2008

NPC Natural Product Communications Vol. 3

No. 5
829 - 832
Seasonal Variation in the Leaf Essential Oil Composition of
Sassafras albidum
Kristi M. Kaler and William N. Setzer*

Department of Chemistry, University of Alabama in Huntsville, Huntsville, Alabama 35899, USA

wsetzer@chemistry.uah.edu

Received: January 27th, 2008; Accepted: March 24th, 2008

The seasonal variation in the chemical composition of the leaf essential oil of Sassafras albidum has been analyzed by GC-MS.
Three individual trees were sampled four times during the course of the growing season. The leaf oils were made up of 44
components, with geranial (11%-27%) and neral (10%-18%) dominating. S. albidum showed a general trend of diminishing
monoterpenoid concentrations and increasing sesquiterpenoid concentrations during the season.

Keywords: Sassafras albidum, Lauraceae, seasonal variation, essential oil composition, GC-MS, geranial, neral, limonene,
(E)-caryophyllene, α-pinene.

Sassafras albidum (Nutt.) Nees is a medium-sized Table 1: Leaf essential oil yields of Sassafras albidum.
tree that ranges from central Florida, north to New Tree
Date Mass Fresh Mass Ess.
Collected/Extracted Leaves (g) Oil (mg)
England and the Great Lakes, and west to eastern 21-May-04 25.1 21.8
Texas and Oklahoma [1]. S. albidum, along with A
10-Jul-04 27.8 45.3
4-Sept-04 23.4 52.1
other members of the Lauraceae, Lindera benzoin, 30-Oct-04 12.6 7.3
Cinnamomum camphora, and Persea borbonia, is a 22-May-04 26.5 31.9
host plant of the spicebush swallowtail butterfly, 11-Jul-04 28.1 50.3
B
5-Sept-04 22.5 60.1
Papilio troilus [2]. In this paper, we present the 31-Oct-04 15.1 19.2
variation of the chemical composition of the leaf 23-May-04 24.8 19.5
12-Jul-04 27.9 27.0
essential oils of S. albidum growing in Huntsville, C
6-Sept-04 27.5 23.5
Alabama, during the growing season of 2004. To our 31-Oct-04 17.9 4.3
knowledge, this report is the first to present the
seasonality of sassafras leaf oil. eugenol, and citral [6]. The leaf essential oil
composition of S. albidum, as revealed in this current
The essential oil yields for the three different trees on study, differs somewhat from that reported previously
the four different dates of collection are summarized from a sample from Delaware [7]. Thus, the major
in Table 1. The essential oil yields show a gradual components in this work were geranial [= (E)-citral]
increase from May into September, but then a (ranges between 10.7% and 26.5%), neral [= (Z)-
dramatic decrease by the end of October. Similar citral] (9.9%-18.1%), limonene (5.7%-16.4%), (E)-
variation in essential oil yields have been previously caryophyllene (5.1%-12.5%), α-pinene (3.2%-
observed in a number of plants, including 12.2%), (Z)-3-hexenol (2.5%-9.9%), linalool (3.5%-
Micromeria fruticosa from Israel [3], and Thymus 6.7%), and caryophyllene oxide (up to 19.0%). The
hyemalis from Spain [4]. Delaware sample showed no geranial, neral,
caryophyllene oxide, or (Z)-3-hexenol. Previous
The chemical compositions of the S. albidum leaf oils investigations of S. albidum have shown the root bark
are summarized in Table 2. S. albidum leaves have oil to be composed largely of safrole (80-85%), with
been reported to contain α-pinene, α-phellandrene, smaller amounts of camphor (1-3%) and
myrcene, linalool, and geraniol [5], while extracts methyleugenol (1-13%) [7,8]. The leaf chemistry of
of leaf litter have revealed α-pinene, α-phellandrene, S. albidum is remarkably different from that of the
830 Natural Product Communications Vol. 3 (5) 2008 Kaler and Setzer

Table 2: Chemical composition of Sassafras albidum leaf essential oil.

