How does chromatography play a role in separating and analyzing phytochemicals?

Chromatography is the lynchpin of phytochemistry and is the key to

obtaining pure compounds for structure elucidation, for
pharmacological testing or for development into therapeuticals. It also
plays a fundamental role as an analytical technique for quality control
and standardisation of phytotherapeuticals. Although liquid
chromatography is barely 100 years old, an extraordinary variety of
instrumental and ancillary equipment is available, notably in the
domain of high-performance liquid chromatography. It is impossible
to touch all areas of chromatography in such a review but certain areas
are worthy of mention: HPLC, HPTLC, UPLC and countercurrent
chromatography.
Another important addition has been the development of hyphenated
techniques involving HPLC: LC/UV, LC/MS, LC/MS and LC/NMR. n

These are indispensable nowadays for the early detection and

identification of compounds in crude plant extracts.

Graphical abstract

Chromatography is essential to phytochemistry and is the key to

obtaining pure compounds for structure
elucidation (by countercurrent chromatography, for example), for
pharmacological testing or for development into therapeuticals. It also
plays a fundamental role as an analytical technique for quality control
and standardisation of phytotherapeuticals.

1. Download : Download full-size image

Introduction

Over the past 100 years, chromatography has developed from a

rudimentary tool for the separation of pigments into an array of
techniques capable of dealing with the most complex analytical and
purification problems in phytochemistry. The developments can
basically be divided into three important landmarks: the first
introduction of chromatography; the contribution played by Martin in
the 1950s; and the introduction of commercially-available high-
performance liquid chromatography (HPLC) equipment in the 1970s.
A list of the most frequently employed chromatographic methods is
given in Table 1.
In a review of this type, it is impossible to include all aspects of
chromatography and attention will be concentrated on certain key
areas, such as HPLC.

Section snippets

Historical aspects

Tswett introduced column adsorption chromatography at the

beginning of the twentieth century, initially for the separation of plant
pigments. The first printed description of the method came with a
lecture in 1903 at the Warsaw Society of Natural Sciences entitled “On
a new category of adsorption phenomena and their application to
biochemical analysis” (Engelhardt, 2004). This was the starting point
of liquid chromatography and in a paper published in 1906, Tswett
took the subject further with
High-performance thin-layer chromatography (HPTLC)

Thin layer chromatography (TLC) is certainly not to be forgotten. It is

the only chromatographic method offering the option of presenting
the result as an image. Furthermore, TLC is the sole technique in
which all the components of the sample are included in the
chromatogram. In contrast, HPLC and GC are selective and not all of
the compounds in the sample are included in the display.
The real breakthrough in TLC came through the work of Stahl, who
introduced the use of calcium sulphate as
High-performance liquid chromatography (HPLC)

The most remarkable advances in chromatography have occurred in

the domain of HPLC, despite the fact that the technique itself has only
been in existence for about 40 years. The year 1967 was a landmark in
the introduction of HPLC, with papers from Horvath, Huber and
Scott, but the first automatic liquid chromatograph with gradient
elution was an amino acid analyser described by Moore and co-
workers in 1958 (Spackman et al., 1958).
Until the advent of HPLC, most phytochemical separations were
Capillary electrophoresis (CE)

Capillary electrophoresis is an analytical technique which provides

high separation efficiency and short run times. Several modes of CE
are available: (1) capillary zone electrophoresis (CZE), (2) micellar
electrokinetic chromatography (MEKC), (3) capillary gel
electrophoresis (CGE), (4) capillary isoelectric focusing, (5) capillary
isotachophoresis, (6) capillary electrochromatography (CEC) and (7)
non-aqueous CE. The simplest and most versatile CE mode is CZE, in
which the separation is based
Countercurrent chromatography (CCC)

Countercurrent chromatography is another advance of the twentieth

century with tremendous implications for phytochemistry. Unlike
HPLC, there is no solid column packing involved. It is basically a
development of countercurrent distribution (CCD), a method
developed in the 1940s and 1950s for the batchwise (Craig
Distribution) or continuous (O’Keefe Distribution) fractionation of
mixtures. Countercurrent chromatography (or centrifugal partition
chromatography, as it is sometimes known) is an
Perspectives

