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David Belias, Chem 213

Synthetic #4, FFR
Experiment 21: Steam Distillation of a Spice
Introduction
The extraction of natural products for use in medicine and other industries has been
important since the beginning of recorded history. Some of the earliest civilizations chewed on
herbs to relieve pain and used leaves for wound healing. In ancient Greece, Hippocrates of Cos
collected over 400 natural products for use in treatment of disease. Traditional Chinese medicine
used 247 natural products, as well as 150 combinatorial drug formulas for disease treatment.
More recently, structural analysis and synthesis of natural compounds in organic chemistry has
led to improvement in the effectiveness of medicines. It is estimated that about 60% of all
available drugs today were derived from natural compounds. One theory explaining the
effectiveness of natural compounds as drugs is that the human body is able to metabolize the
natural products into effective drugs. Natural products may provide important clues for the
development of synergistic drugs that are able to deal with complex diseases, which is a vital
aspect of cancer research.
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One specific example of the use of natural products for disease treatment and drug
discovery is they synthesis of heterocycles. At Jiangsu Normal University in China, a three-
component reaction between an amine, an aromatic aldehyde, and tert-butyl 2,4-dioxopiperidine-
1-carboxylate was carried out to yield a tetracyclic heterocycle via multiple condensation
reactions. The specific name for the class of heterocycles produced is naphthyridines, which is
found in biologically active molecules. The main focus of these molecules is their use in the
treatment of cancer. They are also used as inhibitors that help to prevent diseases. Some
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examples of this are the first clinical stage inhibitor of protein kinase CK2 for treatment of
cancer, as well as DNA ligase inhibitors with anti-staphylococcal activity.
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Another method of obtaining useful compounds for treatment of disease is extraction
from natural products. Several important compounds exist as oils in natural products, so
extraction can be difficult sometimes. A useful method for the isolation of oils is steam
distillation, which is the co-distillation of the desired oil with water. High boiling substances,
such as oils, are difficult to distill alone because they would begin to decompose at very high
temperatures. Any liquid that is immiscible with water can be distilled near 100C. Each liquid
in the immiscible mixture contributes a partial pressure to the total vapor pressure. The mixture
boils when the vapor pressure is equal to atmospheric pressure, so any immiscible liquid in the
mixture causes water to boil at a lower temperature than normal. Water vapor is able to sweep
the desired oil out of the distillation mixture and carry it to the condenser.
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This technique was
very useful in this experiment for the isolation of spearmint oil.
Spearmint oil is an aromatic plant of economic importance that has numerous uses in
industry. It is used in fragrances, as well as in pharmaceutical applications. In the food industry,
spearmint is used as an antioxidant, a preservative, and also for enhancing overall nutritional
quality of food. Also, tests have concluded that spearmint has significant antioxidant and
antimicrobial properties. The antioxidant activity was determined to have an inhibition
percentage of 76%. Spearmint showed antimicrobial action against Staphylococcus, Salmonella,
and E. Coli, which may be due to the high phenolic concentration in its leaves.
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Spearmint is
composed of several different compounds, but the main one is carvone.
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Since carvone is the
main component of spearmint oil, many of the properties of spearmint are due to the presence of
carvone.
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Overall, spearmint oil and its constituents have important properties that are useful in the
medical and food industries. The purpose of this experiment was the isolation of spearmint oil
via steam distillation. The composition of spearmint oil was then determined based on analysis
of gas chromatography and gas chromatography-mass spectrometry results.
Experimental
Carvone. Dried spearmint leaves (4 g) were ground and added to a 100-mL round-bottom flask
with distilled water (50 mL). The mixture was stirred with a large stir bar and steam distilled for
two hours to produce a cloudy white distillate. The distillate was then washed with
dichloromethane (3 x 15 mL) and dried over anhydrous sodium sulfate. The remaining solvent
was evaporated to yield carvone, as clear oil (0.019 g, 0.475%); GC (phenyl methyl silicone,
40C to 275C at 10C per min) RT 13.51 min; GC-MS (phenyl methyl silicone, 40C to 275C
at 10C per min) RT 14.65 min, m/z 150.01.
Results and Discussion
This experiment dissolved the isolation of spearmint oil via steam distillation. The oil
was then analyzed by gas chromatography and gas chromatography-mass spectrometry to
determine the components of spearmint oil and confirm a successful isolation. After carrying out
the experiment, it was determined that spearmint oil was isolated and the main component was
carvone.
The gas chromatography and gas chromatography-mass spectrometry spectra suggest that
the isolated oil was spearmint that contained mainly carvone, with traces of other components.
