Andrade, Chemical composition and biological screening of
Capsella bursa-pastoris, Rev. Bras. Farmacogn. 21 (2011) 635–644.
[61] A. Karioti, E. Giocaliere, C. Guccione, G. Pieraccini, E. Gallo, A. Vannacci, A.R.
Bilia, Combined HPLC-DAD-MS, HPLC-MSn and NMR spectroscopy for qual-
ity control of plant extracts: the case of a commercial blend sold as dietary
supplement, J. Pharm. Biomed. Anal. 88 (2014) 7–15.
[62] X. Zhou, L. Tang, Y. Xu, G. Zhou, Z. Wang, Toward a better understanding of
medicinal uses of Carthamus tinctorius L. in traditional Chinese medicine: a
phytochemical and pharmacological review, J. Ethnopharmacol. 151 (2014)
27–43.
[63] H.T. Liu, Y.F. Wang, O. Olaleye, Y. Zhu, X.M. Gao, L.Y. Kang, T. Zhao, Char-
acterization of in vivo antioxidant constituents and dual-standard quality
assessment of Danhong injection, Biomed. Chromatogr. 27 (2013) 655–663.
[64] X.X. Yang, X.X. Zhang, R.M. Chang, Y.W. Wang, X.N. Li, Simultaneous quantifi-
cation of five major active components in capsules of the traditional Chinese
medicine ‘Shu-Jin-Zhi-Tong’ by high performance liquid chromatography, J.
Pharm. Anal. 1 (2011) 284–290.
[65] Y. Chen, Y. Li, X. Chen, L. Wang, C. Sun, W. Yan, X. Liu, Development and vali-
dation of a HPLC method for the determination of five bioactive compounds
in the Xuebijing Injection, Anal. Lett. 43 (2010) 2456–2464.
[66] Y. Li, D. Piao, H. Zhang, T. Kim, S.H. Lee, H.W. Chang, M.H. Woo, J.K. Son,
Quality evaluation of Carthami Flos by HPLC-UV, Arch. Pharm. Res. (2014),
http://dx.doi.org/10.1007/s12272-014-0402-9 (Epub ahead of Print).
[67] L. Fan, H.Y. Zhao, M. Xu, H. Lei, J. Guo, B.R. Han, D. Wang, A. Guo, Qualitative
evaluation and quantitative determination of 10 major active components in
Carthamus tinctorius L. by high-performance liquid chromatography coupled
with diode array detector, J. Chromatogr. A 1216 (2009) 2063–2070.
[68] M.B. Zhao, X.L. Deng, Y.L. Wang, M. Lu, P.F. Tu, Establishment of chromato-
graphic fingerprint and quality assessment of Carthamus tinctorius L. by high
performance liquid chromatography, Acta Pharm. Sin. 39 (2004) 212–216.
[69] D. Shen, Q. Wu, W.J. Sciarappa, J.E. Simon, Chromatographic fingerprints and
quantitative analysis of isoflavones in Tofu-type soybeans, Food Chem. 130
(2012) 1003–1009.
[70] M.C. Nemitz, R.C. Moraes, L.S. Koester, V.L. Bassani, G.L. von Poser, H.F.
Teixeira, Bioactive soy isoflavones: extraction and purification procedures,
potential dermal use and nanotechnology-based delivery systems, Phy-
tochem. Rev. (2014), http://dx.doi.org/10.1007/s11101-014-9382-0.
[71] M.A. Rostagno, A. Villares, E. Guillamon, A. Garcia-Lafuente, J.A. Martinez,
Sample preparation for the analysis of isoflavones from soybeans and soy
foods, J. Chromatogr. A 1216 (2009) 2–29.
[72] D.L. Luthria, S.S. Natarajan, Influence of sample preparation on the assay of
isoflavones, Planta Med. 75 (2009) 704–710.
 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
[73] M.C. Nemitz, F.K.J. Yatsu, J. Bidone, L.S. Koester, V.L. Bassani, C.V. Garcia,
A.S.L. Mendez, von Poser, L. Gilsane, H.F. Teixeira, A versatile, stability-
indicating and high-throughput ultra-fast liquid chromatography method for
the determination of isoflavone aglycones in soybeans, topical formulations,
and permeation assays, Talanta 134 (2015) 183–193.
[74] Q. Wu, M. Wang, W.J. Sciarappa, J.E. Simon, LC/UV/ESI-MS analysis of
isoflavones in Edamame and Tofu soybeans, J. Agric. Food Chem. 52 (2004)
2763–2769.
[75] L. Gu, W. Gu, Characterization of soy isoflavones and screening for
novel malonyl glycosides using high-performance liquid chromatography–
electrospray ionization–mass spectrometry, Phytochem. Anal. 12 (2001)
377–382.
[76] N. Fang, S. Yu, T.M. Badger, Comprehensive phytochemical profile of soy pro-
tein isolate, J. Agric. Food Chem. 52 (2004) 4012–4020.
[77] Caligiani, G. Palla, A. Maietti, M. Cirlini, V. Brandolini, 1 H NMR fingerprinting
of soybean extracts, with emphasis on identification and quantification of
isoflavones, Nutrients 2 (2010) 280–289.
[78] J.A. Hoeck, W.R. Fehr, P.A. Murphy, G.A. Welke, Influence of genotype and
environment on isoflavone contents of soybean, Crop Sci. 40 (2000) 48–51.
[79] M.C. Garcia, M. Torre, M.L. Marina, Characterization of commercial soybean
products by conventional and perfusion reversed-phase high-performance
liquid chromatography and multivariate analysis, J. Chromatogr. A 881 (2000)
47–57.
[80] L.R. Silva, M.J. Pereira, J. Azevedo, R.F. Goncalves, P. Valentao, P. Guedes de
Pinho, P.B. Andrade, Glycine max (L.) Merr., Vigna radiate L. and Medicago
sativa L. sprouts: a natural source of bioactive compounds, Food Res. Int. 50
(2013) 167–175.
[81] Rodrigues, I. Almeida, B. Sarmento, M.H. Amaral, M. Beatriz, Study of the
isoflavone content of different extracts of Medicago spp. as potential active
ingredient Oliveira, Ind. Crop Prod. 57 (2014) 110–115.
[82] S.B. Kundan, S. Anupam, Phytochemical and pharmacological potential of
Medicago sativa: a review, Pharmaceut. Biol. 49 (2011) 211–220.
[83] P.A. Murphy, T. Song, G. Buseman, K. Barua, G.R. Beecher, De. Trainer, J. Holden,
Isoflavones in retail and institutional soy foods, J. Agric. Food Chem. 47 (1999)
2697–2704.
[84] S. Prati, V. Baravelli, D. Fabbri, C. Schwarzinger, V. Brandolini, A. Maietti, P.
Tedeschi, S. Benvenuti, M. Macchia, I. Marotti, A. Bonetti, P. Catizone, G. Ginelli,
Composition and content of seed flavonoids in forage and grain legume crops,
J. Sep. Sci. 30 (2007) 491–501.
[85] C.L. Aguiar, A.S. Baptista, S.M. Alencar, R. Haddad, M.N. Eberlin, Analysis
of isoflavonoids from leguminous plant extracts by RPHPLC/DAD and elec-
trospray ionization mass spectrometry, Int. J. Food Sci. Nutr. 58 (2007)
116–124.
[86] S. Golawska, I. Lukasik, I. Kapusta, B. Janda, Analysis of flavonoids content in
alfalfa, Ecol. Chem. Eng. 17 (2010) 261–267.
[87] B.R. Baggett, J.D. Cooper, E.T. Hogan, J. Carper, N.L. Paiva, J.T. Smith, Profiling
isoflavonoids found in legume root extracts using capillary electrophoresis,
Electrophoresis 23 (2002) 1642–1651.