Without chromatography, it is safe to say that phytochemistry would

be a great deal more tedious; analytical procedures would be more
complex and certain problems would be impossible to tackle, such as
metabolomic analysis. The challenges of proteomics and
metabolomics are already omnipresent, with multi-component
mixtures of high complexity to analyse. Without doubt, advances in
HPLC, via new column technology, such as capillary columns or
UPLC, will provide solutions to these problems, in
Andrew Marston was born in Zambia (1953). He studied chemistry
at University College London and obtained a PhD in the Department
of Organic Chemistry at Liverpool University, U.K. He was then
awarded a Royal Society / NATO fellowship to perform postdoctoral
research on plant-derived tumour promoters at the German Cancer
Research Centre in Heidelberg. He is now a senior research scientist in
the group of Professor Kurt Hostettmann, working in the fields of
pharmacognosy and phytochemistry. This
Special issue articlesRecommended articles
References (36)
 H. Engelhardt
One century of liquid chromatography. From Tswett’s columns to
modern high speed and high performance separations
J. Chromatogr. B
(2004)
 Y. Ito
Golden rules and pitfalls in selecting optimum conditions for high-
speed countercurrent chromatography
J. Chromatogr. A
(2005)
 Y. Ito et al.
pH-zone-refining countercurrent chromatography
J. Chromatogr. A
(1996)
 K. Kobayashi et al.
Isotheasaponins B1–B3 from Camellia sinensis var. sinensis tea leaves
Phytochemistry
(2006)
  A. Marston et al.
Developments in the application of countercurrent chromatography
to plant analysis
J. Chromatogr. A
(2006)
 R. Shellie et al.
Characterisation of lavender essential oils by using gas
chromatography–mass spectrometry with correlation of linear
retention indices and comparison with comprehensive two-
dimensional gas chromatography
J. Chromatogr. A
(2002)
 T. Visser et al.
Gradient elution liquid chromatography-infrared spectrometry at
μg/l level using capillary column switching and addition of a make-up
liquid. A preliminary study
Anal. Chim. Acta
(1997)
 G. Bringmann et al.
HPLC-CD on-line coupling in combination with HPLC-NMR and
HPLC-MS/MS for the determination of the full absolute
stereostructure of new metabolites in plant extracts
Anal. Chem.
(1999)
 P.R. Brown
High-performance liquid chromatography: past developments,
present status and future trends
Anal. Chem.
(1990)
 E.C.Y. Chan et al.
Ultra-performance liquid chromatography/time of flight mass
spectrometry based metabolomics of raw and steamed Panax
notoginseng
Rapid Commun. Mass Spectrom.
(2007)
2 Could you elaborate on the differences between various separation
techniques like distillation, extraction, and chromatography in the
context of phytochemical isolation?

Requests left today: 0

MENU
Close menu
Night mode
Select language

Hello! How can I help you?

Could you elaborate on the differences between various separation

techniques like distillation, extraction, and chromatography in the
context of phytochemical isolation?

Certainly! In the context of phytochemical isolation, different

separation techniques are used to extract and separate specific
compounds from plant materials. Here are some key differences
between distillation, extraction, and chromatography:

Distillation: It is a technique based on differences in boiling points of

the components of a mixture. Distillation is particularly useful for
separating volatile compounds. In phytochemical isolation, steam
distillation is commonly employed to extract essential oils from plants.
The plant material is heated, and the volatile compounds are
vaporized and collected as condensate.

Extraction: Extraction is the process of obtaining desired compounds

by dissolving them in a suitable solvent. Different extraction
techniques, such as maceration, Soxhlet extraction, or ultrasound-
assisted extraction, can be used. In phytochemical isolation, extraction
helps to isolate a wide range of compounds, including non-volatile
ones like polyphenols, alkaloids, or flavonoids.

Chromatography: Chromatography techniques separate mixtures by

exploiting the differences in distribution between the mobile phase
(solvent) and stationary phase (solid support or stationary phase
liquid). There are various chromatography methods like column
chromatography, thin-layer chromatography (TLC), gas
chromatography (GC), or high-performance liquid chromatography
(HPLC). These methods are highly effective for isolating and
analyzing specific plant compounds. For example, HPLC is commonly
used for separating and quantifying different phytochemicals, while
TLC is a quick and inexpensive technique to determine the presence
of specific compounds.

Each technique has its own advantages and limitations. The choice of
technique depends on the nature of the target compound, the
complexity of the mixture, and the desired outcome of the
phytochemical isolation process. It is often necessary to employ a
combination of techniques to achieve optimal separation and
purification of phytochemicals.

3
Are there specific phytochemical groups that are commonly targeted
in research or industry, and if so, why?

Yes, there are several specific phytochemical groups that are

commonly targeted in research and industry due to their potential
pharmacological, nutraceutical, or industrial applications. Here are
some examples:

Polyphenols: Polyphenols are a diverse group of compounds found in

plants, including flavonoids, phenolic acids, and tannins. They are
known for their antioxidant and anti-inflammatory properties and have
been extensively studied for their potential health benefits, such as
cardiovascular protection, anticancer effects, and neuroprotective
properties.

Terpenoids: Terpenoids, also called isoprenoids, are a large class of

compounds derived from isoprene units. They include various classes
such as monoterpenes, sesquiterpenes, diterpenes, and triterpenes.
Terpenoids exhibit a wide range of biological activities, including
antimicrobial, anticancer, antimalarial, and anti-inflammatory effects.
They are also important in the production of natural flavors,
fragrances, and pharmaceuticals.
Alkaloids: Alkaloids are naturally occurring nitrogen-containing
compounds, often with pronounced physiological effects. Many
alkaloids have been identified as bioactive substances with
therapeutic potentials. Examples of notable alkaloids include
morphine, quinine, caffeine, and taxol. They have diverse
pharmacological properties, such as analgesic, anticancer,
antimicrobial, and antimalarial activities.