Percent Composition
RIa Compound Tree A Tree B Tree C
5/21/04 7/10/04 9/4/04 10/30/04 5/22/04 7/11/04 9/5/04 10/31/04 5/23/04 7/12/04 9/6/04 10/31/04
859 (Z)-3-Hexenol 5.8 7.8 6.1 7.6 3.2 4.5 4.5 3.1 2.5 6.2 9.5 9.9
943 α-Pinene 12.2 8.5 4.6 5.7 3.7 3.4 3.2 3.3 4.9 8.7 6.6 8.1
955 Camphene 4.0 0.7 1.6 3.3 1.4 1.3 1.2 0.3 2.0 3.3 2.9 5.4
995 Myrcene traceb trace trace trace trace trace trace trace 0.8 trace trace trace
1026 p-Cymene 1.3 0.9 1.5 1.0 2.3 1.0 1.0 trace 3.8 trace 1.5 1.5
1030 Limonene 16.4 13.1 6.9 7.7 8.6 6.5 6.0 5.7 8.6 14.6 7.7 10.3
1040 (Z)-β-Ocimene trace trace 1.2 trace 0.8 0.6 1.1 trace 0.6 trace trace trace
1050 (E)-β-Ocimene trace trace 1.7 trace 1.6 1.0 0.8 trace 1.0 trace trace trace
1102 Linalool 3.5 5.6 5.3 3.7 5.1 5.7 5.6 4.4 5.4 6.7 6.2 4.2
1143 Camphor trace trace trace trace 0.3 0.3 trace trace trace trace 0.4 trace
1185 (E)-Isocitral --- trace trace trace 0.4 0.7 0.6 trace 0.6 trace trace trace
1191 α-Terpineol trace trace trace trace 0.6 0.5 0.3 trace 0.7 trace trace trace
1200 Estragole 1.7 2.4 3.3 trace 0.2 3.3 3.6 trace 5.1 1.2 0.6 trace
1218 trans-Carveol trace trace 0.8 trace 1.0 0.7 0.6 trace 1.3 trace trace trace
1229 Nerol trace trace 2.0 trace 1.5 2.2 2.5 1.0 1.0 1.0 1.3 trace
1241 Neral 15.7 18.1 13.4 12.8 14.0 17.4 13.4 16.8 9.9 17.2 14.8 10.3
1255 Linalool acetate trace trace 0.8 trace 1.2 1.0 1.2 trace 0.7 trace trace trace
1270 Geranial 22.3 26.5 23.6 14.8 22.3 25.4 18.9 17.8 21.7 22.2 18.4 10.7
1337 δ-Elemene --- --- --- --- 0.4 --- --- --- 0.4 --- --- ---
1391 β-Elemene trace trace 0.5 0.7 0.4 0.4 0.8 0.8 0.3 trace 0.6 trace
1410 (Z)-Caryophyllene --- --- --- --- 0.4 trace trace --- 0.6 --- --- ---
1418 (E)-Caryophyllene 8.9 9.2 8.5 5.7 12.5 8.3 9.5 8.7 6.4 9.3 7.8 5.1
1449 C15H24 trace trace trace trace 0.7 trace 0.6 0.7 0.4 trace trace trace
1452 α-Humulene 0.9 1.2 1.0 trace 1.8 1.5 1.2 0.9 1.1 1.0 1.4 trace
1475 γ-Gurjunene 1.7 2.0 2.4 2.1 1.8 1.9 2.9 3.2 1.3 1.2 1.8 1.5
1484 β-Selinene 2.1 1.7 1.9 2.7 1.9 1.9 2.3 3.8 1.7 1.1 1.5 2.4