[88] A. Abo Markeb, N. Abo El-Maali, Simultaneous quantitation of 5- and 7-
hydroxyflavone antioxidants and their binding constants with BSA using dual
chiral capillary electrophoresis (dCCE) and HPLC with fluorescent detection,
Talanta 119 (2014) 417–424.
[89] H. Colgecen, U.K. Caliskan, M. Kartal, H.N. Buyukkartal, Comprehensive eval-
uation of phytoestrogen accumulation in plants and in vitro cultures of
Medicago sativa L. ‘Elci’ and natural tetraploid Trifolium pratense L, Turk. J.
Biol. 38 (2014) 619–627.
[90] D.V. Huhman, L.W. Sumner, Metabolic profiling of saponins in Medicago sativa
and Medicago truncatula using HPLC coupled to an electrospray ion-trap mass
spectrometer, Phytochemistry 59 (2002) 347–360.
[91] C.N. He, Y. Peng, Y.C. Zhang, L.J. Xu, J. Gu, P.G. Xiao, Phytochemical and bio-
logical studies of paeoniaceae, Chem. Biodivers. 7 (2010) 805–838.
[92] W. Zhang, S.M. Dai, Mechanisms involved in the therapeutic effects of Paeo-
nia lactiflora Pallas in rheumatoid arthritis, Int. Immunopharmacol. 14 (2012)
27–31.
[93] S. Arentz, J.A. Abbott, C.A. Smith, A. Bensoussan, Herbal medicine for
the management of polycystic ovary syndrome (PCOS) and associated
oligo/amenorrhoea and hyperandrogenism; a review of the laboratory evi-
dence for effects with corroborative clinical findings, BMC Complement.
Altern. Med. 18 (2014) 511.
[94] S.L. Li, J.Z. Song, F.F.K. Choi, C.F. Qiao, Y. Zhou, Q.B. Han, H.X. Xu, Chem-
ical profiling of Radix Paeoniae evaluated by ultra-performance liquid
chromatography/photo-diode-array/quadrupole time-of-flight mass spec-
trometry, J. Pharm. Biomed. Anal. 49 (2009) 253–266.
[95] N.C. Luo, W. Ding, J. Wu, D.W. Qian, Z.H. Li, Y.F. Qian, J.M. Guo, J.A. Duan, UPLC-
Q-TOF/MS coupled with multivariate statistical as a powerful technique for
rapidly exploring potential chemical markers to differentiate between Radix
Alba and Radix, Nat. Prod. Commun. 8 (2013) 487–491.
[96] Q. Wang, Z. Liang, Y. Peng, J.L. Hou, S.L. Wei, Z.Z. Zhao, W.Q. Wang, Whole
transverse section and specific-tissue analysis of secondary metabolites in
seven different grades of root of Paeonia lactiflora using laser microdissection
and liquid chromatography-quadrupole/time of flight-mass spectrometry, J.
Pharm. Biomed. Anal. 103 (2015) 7–16.
[97] F. Ahmad, N. Tabassum, S. Rasool, Medicinal uses and phytoconstituents of
Paeonia officinalis, IRJP 3 (2012) 85–87.
[98] L. Yang, S.J. Xu, Q.R. Feng, H. Liu, R.T. Tian, P.S. Xie, A simple thin-layer chro-
matographic fingerprint method for distinguishing between Radix Paeonia
207
rubra and Radix Paeoniae alba, J. Liq. Chromatogr. Relat. Technol. 32 (2009)
2893–2905.
[99] S. Xu, L. Yang, R. Tian, Z. Wang, Z. Liu, P. Xie, Q. Feng, Species differentiation and
quality assessment of Radix (Chi-shao) by means of high-performance liquid
chromatographic fingerprint, J. Chromatogr. A 1216 (2009) 2163–2168.
[100] M. Johnson, V. Irudayaraj, Preliminary phytochemical and anti-bacterial stud-
ies on Passiflora edulis, Ethnobot. Leaflets 12 (2008) 425–432.
[101] S.T. Talcott, S.S. Percival, J. Pittet-Moore, C. Celoria, Phytochemical compo-
sition and antioxidant stability of fortified yellow passion fruit (Passiflora
edulis), J. Agric. Food Chem. 51 (2003) 935–941.
[102] B. Avula, Y.H. Wang, C.S. Rumalla, T.J. Smillie, I.A. Khan, Simultaneous deter-
mination of alkaloids and flavonoids from aerial parts of Passiflora species
and dietary supplements using UPLC-UV-MS and HPTLC, Nat. Prod. Comm. 7
(2012) 1177–1180.
[103] C.D. Birk, G. Provensi, G. Gosmann, F.H. Reginatto, E.P. Schenkel, TLC finger-
print of flavonoids and saponins from Passiflora species, J. Liq. Chromatogr.
Relat. Technol. 28 (2005) 2285–2291.
[104] C.A. Pereira, J.H. Yariwake, F.M. Lancas, J.N. Wauters, M. Tits, L.A. Angenot,
HPTLC densitometric determination of flavonoids from Passiflora alata, P.,
edulis, P. incarnata and P. caerulea and comparison with HPLC method, Phy-
tochem. Anal. 15 (2004) 241–248.
[105] W. Peng, R. Qin, X. Li, H. Zhou, Botany, phytochemistry, pharmacology, and
potential application of Polygonum cuspidatum Sieb.et Zucc.: a review, J.
Ethnopharmacol. 148 (2013) 729–745.
[106] R.Z. Zhao, S. Liu, L.L. Zhou, Rapid quantitative HPTLC analysis, on one plate, of
emodin, resveratrol, and polydatin in the Chinese herb Polygonum cuspidatum,
Chromatographia 61 (2005) 311–314.
[107] M.C. Zhang, X.B. Peng, Y.M. Wang, J.M. Shao, Determination of resveratrol
and the other 3 constituents of Polygonum cuspidatum Sieb, et Zucc by HPLC,
Medicinal Plant 1 (2010) 54–56.
[108] P. Ma, K. Luo, Y. Peng, W. Wang, Z. Zhang, Y. Liu, X. Zhao, L. Xu, P. Xiao, Quality
control of Polygonum cuspidatum by UPLC-PDA and related statistical analysis,
J. Liq. Chromatogr. Relat. Technol. 36 (2013) 2844–2854.
[109] A.V. Lekar, S.N. Borisenko, E.V. Vetrova, S.N. Sushkova, N.I. Borisenko, Extrac-
tion of quercetin from Polygonum hydropiper L. by subcritical water, Am. J.
Agric. Biol. Sci. 9 (2014) 1–5.
[110] M.A. Hawryl, M. Waksmundzka-Hajnos, Micro 2D-TLC of selected plant
extracts in screening of their composition and antioxidative properties, Chro-
matographia 76 (2013) 1347–1352.
[111] X. Yang, B.C. Wang, X. Zhang, S.P. Yang, W. Li, Q. Tang, G.K. Singh, Simultaneous
determination of nine flavonoids in Polygonum hydropiper L. samples using
nanomagnetic powder three-phase hollow fibre-based liquid-phase microex-
traction combined with ultrahigh performance liquid chromatography–mass
spectrometry, J. Pharm. Biomed. Anal. 54 (2011) 311–316.
[112] S. Granica, J.P. Piwowarski, M. Popławska, M. Jakubowska, J. Borzym, A.K. Kiss,
Novel insight into qualitative standardization of Polygoni avicularis herba (Ph.
Eur.), J. Pharm. Biomed. 72 (2013) 216–222.
[113] D.S. Fan, X. Zhou, C. Zhao, H.G. Chen, Y. Zhao, X.J. Gong, Anti-inflammatory,
antiviral and quantitative study of quercetin-3-O-beta-d-glucuronide in Poly-
gonum perfoliatum L, Fitoterapia 82 (2011) 805–810.