Glycosides: Glycosides are compounds consisting of a sugar

molecule (glycone) attached to a non-sugar moiety (aglycone). They
are widespread in plants and have various biological activities. For
instance, cardiac glycosides like digitoxin and digoxin have been used
to treat heart failure. Other glycosides, such as saponins, exhibit
antimicrobial, anticancer, and antifungal properties.

These phytochemical groups are often targeted due to their potential

therapeutic properties, their abundance in plant sources, and the
established knowledge regarding their bioactivities. Research and
industry focus on isolating and characterizing these compounds to
determine their potential applications in drug development,
nutraceutical products, functional foods, cosmetics, and natural
product-based industries.
Skip to Main content
Elsevier logo

Phytochemical
Since the word “phytochemical” is originated from appending the word
“chemical” to the Greek word “phyto,” phytochemical is defined as
non-nutrient plant chemicals with potential health benefits to reduce
the risk of chronic diseases.

From: Advances in Food and Nutrition Research, 2022

Related terms:
NanoparticleAntioxidantCarotenoidPolyphenolAnthocyaninBioactive
CompoundFunctional FoodFlavonoidAntioxidant ActivityMetabolite
View all Topics
Phytochemicals
Monika Thakur, … Renu Khedkar, in Functional and Preservative
Properties of Phytochemicals, 2020

3 Types of phytochemicals
Phytochemicals have great antioxidant potential and are of great
interest due to their beneficial effects on health of human beings, and
they give immense health benefits to the consumers. Epidemiological
and animal trials suggest that the regular consumption of fruits,and
vegetables, and whole grains reduces the risk of various diseases
linked with oxidative damage (Cieslik et al., 2006; Scalbert et al.,
2005; Kris-Etherton et al., 2002). The natural antioxidants are
classified into two categories namely in vitro and in vivo antioxidants.
Free radical scavengers act as hydrogen donors, electron donor,
peroxide decomposer, singlet oxygen quencher, enzyme inhibitor,
synergist, and metal-chelating agents.

Among the phytochemicals mentioned as potentially providing health

benefits are polyphenols, flavonoids, isoflavonoids, anthocyanidins,
phytoestrogens, terpenoids, carotenoids, limonoids, phytosterols,
glucosinolates, and fibers. These antioxidant-rich phytochemicals are
explained below in Table 11.1.

Table 11.1. Antioxidant-rich phytochemicals with their food sources

and health benefits.

Sr. No. Phytochemicals Sources Health benefits References

1 Carotenoids Carrots, tomatoes, parsley, orange and green
leafy vegetables, chenopods, fenugreek, spinach, cabbage, radish,
turnips Antioxidants protect against uterine, prostate, colorectal,
lung, and digestive tract cancersAgarwal and Rao (2000); Britton
(1995); Johnson (2002); Elliott (2005); Ribaya-Mercado and Blumberg
(2004); Prakash et al. (2004); Astrog (1997); Bertram (1999); Paiva
and Russell (1999)
2 Phytosterols Vegetables, nuts, fruits, seeds Suppress the
growth of diverse tumors cell lines via initiation of apoptosis and
concomitant arrest of cells in the G1 phase of the cell cycle John et al.
(2007); von Bergmann et al. (2005); Dillard and German (2000)
3 Limonoids Citrus fruits Inhibiting phase I enzymes and
inducing phase II detoxification enzymes in liver, provide protection to
lung tissue. Detoxify enzymes Ozaki et al. (1995); Lam et al. (1994);
Willcox et al. (2004)
4 Polyphenols
➢Flavonoids
➢Isoflavonoids
➢Anthocyanidins Fruits, vegetables, cereals, beverages, legumes,
chocolates, oilseeds Action against free radicals, free radicals
mediated cellular signaling, inflammation, allergies, platelet
aggregation, and hepatotoxins Kaul and Kapoor (2001); Scalbert et
al. (2005); Cieslik et al. (2006); Prakash and Kumar (2011); Packer
and Weber (2001); Gee and Johnson (2001); Willcox et al. (2004);
Arts and Hollman (2005); Andjelkovic et al., (2006); Prakash et al.
(2007); Singh et al. (2009); Anagnostopoulou et al. (2006); Sharma et
al. (2003); Singh et al. (2003, 2004, 2008a, 2008b); Kris-Etherton et
al. (2002); Dore (2005); Croteau et al. (2000); Ko et al. (2010); Patel et
al. (2001).
5 Glucosinolates Cruciferous vegetables Protection against
cancer of colon, rectum, and stomach Conaway et al. (2001); Cartea
and Velasco (2008); Hayes et al. (2008); Vig et al. (2009); Traka and
Mithen (2009)
6 Phytoestrogen Legumes, berries, whole grains, cereals, red
wine, peanuts, red grapes Protection against bone loss and heart
disease, cardiovascular diseases, breast and uterine cancers
Morabito et al. (2002); Prakash and Gupta (2011); Sakamoto et
al. (2010); Dip et al. (2009); Mense et al. (2008); Fremont (2000); Cos
et al. (2003)
7 Terpenoids (Isoprenoids) Mosses, liverworts, algae, lichens,
mushrooms Antimicrobial, antiparasitic, antiviral, antiallergic,
antiinflammatory, chemotherapeutic, antihyperglycemic,
antispasmodic Tholl (2006); Langenheim (1994); Lee et al. (2003);
Paduch et al. (2007); Hammer et al. (2003); Singh et al. (1999);
Prakash and Kumar (2011); Prakash and Gupta (2009); Ikeda et al.
(2002)
8 Fibers Fruits and vegetables (green leafy), oats Reduces
blood cholesterol, cardiovascular disease Narasinga Rao (1988,
2003); Schnecman (1989); Dillard and German (2000); Packer and
Weber (2001)
9 Polysaccharides Fruits and vegetablesAntimicrobial,
antiparasitic, antiviral, antiallergic, antiinflammatory, lowering serum,
enhances defense mechanisms Bnouham et al. (2006); Lopez (2007);
Atherton (2002); Schmidgall et al. (2000)
10 Saponins Oats, leaves, flowers, and green fruits of tomato
Protection against pathogens, antimicrobial, antiinflammatory,
antiulcer agent Hostettmann and Marston (1995); Morrissey and
Osbourn (1999); Price et al. (1987); Mert-Turk (2006)
Phytochemicals
Manoj Kumar Singh, … Ajay Kumar, in Functional and Preservative
Properties of Phytochemicals, 2020