1490 Unknown trace trace 1.1 trace 0.6 0.8 1.4 1.4 trace trace 1.0 trace
1493 α-Selinene 1.3 1.3 1.7 1.9 1.5 1.6 2.1 2.9 1.2 0.8 1.3 1.4
1497 α-Zingiberene trace trace 1.5 trace 2.0 1.4 1.6 trace 1.5 trace 1.5 trace
1510 (E,E)-α-Farnesene trace trace 0.6 8.2 0.7 0.4 0.5 7.6 0.5 trace trace 9.3
1513 C15H24O trace trace 2.2 --- 1.5 2.0 3.3 trace 4.5 1.2 1.9 ---
1523 δ-Cadinene trace trace 0.4 trace 0.8 0.5 0.5 trace 0.7 trace trace trace
1545 Unknown trace --- --- --- 0.9 trace trace trace 0.5 --- --- ---
1564 (E)-Nerolidol trace trace 1.7 1.1 2.6 2.1 2.5 trace 4.0 1.0 1.4 trace
1579 Caryophyllene oxide 1.7 1.1 1.0 11.9 0.4 0.7 1.1 15.8 trace 2.2 4.9 19.0
1604 Humulene epoxide II --- --- --- trace --- --- --- trace --- --- trace 0.9
1623 C15H26O trace trace 1.0 0.6 0.5 0.5 1.1 0.7 0.7 trace 0.7 trace
1628 C15H24O trace trace 1.1 0.3 trace trace 0.7 trace 1.5 trace 0.8 trace
1634 C15H24O trace trace trace 1.6 trace trace trace trace trace trace trace trace
1652 Kongol 0.6 trace 0.7 1.1 0.5 0.6 0.9 1.0 1.0 trace 0.5 trace
1656 C15H24O --- trace trace 3.2 trace trace 2.4 trace trace 1.1 3.2 trace
1670 C15H24O --- --- --- --- --- --- trace trace 1.0 --- --- ---
1715 Pentadecanal --- --- --- 2.3 --- --- --- trace --- --- --- trace
Total identified 100.0 100.0 94.7 94.2 95.8 96.8 90.5 97.2 91.4 97.7 92.5 100.0
Fatty acid derivatives 5.8 7.8 6.1 9.8 3.2 4.5 4.5 3.1 2.5 6.2 9.5 9.9
Monoterpene hydrocarbons 33.8 23.1 17.4 17.8 18.4 13.7 13.3 9.3 21.6 26.6 18.7 25.4
Oxygenated monoterpenoids 41.5 50.1 45.9 31.3 46.5 54.0 43.2 40.1 41.3 47.0 41.1 25.2
Sesquiterpene hydrocarbons 14.8 15.5 18.5 21.3 24.7 17.9 22.1 28.6 16.1 13.5 15.7 19.6
Oxygenated sesquiterpenoids 2.3 1.1 7.7 19.8 5.5 5.8 12.0 17.5 12.8 5.5 13.4 19.9
Phenylpropanoids 1.7 2.4 3.3 trace 0.2 3.3 3.6 trace 5.1 1.2 0.6 trace
Unidentified trace trace 1.1 trace 1.5 0.8 1.4 1.4 0.5 trace 1.0 trace
a
RI = “Retention Index” based on a homologous series of normal alkanes.
d
trace, <0.1%