[114] N.A. Yunuskhodzhaeva, K.A. Eshbakova, V.N. Abdullabekova, Flavonoid com-
position of the herb Polygonum aviculare, Chem. Nat. Compd. 46 (2010)
803–804.
[115] N. Deshkar, S. Tilloo, V. Pande, A comprehensive review of Rubia cordifolia
Linn, Pharmacogn. Rev. 2 (2008) 124–134.
[116] N.P. Mishchenko, S.A. Fedoreev, V.M. Bryukhanov, Y.F. Zverev, V.V. Lampa-
tov, O.V. Azarova, Y.N. Shkryl’, G.K. Chernoded, Chemical composition and
pharmacological activity of anthraquinones from Rubia cordifolia cell culture,
Pharm. Chem. J. 4 (2007) 605–609.
[117] A. Laiqa, A. Javed, M. Mohd, A. Altaf, A validated quantitative determination
of alizarin in Rubia cordifolia Linn. by isocratic RP-HPLC, Int. J. Pharm. Biol. Sci.
5 (2014) 1–6.
[118] H.K. Soni, B.R. Shah, S.B. Bhatt, N.R. Sheth, Standardization of single herb cap-
sules of Manjistha, Kokam and Punarnava with assessment of their nutritional
value, J. Pharm. Res. 3 (2010) 1899–1902.
[119] T. Sabudak, N. Guler, Trifolium L. – a review on its phytochemical and phar-
macological profile, Phytother. Res. 23 (2009) 439–446.
[120] E. De Rijge, A. Zarfa Gomez, FTh. Arise, A. Brinkman, C. Gooljer, Determination
of isoflavone glucoside malonates in Trifolium pretense L. (red clover) extracts,
J. Chromatogr. A 932 (2001) 55–64.
[121] L.V. Gu, W.Y. Gu, Characterisation of soy isoflavones and screen-
ing for novel malonyl glycosides using high-performance liquid
chromatography–electrospray ionisation–mass spectrometry, Phytochem.
Anal. 12 (2001) 377–382.
[122] L. Krenn, I. Unterrieder, R. Ruprechter, Quantification of isoflavones in red
clover by high-performance liquid chromatography, J. Chromatogr. B 777
(2002) 123–128.
[123] C.E. Piersen, N.L. Booth, Y. Sun, W. Liang, J.E. Burdette, R.B. van Breemen, S.E.
Geller, C. Gu, S. Banuvar, L.P. Shulman, J.L. Bolton, N.R. Farnsworth, Chemical
and biological characterization and clinical evaluation of botanical dietary
supplements: a phase I red clover extract as a model, Curr. Med. Chem. 11
(2004) 1361–1374.
[124] Y.W. Wu, S.Q. Sun, Q. Zhou, J.X. Tao, I. Noda, Volatility-dependent 2D
IR correlation analysis of traditional Chinese medicine Red Flower Oil
preparation from different manufacturers, J. Mol. Struct. 882 (2008)
107–115.
208 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
[125] Q. Chen, P. Li, B. Li, X. Li, J. Zhu, F. Chen, Simultaneous determination of
formononetin, biochanin A, daidzen and genistein in Trifolium pratense (red
clover) by HPLC, LC-GC Eur. 23 (2010) 406–411.
[126] J. Kasper, M.F. Melzig, HPTLC method for the quantification of isoflavones in
nutritional supplements of red clover (Trifolium pratense L.), J. Planar Chro-
matogr. Mod. TLC 24 (2011) 373–375.
[127] S. Waldmann, M. Almukainzi, N.A. Bou-Chacra, G.L. Amidon, B.J. Lee, J. Feng,
I. Kanfer, J.Z. Zuo, H. Wei, M.B. Bolger, R. Lobenberg, Provisional biopharma-
ceutical classification of some common herbs used in western medicine, Mol.
Pharmaceut. 9 (2012) 815–822.
[128] Y.L. Wang, Y.Z. Liang, B.M. Chen, High-performance liquid chromatography
with atmospheric pressure chemical ionization and electrospray ionization
mass spectrometry for analysis of Angelica sinensis, Phytochem. Anal. 18
(2007) 265–274.
[129] X. Li, X. Wu, L. Huang, Correlation between antioxidant activities and phe-
nolic contents of Radix Angelicae sinensis (Danggui), Molecules 14 (2009)
5349–5361.
[130] Z.Y. Li, Z.Z. Zhang, G.H. Du, X.M. Qin, Comparative analysis of Danggui and
European Danggui using nuclear magnetic resonance-based metabolic fin-
gerprinting, J. Pharm. Biomed. Anal. 103 (2015) 44–51.
[131] L.Z. Yi, Y.Z. Liang, H. Wu, D.L. Yuan, The analysis of Radix Angelicae sinensis
(Danggui), J. Chromatogr. A 1216 (2009) 1991–2001.
[132] Y.J. Bai, M. Kong, J.D. Xu, X.L. Zhang, S.S. Zhou, X.N. Wang, L.F. Liu, S.L. Li,
Effect of different drying methods on the quality of Angelicae sinensis Radix
evaluated through simultaneously determining four types of major bioactive
components by high performance liquid chromatography photodiode array
detector and ultra-high performance liquid chromatography quadrupole
time-of-flight mass spectrometry, J. Pharm. Biomed. Anal. 94 (2014) 77–83.
[133] X. Wang, X. Shi, F. Li, J. Liu, C. Cheng, Application of analytical and prepar-
ative high-speed counter-current chromatography for the separation of
Z-ligustilide from a crude extract of Angelica sinensis, Phytochem. Anal. 19
(2008) 193–197.
[134] P. Li, S.P. Li, S.C. Lao, C.M. Fu, K.W. Kelvin, Y.T. Kan Wang, Optimization of pres-
surized liquid extraction for Z-ligustilide, Z-butylidenephthalide and ferulic
acid in Angelica sinensis, J. Pharm. Biomed. Anal. 40 (2006) 1073–1079.
[135] Y. Zhao, J. Sun, L.L. Yu, P. Chen, Chromatographic and mass spectrometric
fingerprinting analyses of Angelica sinensis (Oliv.) Diels-derived dietary sup-
plements, Anal. Bioanal. Chem. 405 (2013) 4477–4485.
[136] Filipiak-Szok, M. Kurzawa, E. Szlyk, Evaluation of antioxidants in Dong quai
(Angelica sinensis) and its dietary supplements, Chem. Pap. 68 (2014) 493–503.
[137] J. Suresh, K. Mruthunjaya, N. Paramakrishnan, M.N. Naganandhini, Determi-
nation of Artemisinin in Artemisia abrotanum and Artemisia pallens by LC/MS
method, Int. J. Clin. Pharmacol. Res. 3 (2011) 49–52.
[138] J.A. Pino, R. Marbot, M.P. Marti, Leaf oil of Artemisia abrotanum L. grown in
Cuba, J. Essent. Oil Res. 23 (2011) 119–120.
[139] J.H. Kim, S.H. Jung, Y.I. Yang, J.H. Ahn, J.-G. Cho, K.T. Lee, N.I. Baek, J.H.
Choi, Artemisia leaf extract induces apoptosis in human endometriotic cells
through regulation of the p38 and NF-kB pathways, J. Ethnopharmacol. 145
(2013) 767–775.
[140] Y. Zhao, C.A. Geng, Y.B. Ma, X.Y. Huang, H. Chen, T.W. Cao, K. He, H. Wang,
X.M. Zhang, J.J. Chen, UFLC/MS-IT-TOF guided isolation of anti-HBV active
chlorogenic acid analogues from Artemisia capillaris as a traditional Chinese
herb for the treatment of hepatitis, J. Ethnopharmacol. 156 (2014) 147–154.
[141] B. Avula, Y.H. Wang, T.J. Smillie, W. Mabusela, L. Vincent, F. Weitz, I.A. Khan,
Comparison of LC–UV, LC-ELSD and LC–MS methods for the determination of
sesquiterpenoids in various species of Artemisia, Chromatographia 70 (2009)
797–803.