1.4 Use claim of phytochemicals in patents

The phytochemicals are used as functional food, soft drinks, and
many other food items, which are having good nutrient value and
significant importance, economically. To this Szilbereky et al. (2018)
(Fitorex Kft., EP2908664B1) have published a raw material for
functional foods in which the seeds of cereals, leguminous plants, soy-
beans, rice, wheat, oat, chick pea, or maize have been used.
Similarly, Seth (2018) (US20180352834A1) has disclosed a food
composition of palmitic acid, linoleic acid, p-coumaric acid, oleic acid,
caffeic acid, pantothenic acid, and antioxidative agents including
vitamins E, C, B1, or B6. The claimed food composition is applicable
for most of the mammals (e.g., dog food).

In India, phytochemicals, as well as medicinal plants, have remained

the most abundant source of health care and life improvement since
very long. India is the richest source of traditional herbal plants with
their prescriptions. In India, Ayurvedic, Unani and Siddha medico-
therapeutics are playing a very important role in the society since
ancient time. Ayurveda is approximately 5000 years old and
predominantly uses phytochemicals in their preparations and
formulations. Now in modern era, about 24%–27% drugs are derived
from the plant sources. Several synthetic drugs also have been
developed as the analogs/prototype of the natural phytochemicals,
which serve as lead compounds for these synthetic drugs.

India, being one of the richest plant biodiversity countries in the world,
has Western Ghats and Himalayas as the regions rich in plant
biodiversity in the country. About 7500 plant species out of 43,000
(that exist in the country) are recorded in various medicines, and
∼1700 species are acknowledged in the Ayurvedic literature. In India,
phytochemicals are not limited to medicinal use only, but also they
have been used in cosmetics, health and hygiene, fragrance, and food
supplements.

The IPR aspect for phytochemicals developed in two schemes: (a)

The phytochemicals which are extracted from the natural plants and
their use; and (b) Scientifically advanced version of medicinal plants
(improved plant varieties) and their extracted phytochemicals. The
improvement has been achieved by several private agencies in terms
of yield, intrinsic quality, and more resistance or tolerance against pest
and diseases. To improve/produce the phytochemicals from the
plants, several advanced technologies have been used such as
genetic engineering, molecular techniques, etc. Hence, more legal
protection has been required for various processes and products of
molecular breeding.

A review on marine-based phytochemicals and their application in

biomedical research
Rousan Khatun, … Alok Prasad Das, in Recent Frontiers of
Phytochemicals, 2023

Abstract
Phytochemicals are involved in specific biological activities, and they
are the natural bioactive substances which are derived from
phytonutrients (i.e., plant foods). In the field of biomedicine, various
phytochemicals may play a protective role against diseases and
infections. Phytochemicals like phenolics, alkaloids, terpenes, and
organosulfides are considered secondary metabolites of plant foods.
Currently, marine natural products have been recognized as the
utmost consequential origin of bioactive substances. Biomedicines
include lots of marine-based phytochemical substances associated
simultaneously. The diagrammatic dissimilarities of these marine
phytochemical substances come from various alteration reactions of
the main fluxion. For decades, various phytochemicals have illustrated
distinct biomedical and therapeutic effects on the health of human
beings such as immune response and antimicrobial. The ongoing
headway shows the direction to appreciating the gradual development
to recognize narrative dominating properties of marine-based
phytochemical substances for biomedical. This chapter summarizes
stocking research of marine-based phytochemical substances, which
are directly applicable to biomedical as well as therapeutics. All these
phytochemicals are stably visualized in various considerable studies
(i.e., in vitro, in vivo, and clinical studies) with high potential
protectiveness and therapeutics on many infections and diseases that
subsequently relate to the biomedicals.