root; in this present study, we find no trace of safrole,
while neither geranial nor neral has been found in the
root oil.
Although citral and geranial are not typically found
in leaf oils of the Lauraceae, they are both abundant
in the leaf essential oil of Cinnamomum porrectum
[9] and neral is abundant in C. osmophloeum leaf
oil [10]. Limonene and α-pinene are found in many
members of the Lauraceae, and (E)-caryophyllene
seems to be virtually omnipresent in leaf oils
of the family [11]. Interestingly, α-copaene and
germacrene D, which are generally common to
abundant components of leaf oils of the Lauraceae,
were not detected in S. albidum from Alabama;
germacrene D, however, was found in the Delaware
sample [7], and α-humulene was found in both.
Seasonal variation of sassafras leaf oil Natural Product Communications Vol. 3 (5) 2008 831

Figure 2: Seasonal variation in geranial + citral concentrations in

Sassafras albidum leaf oils.
Figure 1: Average total monoterpenoid and sesquiterpenoid
concentrations in Sassafras albidum leaf oils.
185 m elevation), were collected at four different
times of the year (mid May, mid July, early
The most notable changes in leaf oil chemical
September, and late October) during 2004. The
composition during the season were the decrease in
leaves were collected in the morning (approximately
total monoterpenoid concentration with concomitant
8:00 am), immediately chopped, and hydrodistilled
increase in sesquiterpenoid concentration (Figure 1).
for 4 h using a Likens-Nickerson hydrodistillation-
This phenomenon has been observed in tropical
extraction apparatus. Essential oil yields are
Virola surinamensis from Brazil during the transition
summarized in Table 1.
from dry season to wet season [12]. In addition, the
sum of geranial and neral concentrations showed a
GC-MS analysis: The leaf oils of S. albidum were
rise in July and then a steady drop later in the season
subjected to GC-MS analysis using an Agilent 6890
(Figure 2). The increase in geranial and neral may be
GC with Agilent 5973 mass selective detector, fused
in response (albeit unsuccessful) to herbivory by
silica capillary column (HP-5ms, 30 m x 0.25 mm),
Japanese beetles (Popillia japonica) that heavily
helium carrier gas, 1.0 mL/min flow rate; inj temp
infested the trees during July, but not during the other
200°C, oven temp prog: 40°C initial temperature,
collection periods. Geranial has been shown to be
held for 10 min; increased at 3°/min to 200°C;
the repellent agent in lemongrass (Cymbopogon
increased 2°/min to 220°C, and interface temp
citratus) oil [13]. Interestingly, the concentration of
280°C; EIMS, electron energy, 70 eV. The sample
linalool, also observed to show insect repellent
was dissolved in CHCl3 to give a 1% w/v solution;
activity [14], increased during the Japanese beetle
1-μL injections using a splitless injection technique
infestation. There was a dramatic increase in the
were used. Identification of oil components was
concentration of caryophyllene oxide in October,
achieved based on their retention indices (determined
reflected in a small decrease in (E)-caryophyllene
with reference to a homologous series of normal
concentration.
alkanes), and by comparison of their mass spectral
fragmentation patterns with those reported in the
Experimental
literature [15] and stored on the MS library [NIST
Plant material: A sampling of mature leaves from database (G1036A, revision D.01.00)/ChemStation
different locations on three different trees data system (G1701CA, version C.00.01.08)]. The
growing on the campus of the University of Alabama chemical compositions of the S. albidum leaf oils are
in Huntsville, Alabama (34o 43.32' N, 86o 38.30' W, summarized in Table 2.

References
[1] (a) Griggs MM. (1990) Sassafras albidum (Nutt.) Nees sassafras. In Silvics of North America, Vol. 2, Hardwoods, Agriculture
Handbook 654, Burns RM, Honkala BH (Eds). U.S. Department of Agriculture, Forest Service, Washington, DC. 773-777; (b)
Maisenhelder LC. (1965) Sassafras (Sassafras albidum (Nutt.) Nees. In Silvics of Forest Trees of the United States, Agriculture
Handbook 271, Fowells HA (Ed). U.S. Department of Agriculture, Washington, DC. 654-655.
[2] (a) Nitao JK, Ayres MP, Lederhouse RC, Scriber JM. (1991) Larval adaptation to lauraceous hosts: geographic divergence in the
spicebush swallowtail butterfly. Ecology, 72, 1428-1435; (b) Lederhouse RC, Ayres MP, Nitao JK, Scriber JM. (1992) Differential
use of lauraceous hosts by swallowtail butterflies, Papilio troilus and P. palamedes (Papilionidae). OIKOS, 63, 244-252.
832 Natural Product Communications Vol. 3 (5) 2008 Kaler and Setzer