[142] Yang, D.Y. Lee, M. Jeon, Y. Suh, S.H. Sung, Determination of five active
compounds in Artemisia princeps and A. capillaris based on UPLC-DAD and
discrimination of two species with multivariate analysis, Arch. Pharmacal.
Res. 37 (2014) 617–625.
[143] D. Lopes-Lutz, D.S. Alviano, C.S. Alviano, P.P. Kolodziejczyk, Screening of
chemical composition, antimicrobial and antioxidant activities of Artemisia
essential oils, Phytochemistry 69 (2008) 1732–1738.
[144] S. Gairola, M. Maithani, V. Gupta, P. Bansal, P.j. Ghaiye, Pharmacological poten-
tial and chemical constituents of Artemisia vulgaris, J. Pharm. Res. Clin. Pract.
1 (2011) 82–87.
[145] H.J. de Boer, C. Cotingting, Medicinal plants for women’s healthcare in south-
east Asia: a meta-analysis of their traditional use, chemical constituents, and
pharmacology, J. Ethnopharmacol. 151 (2014) 747–767.
[146] D. Melguizo-Melguizo, E. Diaz-de-Cerio, R. Quirantes-Piné, J. Švarc-Gajicı́,
A. Segura-Carretero, The potential of Artemisia vulgaris leaves as a
source of antioxidant phenolic compounds, J. Funct. Foods 10 (2014)
192–200.
[147] M. Wang, B. Avula, Y.H. Wang, J. Zhao, C. Avonto, J.F. Parcher, V. Raman, J.A.
Zweigenbaum, P.L. Wylie, I.A. Khan, An integrated approach utilising chemo-
metrics and GC/MS for classification of chamomile flowers, essential oils and
commercial products, Food Chem. 152 (2014) 391–398.
[148] C.M. Ma, L. Winsor, M. Daneshtalab, Quantification of spiroether isomers
and herniarin of different parts of Matricaria matricarioides and flowers of
Chamaemelum nobile, Phytochem. Anal. 18 (2006) 42–49.
[149] B. Avula, Y.H. Wang, M. Wang, C. Avonto, J.P. Zhao, T.J. Smilie, D. Rua, I.A.
Khan, Quantitative determination of phenolic compounds by UHPLC-UV-MS
and use of partial least-squares discriminant analysis to differentiate chemo-
types of chamomile/chrysanthemum flower heads, J. Pharm. Biomed. Anal.
88 (2014) 278–288.
[150] K. Dhanya, S. Syamkumar, S. Siju, B. Sasikumar, Sequence characterized ampli-
fied region markers: a reliable tool for adulterant detection in turmeric
powder, Food Res. Int. 44 (2011) 2889–2895.
[151] C. Jin, W. Kong, Y. Luo, J. Wang, H. Wang, Q. Li, X. Xiao, Development and
validation of UPLC method for quality control of Curcuma longa Linn.: fast
simultaneous quantitation of three curcuminoids, J. Pharm. Biomed. Anal. 53
(2010) 43–49.
[152] H.H. Tonnesen, A.L. Grislingaas, J. Karlsen, Studies on curcumin and cur-
cuminoids. XIX. Evaluation of thin-layer chromatography as a method for
quantitation of curcumin and curcumin, Z. Lebensm. Unters. Forsch. 193
(1991) 548–550.
[153] M. Paramasivam a, R. Poi, H. Banerjee, A. Bandyopadhyay, High-performance
thin layer chromatographic method for quantitative determination of cur-
cuminoids in Curcuma longa germplasm, Food Chem. 113 (2009) 640–644.
[154] K.K. Rout, S. Parida, S.K. Mishra, Standardization of the ayurvedic formu-
lation Haridra Khanda using high-performance thin-layer chromatography
densitometry, J. AOAC Int. 91 (2008) 1162–1168.
[155] K. Tanaka, Y. Kuba, T. Sasaki, F. Hiwatashi, K. Komatsu, Quantitation of cur-
cuminoids in curcuma rhizome by near-infrared spectroscopic analysis, J.
Agric. Food Chem. 56 (2008) 8787–8792.
[156] T. Nhujak, W. Saisuwan, M. Srisa-art, A. Petsom, Microemulsion electrokinetic
chromatography for separation and analysis of curcuminoids in turmeric
samples, J. Sep. Sci. 29 (2006) 666–676.
[157] M. Lechtenberg, B. Quandt, A. Nahrstedt, Quantitative determination of
curcuminoids in Curcuma rhizomes and rapid differentiation of Curcuma
domestica Val. and Curcuma xanthorrhiza Roxb. by capillary electrophoresis,
Phytochem. Anal. 15 (2004) 152–158.
[158] U.K.S. Sanagi, M. Marsin, R.M. Ahmad, Application of supercritical fluid extrac-
tion and chromatography to the analysis of turmeric, J. Chromatogr. Sci. 31
(1993) 20–25.
[159] H.J. Kim, Y.P. Jang, Direct analysis of curcumin in turmeric by DART-MS, Phy-
tochem. Anal. 20 (2009) 372–377.
[160] H. Jiang, A. Somogyi, N.E. Jacobsen, B.N. Timmermann, D.R. Gang, Analysis of
curcuminoids by positive and negative electrospray ionization and tandem
mass spectrometry, Rapid Commun. Mass Spectrom. 20 (2006) 1001–1012.
[161] K. Inoue, S. Hamasaki, Y. Yoshimura, M. Yamada, M. Nakamura, Y. Ito, H.
Nakazawa, Validation of LC/Electrospray-MS for determination of major cur-
cuminoids in foods, J. Liq. Chromatogr. Relat. Technol. 26 (2003) 53–62.
[162] C.G. Ahmet, C. Simay, C. Muhiddin, B. Gökhan, Rapid quantitation of curcumin
in turmeric via NMR and LC–tandem mass spectrometry, Food Chem. 113
(2009) 1239–1242.
[163] C. Mathon, M. Duret, M. Kohler, P. Edder, S. Bieri, P. Christen, Multi-targeted
screening of botanicals in food supplements by liquid chromatography with
tandem mass spectrometry, Food Chem. 138 (2013) 709–717.
[164] A. Karioti, E. Fani, F.F. Vincieri, A.R. Bilia, Analysis and stability of the con-
stituents of Curcuma longa and Harpagophytum procumbens tinctures by
HPLC-DAD and HPLC-ESI-MS, J. Pharm. Biomed. Anal. 55 (2011) 479–486.
[165] J. Cheng, K. Weijun, L. Yun, W. Jiabo, W. Haitao, L. Qingmiao, X. Xiaoh,
Development and validation of UPLC method for quality control of Curcuma
longa Linn.: fast simultaneous quantitation of three curcuminoids, J. Pharm.
Biomed. Anal. 53 (2010) 43–49.
[166] B. Avula, Y.H. Wang, I.A. Khan, Quantitative determination of curcuminoids
from the roots of Curcuma longa, Curcuma species and dietary supplements
using an UPLC-UV-MS method, J. Chromatograph. Separat. Techniq. 3 (2012)
120–125.
[167] A. Jitoe, T. Masuda, I.G.P. Tengah, D.N. Suprapta, I.W. Gara, N. Nakatani,
Antioxidant activity of tropical ginger extracts and analysis of the contained
curcuminoids, J. Agric. Food Chem. 40 (1992) 1337–1340.
[168] S. Ruslay, F. Abas, K. Shaari, Z. Zainal, H. Maulidiani, D.A. Sirat, N.H. Israf, H. Lajis
Nordin, Characterization of the components present in the active fractions of
health gingers (Curcuma xanthorrhiza and Zingiber zerumbet) by HPLC-DAD-
ESIMS, Food Chem. 104 (2007) 1183–1191.