Date palm (Phoenix dactylifera L.) secondary metabolites: Bioactivity

and pharmaceutical potential
Heba I. Mohamed, … Jameel M. Al-Khayri, in Phytomedicine, 2021

2.8 Phytochemical compounds in dates

Phytochemicals are non-nutrient bioactive components that are
primarily responsible for scavenging toxic radicals after oxidative
stress by generating antioxidants, the main cause of most chronic
diseases (Al-Harrasi et al., 2014). Fruit phytochemicals displayed high
antioxidant capacities linked to lower incidence of degenerative
diseases and lower mortality average in humans (Baliga et al., 2011;
Wang et al., 2013). Phenolic acids, flavonoids, tannins, carotenoids,
isoflavons, sterols, and lignans are some of date phytochemicals of
the fruits that are considered bioactive. Table 1 includes the role of
these phytochemicals and their levels in date fruits when it comes to
disease prevention.

Table 1. Phytochemicals and the role of date varieties (Gnanamangai

et al., 2019).
Phytochemical Quantity Function References
Carotenoid:
Β Carotene 3%–10% Protects testicular functions and have
gonadotropic and hepatoprotective activity Boudries, Kefalas, and
Hornero-Mendez (2007), Domitrovic, Jakovac, Grebic, Milin, and
Radosevic-Stasic (2008), Eustache et al. (2009), Jana et al. (2008),
Janbaz, Saeed, and Gilani (2005), Said, Banni, Kerkeni, Said, and
Messaoudi (2010)
Lutein 89%–94%
Minor Carotenoids 2%–8%
Phenols:
Cinnamic acids, acetylated flavonols, caffeoglshikimic acid hexoside,
caffeoylsinapoyl monohexoside, dihexoside, hydroxyl benzoates,
hydroxyl cinnamates 30,000 ppm (3.0 g/100 g) Less glucose
absorbance, antioxidant activity, inhibits Angiotensin II converting
enzymes and reduces hypertension and anticancer Neori et al.
(2013), Surh (2003)
Phenolic acids:
Gallic acid, protocatechuic, phydroxyl benzoic, vanillic, caffeic,
dactyliferic acid, ellagic acid, syringic, p-coumaric ferulic, o-coumaric
14.18–49.67 mg/100 g (bound) Hepatoprotective Vayalil,
(2002), Al-Farsi and Lee (2008), Al-Farsi, Alasalvar, Morris, Baron,
and Shahidi (2005), Janbaz et al. (2005), Domitrovic et al. (2008)
6.1–14.8 mg/100 g (free)
Tannins:
Flavanoids, flavones, flavonols, flavanones, procynidines,
proanthocyanidins, flavonoid glycosides (luteolin, quercetin, apigenin),
Chrysoeriol, isohamnetic, anthocyanins 0.0162–5446 g/kg
Hepatoprotective Hong, Tomas-Barberan, Kader, and
Mitchell (2006), Al-Hooti et al. (1998), Janbaz et al. (2005), Domitrovic
et al. (2008)
Sterols:
Phytosterols:
Campesterol, β-sitosterol, lupenone, lupeol, 24-methylene cy-
cloartanol, propylidene cholesterol 1.83%–2.57%
Hepatoprotective, inhibits cholesterol synthesis and causes
hypocholestrolemia John, Sorokin, and Thompson (2007), Liolios,
Sotiroudis, and Chinou (2009)
Phytoestrogens:
Isoflavones, lignans, genistein, daidzein Anticancer and
antidiabetic Bhathena and Velasquez (2002)
Phytochemicals and cancer
Mayuri Iyer, … Vijay Upadhye, in Recent Frontiers of Phytochemicals,
2023

Abstract
Phytochemicals are the natural chemicals present in plants. These
chemicals help protect plants from germs, fungi, bugs, and other
threats. On the basis of their chemical properties and structures,
phytochemicals are broadly classified into six categories:
carbohydrates, lipids, phenolics, terpenoids, alkaloids, and other
nitrogen-containing compounds. These include antioxidants, dietary
fibers, immune-activating agents, and detoxifying agents. In the last
30 years, phytochemicals have been researched for their essential
role in preventing various chronic diseases like diabetes, arthritis,
coronary heart disease, Alzheimer’s, and cancer. Cancer is one of the
deadliest and leading causes of death worldwide, and the treatment
involves the use of anticancer drugs having various side effects and
toxicity. Extensive research is going on to discover new strategies or
drugs that can prevent chemotoxicity by increasing a cancer patient’s
survivability and quality of life. Recently, many reports describe
studies on assessing the chemoprotective effects of phytochemicals
and thereby potential in ameliorating cancer progression.