First, gas chromatography (Figure 1) was run on the isolated oil, producing several peaks. The
tall peak at a retention time of 1.22 minutes most likely corresponds to dichloromethane, which
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was the solvent used. The next significant peak had a retention time of 12.75 minutes and an
area of 9,475. This most likely corresponds to a minor component of spearmint oil, possibly
limonene.
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It has the lowest retention time of the three major peaks due to a lower polarity,
which means a lower boiling point, so it will elute the quickest. The next major peak is at a
retention time of 13.32 minutes and has an area of 6,107. This peak also most likely represents a
minor component of spearmint oil, which could be 1,8-cineol.
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It would elute after limonene,
but before carvone because cineol contains an oxygen atom, increasing polarity of the molecule.
However, carvone contains a carbonyl group, which is slightly more polar than an ether group,
so carvone would elute after cineol. The final significant peak is at a retention time of 13.51
minutes and an area of 44,754, corresponding to carvone. Carvone has the highest boiling point
of these three compounds due to a higher polarity, so it would elute last and have the highest
retention time. There were several other small-intensity peaks that most likely correspond to
other components of spearmint since over 19 different compounds have been detected in
spearmint.
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From the gas chromatography spectrum, a percent composition of the spearmint oil was
calculated. The total area of the three significant peaks was 60,336, which means a percent
composition of 74.2% for carvone. This is expected because carvone is known to be the main
component of spearmint oil. The percent composition for limonene was 15.7% and 10.1% for
1,8-cineol. The two smaller peaks could be due to one of the nineteen other components of
spearmint, but based on the retention times, it was predicted that these two were the detected
compounds.
After analyzing the gas chromatography-mass spectrometry results (Figure 2), another
explanation for the gas chromatography results was possible. On the gas chromatography, only
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one peak was detected at a retention time of 14.65 minutes and an area of 3,499,364. When
looking at the library result from NIST, this retention time is consistent with 2-cyclohexen-1-
one, 2-methyl-5-(1-methylethenyl). This compound is also known as carvone, suggesting that
the correct product was isolated. Since there was only one peak on the spectrum, the percent
composition of carvone in the product was calculated to be 100%. This suggests a different
explanation for the results from the first gas chromatography. The gas chromatography result
could be due to the presence of air bubbles in the syringe. Air bubbles would cause the liquid to
be injected in multiple fractions, so the different retention times could all correspond to one
compound. All three retention times of the three peaks are very similar, so this could be an
explanation for the different results on the two spectra.
The mass spectrometry results also suggest that the compound in the spearmint oil is
carvone. The mass to charge value of the spectrum for the molecular ion fragment was 150.01,
which was very close to the literature molecular mass of carvone (150.22 g/mol).
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When looking
at the NIST library mass spectrometry data for carvone, the observed spectrum for the isolated
product was very similar. Overall, the gas chromatography and gas chromatography-mass
spectrometry data suggest the successful isolation of spearmint oil.


References
(1) Ji, H.; Li, X.; Zhang, H. Natural Products and Drug Discovery: Can Thousands of Years
of Ancient Medical Knowledge Lead Us to New and Powerful Drug Combinations in the
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Fight Against Cancer and Dementia? US National Library of Medicine. 2009, 10, 194-
200.
(2) Li, C.; Mu, X.; Li, Y.; Liu, Y.; Wang, X. A Combinatorial Synthesis of Fused Tetracyclic
Heterocycles Containing [1,6]Naphthyridine Derivatives under Catalyst Free Conditions.
ACS Combinatorial Science. 2013,
(3) Miles, W.H.; Rummel, S.A.; Smiley, P.M. Steam Distillation of a Spice. Journal of
Chemical Education. 2002, 79, 90.
(4) Dhifi, W.; Jelali, N.; Mnif, W.; Litaiem, M.; Hamdi, N. Chemical Composition of the
Essential Oil of Mentha Spicata L. From Tunisia and its Biological Activities. Journal of
Food Biochemistry. 2012, 37, 524-527.
(5) Hussain, A.; Anwar, F.; Shahid, M. Chemical Composition, and Antioxidant and
Antimicrobial Activities of Essential Oil of Spearmint (Mentha spicata L.) From
Pakistan. Journal of Essential Oil Research. 2010, 22, 78-84.
(6) (R)-()-Carvone; MSDS No. 124931 [Online]; Sigma Aldrich; St. Louis, MO, Nov 23,
2012. http://www.sigmaaldrich.com/MSDS/MSDS (accessed April 23, 2013).