[169] S. Paramapojn, W. Gritsanapan, Free radical scavenging activity determina-
tion and quantitative analysis of curcuminoids in Curcuma zedoaria rhizome
extracts by HPLC method, Curr. Sci. 97 (2009) 1069–1073.
[170] J. Zhang, S. Jinnai, R. Ikeda, M. Wada, S. Hayashida, K. Nakashima, A simple
HPLC-fluorescence method for quantitation of curcuminoids and its applica-
tion to turmeric products, Anal. Sci. 25 (2009) 385–388.
[171] N.Y. Qin, F.Q. Yang, Y.T. Wang, S.P. Li, Quantitative determination of eight com-
ponents in rhizome (Jianghuang) and tuberous root (Yujin) of Curcuma longa
using pressurized liquid extraction and gas chromatography–mass spectrom-
etry, J. Pharm. Biomed. Anal. 43 (2007) 486–492.
[172] J. Ibrahim bin, A. Abu Said, A. Nor Azah Mohd, A. Abdul Rashih, I. Halijah, Chem-
ical composition of the rhizome oils of four Curcuma species from Malaysia,
J. Essent. Oil Res. 11 (1999) 719–723.
[173] I. Jacobson, A.K. Joensson, B. Gerden, S. Haegg, Spontaneously reported
adverse reactions in association with complementary and alternative
medicine substances in Sweden, Pharmacoepidemiol. Drug Saf. 18 (2009)
1039–1047.
[174] A. Olalde Rangel, Synergistic phytoceutical compositions for treatment of cir-
culatory disorders, feminine endocrine disorders, and dermal disorders, U.S.
Pat. Appl. Publ. (2007).
[175] J. Barnes, L.A. Anderson, S. Gibbons, J.D. Phillipson, Echinacea species (Echi-
nacea angustifolia (DC.) Hell., Echinacea pallida (Nutt.) Nutt., Echinacea
purpurea (L.) Moench): a review of their chemistry, pharmacology and clinical
properties, J. Pharm. Pharmacol. 57 (2005) 929–954.
 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
[176] X.B. Luo, B. Chen, S.Z. Yao, J.G. Zeng, Simultaneous analysis of caffeic
acid derivatives and alkamides in roots and extracts of Echinacea purpurea
by high performance liquid chromatography–photodiode array detection-
electrospray mass spectrometry, J. Chromatogr. A 986 (2003) 73–81.
[177] W. Li, Y. Sun, R.B. van Breemen, J.F. Fitzloff, Evaluation of major caffeic acid
derivatives in Echinacea purpurea dietary supplements by HPLC with photodi-
ode array detection and electrospray mass spectrometry, J. Liq. Chromatogr.
Relat. Technol. 27 (2004) 2507–2529.
[178] N.B. Cech, M.S. Eleazer, L.T. Shoffner, M.R. Crosswhite, A.C. Davis, A.M.
Mortenson, High performance liquid chromatography/electrospray ioniza-
tion mass spectrometry for simultaneous analysis of alkamides and caffeic
acid derivatives from Echinacea purpurea extracts, J. Chromatogr. A 1103
(2006) 219–228.
[179] C. Bergeron, S. Gafner, Quantitative analysis of the polysaccharide and
glycoprotein fractions in Echinacea purpurea and Echinacea angustifolia by
HPLC-ELSD for quality control of raw material, Pharm. Biol. 45 (2007)
98–105.
[180] E. Mudge, D. Lopes-Lutz, P. Brown, A. Schieber, Analysis of alkylamides in
echinacea plant materials and dietary supplements by ultrafast liquid chro-
matography with diode array and mass spectrometric detection, J. Agric. Food
Chem. 59 (2011) 8086–8094.
[181] S. Agatonovic-Kustrin, C.M. Loescher, R. Singh, Quantification of phenyl-
propanoids in commercial echinacea products using TLC with video
densitometry as detection technique and ANN for data modelling, Phytochem.
Anal. 24 (2013) 303–308.
[182] M. Sandasi, I. Vermaak, W. Chen, A.M. Viljoen, Hyperspectral imaging and
chemometric modeling of Echinacea – a novel approach in the quality control
of herbal medicines, Molecules 19 (2014) 13104–13121.
[183] P. Zanoli, M. Zavatti, Pharmacognostic and pharmacological profile of Humu-
lus lupulus L, J. Ethnopharmacol. 116 (2008) 383–396.
[184] P.J. Magalhães, J.S. Vieira, L.M. Goncalves, J.G. Pacheco, L.F. Guido,
A.A. Barros, Isolation of phenolic compounds from hop extracts using
polyvinylpolypyrrolidone: characterization by high-performance liquid
chromatography–diode array detection–electrospray tandem mass spec-
trometry, J. Chromatogr. A 1217 (2010) 3258–3268.
[185] D. Nikoli, R.B. Van Breemen, Analytical methods for quantitation of prenylated
flavonoids from hops, Curr. Anal. Chem. 9 (2013) 71–85.
[186] L. Ceslova, M. Holcapek, M. Fidler, J. Drstickova, M. Lisa, Characterization of
prenylflavonoids and hop bitter acids in various classes of Czech beers and hop
extracts using high-performance liquid chromatography–mass spectrome-
try, J. Chromatogr. A 1216 (2009) 7249–7257.
[187] F.P. Prencipe, V. Brighenti, M. Rodolfi, A. Mongelli, C. dall’Asta, T. Ganino, R.
Bruni, F. Pellati, Development of a new high-performance liquid chromatogra-
phy method with diode array and electrospray ionization-mass spectrometry
detection for the metabolite fingerprinting of bioactive compounds in Humu-
lus lupulus L, J. Chromatogr. A 1349 (2014) 50–59.
[188] J. Olšovská, Z. Kameník, P. Čejka, M. Jurková, A. Mikyška, Ultra-high-
performance liquid chromatography profiling method for chemical screening
of proanthocyanidinsin Czech hops, Talanta 116 (2013) 919–926.
[189] M.A. Farag, E.A. Mahrous, T. Lubken, A. Porzel, L. Wessjohann, Classification of
commercial cultivars of Humulus lupulus L. (hop) by chemometric pixel anal-
ysis of two dimensional nuclear magnetic resonance spectra, Metabolomics
10 (2014) 21–32.
[190] Y. Jiang, J. Vaysse, V. Gilard, S. Balayssac, S. Dejean, M. Malet-Martino, B. David,
C. Fiorini, Y. Barbin, Quality assessment of commercial Magnoliae officinalis
cortex by 1 H-NMR-based metabolomics and HPLC methods, Phytochem. Anal.
23 (2012) 387–395.
[191] Wang, K. Yuan, W.W. Yu, J. Wang, Evaluation and discrimination of cor-
tex Magnolia officinalis produced in Zhejiang province (Wen-Hou-Po) by
UPLCDAD-TOF-MS fingerprint, Nat. Prod. Comm. 5 (2010) 1631–1638.
[192] M.S.A. Marzouk, F.A. Moharram, M.A. Mohamed, A.M. Gamal-Eldeend, E.A.
Aboutabl, Anticancer and antioxidant tannins from Pimenta dioica leaves, Z
Naturforsch C. 62 (2007) 526–536.
[193] K.P.P. Amma, M.P. Rani, I. Sasidharan, M.M. Sreekumar, Comparative chemical
composition and in vitro antioxidant activities of essential oil isolated from
the leaves of Cinnamomum tamala and Pimenta dioica, Nat. Prod. Res. 27 (2013)
290–294.
[194] Z.L. Cardeal, M.D.R. Gomes da Silva, P.J. Marriott, Comprehensive two-
dimensional gas chromatography/mass spectrometric analysis of pepper
volatiles, Rapid Commun. Mass Spectrom. 20 (2006) 2823–2836.