This chapter aims to summarize the current knowledge on anticancer

phytochemicals. The first part describes different types of
phytochemicals from edible and nonedible plant materials possessing
anticancer properties, their mechanism of action, and molecular
targets. The second part is a compilation of various in vitro, in vivo,
and clinical studies on phytochemicals’ use in different cancer types.
The last part describes recent developments and gaps in
phytochemical-based drug discovery. In the end, this chapter
accentuates the importance of phytochemicals and their usage in daily
diet.

Phytochemicals: recent trends in food, pharmacy, and biotechnology

Ayushman Gadnayak, Budheswar Dehury, in Recent Frontiers of
Phytochemicals, 2023

5.1 Introduction
Phytochemicals, often considered as phytonutrients, are bioactive and
natural compounds rich in various foods like fruits, nuts, whole grain
products as well as legumes, and dark chocolates. There are
numerous phytochemicals in which only a small amount is being
isolated and identified from the plants (Gómez-García et al., 2020).
The most common phytochemicals found in food are carotenoids,
flavonoids, ligands, saponins, cyanidin, isoflavones, polyphenols, and
indole. The biodiversity in the resources of the phytochemicals has
given a renewable and unique resource to discover the potential of
using such agents in the discovering novel drugs, development of
therapeutics against cancer, and supplementary foods. Much of the
present evidence on the advantages of the phytochemicals have
come from previous researches and their clinical applications. After
intake of phytochemicals in their diet, individuals have shown
significantly lower risk of certain types of cancers as well as
cardiovascular diseases. However, there is no accurate evidence of
using any specific phytochemical to eliminate the chances of cancer.
However, there are promising evidences that indicates the potential
use of phytochemicals to:

•
Protect the cells and DNA from damage that in turn leads to lower the
risk of cancer,

•
Enhance the immune system,

•
Reduce inflammations,
•
Slow rate of growth for some of the cancer cells,

•
Regulate the hormones, and

•
Lowers the risks of chronic diseases such as cardiovascular diseases
and obesity.

Phytochemicals including carotenoid, flavonoids, anthocyanin,

isothiocyanates, and zeaxanthin helps in inhibiting the growth of
cancer cells and lowers the occurrence of cardiovascular diseases.
Carotenoid can be found in cooked tomatoes, carrots, orange, and
sweet potatoes. On the other hand, flavonoids that are found in
berries, apples, soybeans, coffee, walnuts, and whole grains have the
capability to fight inflammation, reduce tumor growth, and decrease
the chances of DNA damage. Anthocyanins, found in berries help in
lowering blood pressure, therefore, they can be used as a daily
remedy for maintaining hypertension. Lastly, zeaxanthin is found in
leafy green lie chard, and spinach is demonstrated to promote eye
health (Roswellparki, 2019).

Novel extraction and characterization methods for phytochemicals

Ratnnadeep C. Sawant, … S. Abdul Kader, in Recent Frontiers of
Phytochemicals, 2023

4.4.1.2 Phytochemical screening assay

“Phytochemicals” are chemical compounds found in plants, and the
term is often used to describe a large number of secondary
metabolites derived from the plants. Phytochemical screening assay is
a simple, quick, and inexpensive procedure that gives a quick answer
to the researcher about the presence or absence of various types of
phytochemicals as mentioned above in the given sample. It is an
important tool in bioactive compound analyses. After obtaining the
crude extract or active fraction from the plant material, phytochemical
screening test can be performed using the experimental procedures
given in Table 4.3 (Gul et al., 2017) to get an idea regarding the type
of phytochemicals present in the sample extract mixture or fraction.

Table 4.3. Preliminary phytochemical screening tests.