[3] Dudai N, Larkov O, Ravid U, Putievsky E, Lewinsohn E. (2001) Developmental control of monoterpene content and composition
in Micromeria fruticosa (L.) Druce. Annals of Botany, 88, 349-354.
[4] Jordán MJ, Martínez RM, Goodner KL, Baldwin EA, Sotomayor JA. (2006) Seasonal variation of Thymus hyemalis Lange and
Spanish Thymus vulgaris L. essential oils composition. Industrial Crops and Products, 24, 253-263.
[5] Kleber C. (1899) The chemistry of sassafras. Americal Journal of Pharmacy, 71, 27-32.
[6] Gant RE, Clebsch EEC. (1975) The allelopathic influences of Sassafras albidum in old-field succession in Tennessee. Ecology, 56,
604-615.
[7] Tucker AO, Maciarello MJ, Broderick CE. (1994) Filé and the essential oils of the leaves, twigs, and commercial root teas of
Sassafras albidum (Nutt.) Nees (Lauraceae). In Spices, Herbs, and Edible Fungi, Charalambous G (Ed). Elsevier, Amsterdam.
[8] (a) Kamdem DP, Gage DA. (1995) Chemical composition of essential oil from the root bark of Sassafras albidum. Planta Medica,
61, 574-575; (b) Simic A, Sokovic MD, Ristic M, Grujic-Jovanovic S, Vukojevic J, Marin PD. (2004) The chemical composition of
some Lauraceae essential oils and their antifungal activities. Phytotherapy Research, 18, 713-717.
[9] Zhu LF, Lu BY, Li YJ. (1984) Studies on chemical constituents of essential oil from leaves of jiang-zhang (C. porrectum). Acta
Botanica Sinica, 26, 638-643.
[10] Chang ST, Chen PF, Chang SC. (2001) Antibacterial activity of leaf essential oils and their constituents from Cinnamomum
osmophloeum. Journal of Ethnopharmacology, 77, 123-127.
[11] (a) Setzer WN, Haber WA. (2007) Leaf essential oil composition of five species of Beilschmiedia from Monteverde, Costa Rica.
Natural Product Communications, 2, 79-83; (b) Takaku S, Haber WA, Setzer WN. (2007) Leaf essential oil composition of ten
species of Ocotea (Lauraceae) from Monteverde, Costa Rica. Biochemical Systematics & Ecology, 35, 525-532; (c) Setzer WN,
Stokes SL, Penton AF, Takaku S, Haber WA, Hansell E, Caffrey CR, McKerrow JH. (2007) Cruzain inhibitory activity of leaf oils
of neotropical Lauraceae and essential oil components. Natural Product Communications, 2, 1203-1210.
[12] Lopes NP, Kato MJ, Andrade EhdA, Maia JGS, Yoshida M. (1997) Circadian and seasonal variation in the essential oil from Virola
surinamensis leaves. Phytochemistry, 46, 689-693.
[13] Leal WS, Uchida K. (1998) Application of GC-EAD to the determination of mosquito repellents derived from a plant, Cymbopogon
citratus. Journal of Asia Pacific Entomology, 1, 217–221.
[14] (a) Chapman RF, Bernays EA, Simpson SJ. (1981) Attraction and repulsion of the aphid Cavariella aegopodii by plant odors.
Journal of Chemical Ecology, 7, 881-888; (b) Cook SM, Jönsson M, Skellern MP, Murray DA, Anderson P, Powell W. (2007)
Responses of Phradis parasitoids to volatiles of lavender, Lavendula angustifolia – a possible repellent for their host, Meligethes
aeneus. BioControl, 52, 591-598.
[15] Adams RP. (2007) Identification of Essential Oil Components by Gas Chromatography/Mass Spectrometry, 4th Ed. Allured
Publishing, Carol Stream, Illinois.