[195] M. Yoshimura, Y. Amakura, T. Yoshida, Polyphenolic compounds in clove and
pimento and their antioxidative activities, Biosci. Biotechnol. Biochem. 75
(2011) 2207–2212.
[196] A.R. Swain, S.P. Dutton, A.S. Truswell, Salicylates in foods, J. Am. Diet. Assoc.
85 (1985) 950–960.
[197] A.C. Martin, E. Johnston, C. Xing, A.D. Hegeman, Measuring the chemical and
cytotoxic variability of commercially available kava (Piper methysticum G.
Forster), PLoS ONE 9 (2014) e111572.
[198] Sidani, J. Campbell, Gynecology: select topics, Prim. Care 29 (2002) 297–321.
[199] J.W. Jhoo, J.P. Freeman, C.Y.W. Ang, J.J. Mihalov, Assessment of kavalactones
in kava beverage products and aqueous infusions, J. Food Sci. 71 (2006)
C345–C351.
[200] A.H. Schmidt, I. Molnar, Computer-assisted optimization in the development
of a high-performance liquid chromatographic method for the analysis of
kava pyrones in Piper methysticum preparations, J. Chromatogr. A 948 (2002)
51–63.
209
[201] A.R. Bilia, L. Scalise, M.C. Bergonzi, F.F. Vincieri, Analysis of kavalactones from
Piper methysticum (kava-kava), J. Chromatogr. B: Analyt. Technol. Biomed. Life
Sci. 812 (2004) 203–214.
[202] M.T. Cimino, DNA-based plant species identification in nutritional supple-
ments by designed nucleic acid probes, and supplement quality control uses,
U.S. Pat. Appl. Publ., US 20,070,243,533 A1 20071018 (2007).
[203] P. Lasme, F. Davrieux, D. Montet, V. Lebot, Quantification of kavalactones and
determination of kava (Piper methysticum) chemotypes using near-infrared
reflectance spectroscopy for quality control in Vanuatu, J. Agric. Food Chem.
56 (2008) 4976–4981.
[204] W. Li, S. Chen, D. Fabricant, C.K. Angerhofer, H.H.S. Fong, N.R. Farnsworth, J.F.
Fitzloff, High-performance liquid chromatographic analysis of Black Cohosh
(Cimicifuga racemosa) constituents with in-line evaporative light scattering
and photodiode array detection, Anal. Chim. Acta 471 (2002) 61–75.
[205] M. Zhao, L.J. Xu, C.T. Che, Alisolide, alisols O and P from the rhizome of Alisma
orientale, Phytochemistry 69 (2008) 527–532.
[206] L. Chen, J. Qi, Y.X. Chang, D. Zhu, B. Yu, Identification and determination of
the major constituents in Traditional Chinese Medicinal formula Danggui-
Shaoyao-San by HPLC-DAD-ESI-MS/MS, J. Pharm. Biomed. Anal. 50 (2009)
127–137.
[207] X. Liu, S.L. Li, Y. Zhou, J.Z. Song, Y.F. Zheng, G.P. Peng, H.X. Xu, Characterization
of protostane triterpenoids in Alisma orientalis by ultra-performance liquid
chromatography coupled with quadrupole time-of-flight mass spectrometry,
Rapid Commun. Mass Spectrom. 24 (2010) 1514–1522.
[208] C.M. Loescher, D.W. Morton, S. Razic, S. Agatonovic-Kustrin, High perfor-
mance thin layer chromatography (HPTLC) and high performance liquid
chromatography (HPLC) for the qualitative and quantitative analysis of Calen-
dula officinalis. Advantages and limitations, J. Pharm. Biomed. Anal. 98 (2014)
52–59.
[209] H. Neukirch, M. D’Ambrosio, J. Dalla Via, A. Guerriero, Simultaneous quan-
titative determination of eight triterpenoid monoesters from flowers of 10
Varieties of Calendula officinalis L. and characterisation of a new triterpenoid
monoester, Phytochem. Anal. 15 (2004) 30–35.
[210] V. Kaškonienė, P. Kaškonas, M. Jalinskaitė, A. Maruška, Chemical composition
and chemometric analysis of variation in essential oils of Calendula officinalis
L. during vegetation stages, Chromatographia 73 (2011) S163–S169.
[211] B. Avula, Y.H. Wang, C. Swamy Rumalla, Z. Ali, T.J. Smillie, I.A. Khan, Analyt-
ical methods for determination of magnoflorine and saponins from roots of
Caulophyllum thalictroides (L.) Michx. Using UPLC, HPLC and HPTLC, J. Pharm.
Biomed. Anal. 56 (2011) 895–903.
[212] J.M. Betz, A. Denis, T. Annette, C.E. Ryan, W.R. Obermeyer, S.W. Page, T.Z.
Woldemariam, Gas chromatographic determination of toxic quinolizidine
alkaloids in blue cohosh Caulophyllum thalictroides (L) Michx, Phytochem.
Anal. 9 (1998) 232–236.
[213] T.Z. Woldemariam, J.M. Betz, P.J. Houghton, Analysis of aporphine and quino-
lizidine alkaloids from Caulophyllum thalictroides by densitometry and HPLC,
J. Pharm. Biomed. Anal. 15 (1997) 839–843.
[214] M. Ganzera, H.R. Dharmaratne, N.P. Nanayakkara, I.A. Khan, Determination
of saponins and alkaloids in Caulophyllum thalictroides (blue cohosh) by
high-performance liquid chromatography and evaporative light scattering
detection, Phytochem. Anal. 14 (2003) 1–7.
[215] S. Subramaniam, E. Grundel, J. Roach, K.D. White, E. Mazzola, M. Ganzera, J.I.
Rader, Alkaloids and saponins in dietary supplements of blue cohosh (Caulo-
phyllum thalictroides), J. AOAC Int. 91 (2008) 21–32.
[216] M. Ganzera, E. Bedir, I.A. Khan, Separation of Cimicifuga racemosa triterpene
glycosides by reversed phase high performance liquid chromatography and
evaporative light scattering detection, Chromatographia 52 (2000) 301–304.
[217] B. Avula, Z. Ali, I.A. Khan, Chemical fingerprinting of Actaea racemosa (black
cohosh) and its comparison study with closely related actaea species (A.
pachypoda, A., podocarpa, A. rubra) by HPLC, Cromatographia 66 (66) (2007)
757–762.
[218] B. Jiang, F. Kronenberg, P. Nuntanakorn, M.H. Qiu, E.J. Kennelly, Evaluation of
the botanical authenticity and phytochemical profile of black cohosh products
by high-performance liquid chromatography with selected ion monitoring
liquid chromatography–mass spectrometry, J. Agric. Food Chem. 54 (54)
(2006) 3242–3253.
[219] K. He, B. Zheng, C.H. Kim, Direct analysis and identification of triterpene
glycosides by LC/MS in black cohosh, Cimicifuga racemosa, and in several com-
mercially available black cohosh products, Planta Med. 66 (2000) 635–640.
[220] H.K. Wang, N. Sakurai, C.Y. Shih, K.H. Lee, LC/TIS-MS fingerprint profiling of
Cimicifuga species and analysis of 23-Epi-26-deoxyactein in Cimicifuga race-
mosa commercial products, J. Agric. Food Chem. 53 (53) (2005) 1379–1386.
[221] C. Ma, A.R. Kavalier, B. Jiang, E.J. Kennelly, Metabolic profiling of Actaea species
extracts using high performance liquid chromatography coupled with elec-
trospray ionization time-of-flight mass spectrometry, J. Chromatogr. A 1218
(2011) 1461–1476.
[222] B. Jiang, C. Ma, T. Motley, F. Kronenberg, E.J. Kennelly, Phytochemical finger-
printing to thwart black cohosh adulteration: a 15 Actaea species analysis,
Phytochem. Anal. 22 (2011) 339–351.