S. no. Phytochemicals Procedure Results

1 Anthraquinones Take 6 g of the crude drug powder sample in a
conical flask. Add 10 mL of benzene, and filter it after 10 min. Again,
add 10 mL of 10% ammonia solution to the filtrate, and shake
vigorously for 30 s. Appearance of pink, violet, or red color indicates
the presence of anthraquinones.
2 Tannins Add 10 mL of bromine water to the 0.5 g aqueous
extract. Decoloration of bromine water indicates the presence of
tannins.
3 Saponins Take a small quantity of aqueous crude extract in a
test tube. Add 5 mL of distilled water and mix thoroughly. Again add a
few drops of olive oil and again mix vigorously. Formation of foam
indicates the presence of saponins.
4 Flavonoids Shinoda test: Mix pieces of magnesium ribbon
and conc. HCl with aqueous crude extract. Alkaline reagent test: To
the aqueous crude extract, add 2 mL of 2.0% NaOH. When
concentrated yellow color appears, add a few drops of diluted acid.
Appearance of pink color after few minutes indicates the
presence of flavonoid. Disappearance of yellow color indicates the
presence of flavonoids.
5 Glycosides Liebermann’s test: Add 2 mL of acetic acid and
2 mL of chloroform to the aqueous crude extract, and allow the
mixture to cool. Then add conc. H2SO4. Keller–Kiliani test: Add 4 mL
of glacial acetic acid and 1 drop of 2% FeCl3 to the 10 mL aqueous
crude extract. Then add 1 mL of conc. H2SO4. Salkowski’s test: Add 2
mL conc. H2SO4 to the aqueous crude extract. Appearance of green
color indicates the presence of glycosides. Formation of a brown ring
between the layers indicates the presence of cardiac glycosides.
Formation of a reddish-brown color indicates the presence of
glycoside.
6 Terpenoids Add 2 mL of chloroform to 5 mL aqueous crude
extract. Evaporate the mixture using water bath and then boil with 3
mL conc. H2SO4. Appearance of gray color indicates the presence
of terpenoids.
7 Steroids Add 2 mL of chloroform and conc. H2SO4 to the 5 mL
aqueous crude extract. Appearance of red color in the lower
chloroform layer indicates the presence of steroids.
8 Phenols Take 20 mL of distilled water in a test tube, add few
powdered samples, boil it, and then filter the mixture. To the filtrate
add 3–4 drops of 0.1% ferric chloride. Appearance of brownish green
or blue color indicates the presence of phenols.
9 Alkaloids Add 1% HCl to the crude extract and warm it. After
filtering, add Mayer’s reagent (mercuric chloride + potassium iodide in
water) to the filtrate. Formation of yellow colored precipitate indicates
the presence of alkaloids.
10 Carbohydrates Molisch’s test: Dissolve the crude sample
powder in 5 mL of distilled water and filter. To this filtrate, add two
drops of alcoholic α´-naphthol solution in a test tube. Then using a
dropper along with side of test tube, disposed tubes and pour drop
wise conc. Sulfuric acid carefully. Formation of violet color at the
junction or interface of two liquids indicates the presence of
carbohydrates.
Biotechnology in Food Processing and Preservation: An Overview
Gargi Ghoshal, in Advances in Biotechnology for Food Industry, 2018

2.5 Essential Phytochemicals

Phytochemicals are precious for human nutrition. Indoles,
isothiocyanates, and sulforaphane from vegetables, such as broccoli,
alylic sulfides from onions and garlic and isoflavonoids from soybeans
are known as plant phytochemicals. These are present in high
concentration in raw foods but intensities are reduced during
processing and handling (Wang and Murphy, 1996). Enhanced
amount of phytochemicals in foods can resolve this difficulty. Two
genes IFS1/IFS2, encoding for isoflavone synthase in soybean are
revealed and expressed in Arabidopsis thaliana, to activate the
synthesis of isoflavonoid genistin (Jung et al., 2000).
Chemical Changes of Bioactive Phytochemicals during Thermal
Processing
Yancui Huang, … Indika Edirisinghe, in Reference Module in Food
Science, 2016

Defining Bioactive Phytochemicals

Phytochemical is a broad term meaning plant (phyto) chemical
referring to a wide variety of compounds that occur naturally in plants.
The term bioactive also has broad meaning. Guaadaoui et al. (2014)
defined bioactive compounds as those that have the ability to interact
with one or more component(s) of a living tissue presenting a wide
range of probable effects. Generally, phytochemicals have been
classified into six major categories based on their chemical structures
and characteristics. These categories include carbohydrate, lipids,
phenolics, terpenoids and alkaloids, and other nitrogen-containing
compounds (Figure 1; Harborne and Baxter, 1993; Campos-Vega and
Oomah, 2013). Within each category, further division based on
biogenesis or biosynthetic origin gives rise to different subcategories.

Sign in to download full-size image

Figure 1. Categorization of phytochemicals.

In recent years the term ‘phytochemical’ has been used to distinguish

plant chemicals that do not meet the classical definition of ‘essential
nutrients.’ Some phytochemicals produce activity in biological
systems, including humans; hence, the term ‘bioactive
phytochemicals.’ Liu (2013) has defined phytochemicals as bioactive
nonnutrient compounds in fruits, vegetables, grains, and other plant
foods that have been linked to reductions in the risk of major
noncommunicable chronic diseases. In this review, nonnutrient
compounds derived from plants that have biological activity in humans
are considered to be bioactive phytochemicals. These include
phenolic compounds, terpenoid compounds, and alkaloids. More than
5000 individual dietary phytochemicals have been identified in plant
foods (i.e., fruits, vegetables, whole grains, legumes, and nuts) with
varying content and composition. However, a large percentage of
these compounds still remain unknown with respect to chemical
structure and/or the biological role in humans (Liu, 2013).

Interactions between Chinese Nutraceuticals and Western Medicines

Noel Chan, … Evette Perez, in Nutraceuticals, 2016

Cytochrome P450
Phytochemicals are largely metabolized by the same enzymes that
metabolize food and drugs. First described and discovered in the
1950s by Julius Axelrod, cytochrome P450 (CYP450) are a set of
enzymes capable of promoting excretion of xenobiotics through
hydroxylation reactions using a heme molecule as a cofactor (Axelrod,
1955; Berg et al., 1976). Phytochemicals interact with CYP450 as
substances to be metabolized, and select phytochemicals are also
capable of inducing and/or inhibiting specific or multiple CYP
enzymes. Naturally, the abundance of specific CYP enzymes varies
by ethnicity and also by individuals. Therefore, effectiveness of
medications for different patients and interactions between
medications also vary across individuals. Ingestion of phytochemicals
from TCMs should be used with caution, especially when patients are
concurrently using western therapeutics because induction and/or
inhibition of enzymes can lead to a decreased or increased dose of
the phytochemical or western medication. Toxic effects are especially
of concern for medications or phytochemicals with a narrow
therapeutic index or high toxicity profiles, such as warfarin or ginkgo
nuts, respectively. As the usage of herbal compounds becomes more
prevalent, even in the United States, it is important for prescribers to
be aware of possible cytochrome interactions that common herbal
decoctions can cause.

Recommended publications

Biochemical Systematics and Ecology

Biochemical Systematics and Ecology
Journal
Industrial Crops and Products
Industrial Crops and Products
Journal
South African Journal of Botany
South African Journal of Botany
Journal
Scientia Horticulturae
Scientia Horticulturae
Journal
Featured Authors

Rauf, Abdur
University of Swabi, Swabi, Pakistan
Citations10,509h-index47Publications180
Iranshahi, Mehrdad
Mashhad University of Medical Sciences, Mashhad, Iran
Citations9,240h-index48Publications70
Ruberto, Giuseppe
Consiglio Nazionale delle Ricerche, Rome, Italy
Citations7,788h-index41Publications51
Lee, Young Chul
Gachon University, Seongnam, South Korea
Citations5,762h-index39Publications63
Elsevier logo with wordmark
About ScienceDirect
Remote access
Shopping cart
Advertise
Contact and support
Terms and conditions
Privacy policy
We use cookies to help provide and enhance our service and tailor
content and ads. By continuing you agree to the use of cookies.

All content on this site: Copyright © 2023 Elsevier B.V., its licensors,
and contributors. All rights are reserved, including those for text and
data mining, AI training, and similar technologies. For all open access
content, the Creative Commons licensing terms apply.
RELX group home page
Feedback

4 How does the choice of solvent impact the efficiency of

phytochemical extraction?

The choice of solvent has a significant impact on the efficiency of

phytochemical extraction. Different solvents possess different
solubility characteristics, polarity, and interactions with target
compounds, which can affect the extraction efficiency and selectivity.
Here are a few important considerations:

Solvent polarity: Polar solvents, such as water and ethanol, are

suitable for extracting polar compounds like phenolics and glycosides.
Conversely, nonpolar solvents like hexane or diethyl ether are more
effective for extracting nonpolar compounds like essential oils or lipids.
Intermediate polarity solvents like methanol or ethanol-water mixtures
can extract a broader range of compounds.

Solvent-solute interactions: Solvent-solute interactions play a crucial

role in extraction efficiency. The solvent should have sufficient affinity
for the target compounds to dissolve and extract them from the plant
matrix. For example, if the target compounds have strong hydrogen
bonding capabilities, a solvent like methanol or ethanol, which can
form hydrogen bonds, might be more effective.

Toxicity and regulatory considerations: Solvents used in extraction

should be safe for human consumption and comply with regulatory
standards. Toxic solvents like chloroform or benzene are generally
avoided, and preferences are given to generally recognized as safe
(GRAS) solvents or solvents approved for use in the pharmaceutical,
food, or cosmetic industries.

Environmental impact: Environmental considerations are important in

solvent choice. Ideally, solvents with low toxicity, low volatility, and low
environmental impacts such as biodegradability should be selected.
Water and ethanol are relatively environmentally friendly choices
compared to some organic solvents.

Process scalability and cost: Solvent choice can impact the scalability
and cost-effectiveness of the extraction process. Some solvents may
be more expensive or difficult to obtain in large quantities. Additionally,
solvent recovery and recycling processes should be considered to
minimize waste and reduce costs.

It is important to note that there is no universally optimal solvent for all

phytochemical extractions. The selection of a solvent depends on the
specific plant matrix, target compounds, intended applications, and
other process considerations. Experimental optimization of solvent
selection and optimization is often necessary to obtain the highest
extraction efficiency and desired phytochemical yield.