[223] J.L. Ríos, M.C. Recio, R.M. Giner, S. Mánez, An update review of saffron and its
active constituents, Phytother. Res. 10 (1996) 189–193.
[224] L.R. Cagliani, N. Culeddu, M. Chessa, R. Consonni, NMR investigations for a
quality assessment of Italian PDO saffron (Crocus sativus L.), Food Control 50
(2015) 342–348.
[225] C.D. Kanakis, D.J. Daferera, P.A. Tarantilis, M.G. Polissiou, Qualitative deter-
mination of volatile compounds and quantitative evaluation of safranal
210 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211
and 4-hydroxy-2,6,6-trimethyl-1-cyclohexene-1-carboxaldehyde (HTCC) in
Greek saffron, J. Agric. Food Chem. 52 (2004) 4515–4521.
[226] H. Sereshti, R. Heidari, S. Samadi, Determination of volatile components of
saffron by optimized ultrasound-assisted extraction in tandem with disper-
sive liquid–liquid microextraction followed by gas chromatography–mass
spectrometry, Food Chem. 143 (2014) 499–505.
[227] S.A. Ordoudi, M. de los Mozos Pascual, M.Z. Tsimidou, On the quality
control of traded saffron by means of transmission Fourier-transform mid-
infrared (FT-MIR) spectroscopy and chemometrics, Food Chem. 150 (2014)
414–421.
[228] L. Maggi, M. Carmona, S.D. Kelly, N. Marigheto, G.L. Alonso, Geographical
origin differentiation of saffron spice (Crocus sativus L. stigmas) – prelimi-
nary investigation using chemical and multi-element (H, C, N) stable isotope
analysis, Food Chem. 128 (2011) 543–548.
[229] A. Torelli, M. Marieschi, R. Bruni, Authentication of saffron (Crocus sativus L.) in
different processed, retail products by means of SCAR markers, Food Control
36 (2014) 126–131.
[230] A. Yilmaz, N.T. Nyberg, P. Mølgaard, J. Asili, J.W. Jaroszewski, 1 H NMR
metabolic fingerprinting of saffron extracts, Metabolomics 6 (2010)
511–517.
[231] E.A. Petrakis, L.R. Cagliani, M.G. Polissiou, R. Consonni, Evaluation of saffron
(Crocus sativus L.) adulteration with plant adulterants by 1 H NMR metabolite
fingerprinting, Food Chem. 173 (2015) 890–896.
[232] H.Y. Chen, Y.H. Lin, I.H. Su, Y.C. Chen, S.H. Yang, J.L. Chen, Investigation
on Chinese herbal medicine for primary dysmenorrhea: implication from a
nationwide prescription database in Taiwan, Compl. Ther. Med. 22 (2014)
116–125.
[233] H.K. Kandikattu, R. Sakina, N. Ilaiyaraja, K. Farhath, Phytochemical analysis
and biological properties of Cyperus rotundus L, Ind. Crop Prod. 52 (2014)
815–826.
[234] I. Essaidi, H. Ben Haj Koubaier, A. Snoussi, H. Casabianca, M.M. Chaabouni, N.
Bouzouita, Chemical composition of Cyperus rotundus L. tubers essential oil
from the South of Tunisia, antioxidant potentiality and antibacterial activity
against foodborne pathogens, J. Essent. Oil Bear. Pl. 17 (2014) 522–532.
[235] E.D. Vijisaral, S. Arumugam, GC–MS analysis of ethanol extract of Cyperus
rotundus leaves, Int. J. Curr. Biotechn. 2 (2014) 19–23.
[236] M. Priya Rani, K.P. Padmakumari, HPTLC and reverse phase HPLC methods for
the simultaneous quantification and in vitro screening of antioxidant poten-
tial of isolated sesquiterpenoids from the rhizomes of Cyperus rotundus, J.
Chromatogr. B: Anal. Technol. Biomed. Life Sci. 904 (2012) 22–28.
[237] Y. Jaiswal, Z. Liang, P. Guo, H.M. Ho, H. Chen, Z. Zhao, Tissue-specific metabo-
lite profiling of Cyperus rotundus L. rhizomes and (+)-nootkatone quantitation
by laser microdissection, ultra-high-performance liquid chromatography-
quadrupole time-of-flight mass spectrometry, and gas chromatography-mass
spectrometry techniques, J. Agric. Food Chem. 62 (2014) 7302–7316.
[238] X. Li, Determination of ferulic acid content in Cyperus rotundus by HPLC, J.
Chem. Pharm. Res. 6 (2014) 1496–1500.
[239] S.B. Badgujar, V.V. Patel, A.H. Bandivdekar, Foeniculum vulgare Mill: a review
of its botany, phytochemistry, pharmacology, contemporary application, and
toxicology, Biomed. Res. Int. 2014 (2014) 842674.
[240] Z. Ghodsi, M. Asltoghiri, The effect of fennel on pain quality, symptoms, and
menstrual duration in primary dysmenorrhea, J. Pediatr. Adolesc. Gynecol. 27
(2014) 283–286.
[241] Piras, D. Falconieri, S. Porcedda, B. Marongiu, M.J. Gonçalves, C. Cavaleiro, L.
Salgueiro, Supercritical CO2 extraction of volatile oils from Sardinian Foenicu-
lum vulgare ssp. vulgare (Apiaceae): chemical composition and biological
activity, Nat. Prod. Res. 28 (2014) 1819–1825.
[242] P.J. Maikhunthod, Marriott, Aroma-impact compounds in dried spice as a
quality index using solid phase microextraction with olfactometry and com-
prehensive two-dimensional gas chromatography, Food Chem. 141 (2013)
4324–4332.
[243] M.A. Strehle, P. Rösch, M. Baranska, H. Schulz, J. Popp, On the way to a qual-
ity control of the essential oil of fennel by means of Raman spectroscopy,
Biopolymers 77 (2005) 44–52.
[244] L. Mattoli, F. Cangi, C. Ghiara, M. Burico, A. Maidecchi, E. Bianchi, E. Ragazzi, L.
Bellotto, R. Seraglia, P. Traldi, A metabolite fingerprinting for the characteri-
zation of commercial botanical dietary supplements, Metabolomics 7 (2011)
437–445.
[245] K. Dhalwal, V.M. Shinde, K.R. Mahadik, A.G. Namdeo, Rapid densitomet-
ric method for simultaneous analysis of umbelliferone, psoralen, and
eugenol in herbal raw materials using HPTLC, J. Sep. Sci. 30 (2007)
2053–2058.
[246] M. Greule, C. Haensel, U. Bauermann, A. Mosandl, Feed additives: authen-
ticity assessment using multicomponent-/multielement-isotope ratio mass
spectrometry, Eur. Food Res. Technol. 227 (2008) 767–776.
[247] A. Pasrija, R. Singh, C.K. Katiyar, Standardization of Fennel (Foeniculum vul-
gare), its oleoresin and marketed ayurvedic dosage forms, Int. J. Pharm. Sci.
Drug Res. 3 (2011) 265–269.
[248] S.K. Seo, Y. Hong, B.H. Yun, S.J. Chon, Y.S. Jung, J.H. Park, S.H. Cho, Y.S. Choi, B.S.
Lee, Antioxidative effects of Korean red ginseng in postmenopausal women:
a double-blind randomized controlled trial, J. Ethnopharmacol. 154 (2014)
753–757.
[249] H. Yang, D.Y. Lee, K.B. Kang, J.Y. Kim, S.O. Kim, Y.H. Yoo, S.H. Sung, Identifica-
tion of ginsenoside markers from dry purified extract of Panax ginseng by a
dereplication approach and UPLCQTOF/MS analysis, J. Pharm. Biomed. Anal.
109 (2015) 91–104.
[250] S.H. Baek, O.N. Bae, J.H. Park, Recent methodology in ginseng analysis, J. Gin-
seng Res. 36 (2012) 119–134.
[251] L. Wang, W.S. He, H.X. Yan, Y. Jiang, K.S. Bi, P.F. Tu, Performance eval-
uation of charged aerosol and evaporative light scattering detection for
the determination of ginsenosides by LC, Chromatographia 70 (2009)
603–608.
[252] W. Wu, F. Song, D. Guo, J. Mi, Q. Qin, Q. Yu, S. Liu, Mass spectrometry-based
approach in ginseng research: a promising way to metabolomics, Curr. Anal.
Chem. 8 (2012) 43–66.
[253] E.J. Lee, R. Shaykhutdinov, A.M. Weljie, H.J. Vogel, P.J. Facchini, S.U. Park,
Y.K. Kim, T.J. Yang, Quality assessment of ginseng by 1 H NMR metabo-
lite fingerprinting and profiling analysis, J. Agric. Food Chem. 57 (2009)
7513–7522.
[254] M.N. Boukhatem, A. Kameli, F. Saidi, Essential oil of Algerian rose-scented
geranium (Pelargonium graveolens): chemical composition and antimicro-
bial activity against food spoilage pathogens, Food Control 34 (2013)
208–213.
[255] R.K. Verma, R.S. Verma, A. Chauhan, A. Singh, L. Rahman, A. Kalra, Biomass
yield, essential oil yield and oil quality of rose-scented geranium (Pelargo-
nium graveolens L.), intercropped with vegetables, Int. J. Agric. Res. 6 (2011)
830–839.
[256] M. Sandasi, G.P.P. Kamatou, C. Gavaghan, M. Baranska, A.M. Viljoen, A qual-
ity control method for geranium oil based on vibrational spectroscopy and
chemometric data analysis, Vib. Spectrosc. 57 (2011) 242–247.
˛
[257] L. Kuźma, D. Kalemba, M. Różalski, B. Różalska, M. Wieckowska-Szakiel, U.
Krajewska, H. Wysokińska, Chemical composition and biological activities
of essential oil from Salvia sclarea plants regenerated in vitro, Molecules 14
(2009) 1438–1447.
[258] S. Nasermoadeli, V. Rowshan, Comparison of Salvia sclarea L. essential oil
components in wild and field population, Int. J. Agric. Crop Sci. 5 (2013)
828–831.
[259] H. Yalcin, I. Ozturk, E. Tulukcu, O. Sagdic, Effect of ␥-irradiation on bioactivity,
fatty acid compositions and volatile compounds of clary sage seed (Salvia
sclarea L.), J. Food Sci. 76 (2011) C1056–C1061.
[260] L. Ciesla, D. Staszek, M. Hajnos, T. Kowalska, M. Waksmundzka-Hajnos, Devel-
opment of chromatographic and free radical scavenging activity fingerprints
by thin-layer chromatography for selected Salvia species, Phytochem. Anal.
22 (2011) 59–65.
[261] M. Lis-Balchin, S.G. Deans, E. Eaglesham, Relationship between bioactivity and
chemical composition of commercial essential oils, Flavour Frag. J. 13 (1998)
98–104.
[262] De Palma, R. Rossi, M. Carai, C. Cabras, G. Colombo, L. Arnoldi, N. Fuzzati, A.
Riva, P. Morazzoni, P.L. Mauri, Pharmaceutical and biomedical analysis of ter-
pene constituents in Salvia miltiorrhiza, Curr. Pharm. Anal. 4 (2008) 249–257.
[263] R. Kasimu, K. Tanaka, Y. Tezuka, Z.N. Gong, J.X. Li, P. Basnet, T.
Namba, S. Kadota, Comparative study of seventeen Salvia plants: aldose
reductase inhibitory activity of water and MeOH extracts and liquid
chromatography–mass spectrometry (LC–MS) analysis of water extracts,
Chem. Pharm. Bull. 46 (1998) 500–504.
[264] Z.L. Ye, C.C. Hu, X.H. Fan, Y.Y. Cheng, Simultaneous analysis of seven major
saponins in compound danshen dropping pills using solid phase extraction
and HPLC with DAD and ESI-MS detectors, J. Liq. Chromatogr. Relat. Technol.
29 (2006) 1575–1587.
[265] G.X. Zhong, P. Li, L.J. Zeng, J. Guan, D.Q. Li, S.P. Li, Chemical characteristics
of Salvia miltiorrhiza (Danshen) collected from different locations in China, J.
Agric. Food Chem. 57 (2009) 6879–6887.
[266] M. Gu, Z. Su, F. Ouyang, Fingerprinting of salvia miltiorrhiza bunge by
thin-layer chromatography scan compared with high speed countercurrent
chromatography, J. Liq. Chromatogr. Relat. Technol. 29 (2006) 1503–1514.
[267] A. Pareek, M. Suthar, G.S. Rathore, V. Bansal, Feverfew (Tanacetum parthenium
L.): a systematic review, Pharmacogn. Rev. 5 (2011) 103–110.
[268] S. Heptinstall, D.V. Awang, B.A. Dawson, D. Kindack, D.W. Knight, J. May,
Parthenolide content and bioactivity of feverfew (Tanacetum parthenium (L.)
Schultz-Bip.). Estimation of commercial and authenticated feverfew products,
J. Pharm. Pharmacol. 44 (1992) 391–395.
[269] E.A. Abourashed, I.A. Khan, Determination of parthenolide in selected fever-
few products by liquid chromatography, J. AOAC Int. 83 (2000) 789–792.
[270] E. Ernst, M.H. Pittler, The efficacy and safety of feverfew (Tanacetum parthe-
nium L.): an update of a systematic review, Public Health Nutr. 3 (2000)
509–514.
[271] N.J. Bailey, J. Sampson, P.J. Hylands, J.K. Nicholson, E. Holmes, Multi-
component metabolic classification of commercial feverfew preparations via
high-field 1 H-NMR spectroscopy and chemometrics, Planta Med. 68 (2002)
734–738.
[272] H. Salapovic, J. Geier, G. Reznicek, Quantification of sesquiterpene lactones in
asteraceae plant extracts: evaluation of their allergenic potential, Sci. Pharm.
81 (2013) 807–818.
[273] J.S. Chaves, F. Batista Da Costa, L.A. Pedro de Freitas, Development of enteric
coated tablets from spray dried extract of feverfew (Tanacetum parthenium
L.), Braz. J. Pharm. Sci. 45 (2009) 573–584.
[274] B. Avula, Y.H. Wang, Z. Ali, T.J. Smillie, I.A. Khan, Chemical fingerprint analysis
and quantitative determination of steroidal compounds from Dioscorea vil-
losa, Dioscorea species and dietary supplements using UHPLC-ELSD, Biomed.
Chromatogr. 28 (2014) 281–294.
[275] K.D. Yoon, Y.W. Chin, M.H. Yang, J. Choi, J. Kim, Application of high speed
countercurrent chromatography-evaporative light scattering detection for
 M. Masullo et al. / Journal of Pharmaceutical and Biomedical Analysis 113 (2015) 189–211 211

the separation of seven steroidal saponins from Dioscorea villosa, Phytochem.
Anal. 23 (2012) 462–468.
[276] D.J. Yang, T.J. Lu, S.H. Lucy, Simultaneous determination of furostanol and
spirostanol glycosides in Taiwanese Yam (Dioscorea spp.) Cultivars by HPLC,
J. Food Drug. Anal. 2 (2003) 271–276.
[277] Y.K. Kwon, E.Y. Jie, A. Sartie, D.J. Kim, J.R. Liu, B.W. Min, S.W. Kim, Rapid
metabolic discrimination and prediction of dioscin content from African yam
tubers using Fourier transform infrared spectroscopy combined with multi-
variate analysis, Food Chem. 166 (2015) 389–396.