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MEDICINAL

CHEMISTRY
RESEARCH

Med Chem Res (2010) 19:1092–1105
DOI 10.1007/s00044-009-9255-z
ORIGINAL RESEARCH

Extraction of essential oils from garlic (Allium sativum)
using ligarine as solvent and its immunity activity
in gastric cancer rat
Rui Li • Wei-chang Chen • Wei-peng Wang
Wen-yan Tian • Xue-guang Zhang

Received: 9 May 2008 / Accepted: 2 September 2009 / Published online: 24 December 2009
Ó Birkha¨user Boston 2009

Abstract The solvent extraction (SE) of garlic essential oil (Allium sativum) was
studied. A multivariate study based on a four-factor, three-level Box–Behnken
design (BBD) was used to evaluate the influence of four major variables affecting
the performance of the SE of garlic essential oil. The yield and the composition of
the essential oils from garlic obtained by SE were determined, and compared with
those obtained by the supercritical fluid extraction (SFE). Statistical treatment of the
results provided by the BBD revealed that the selected parameters, extraction time
and extraction temperature, were significant. The essential oils were analyzed by gas
chromatography–mass spectrometry (GC–MS). Major essential oil components
were 3-vinyl-4H-1,2-dithiin (31.89%), diallyl trisulfide (13.31%), diallyl sulfide
(2.22%), dially disulfide (6.87%), propyl allyl disulfide (13.89%), and dimethyl
disulfide (7.05%). Compared with SFE, the yield of essential oil obtained by SE was
slightly lower, but major essential oil components were quantitatively similar.
Moreover, residual solvent in garlic essential oil was very low (\50 mg/kg). In
addition, a significant increment in the ratio of CD4?/CD8? in serum of gastric
cancer rats was determined after administration of garlic essential oil. It can be

R. Li  W. Chen (&)  W. Tian
Department of Gastroenterology, The First Affiliated Hospital of Soochow University,
Suzhou, China
e-mail: chenwc_prof@yahoo.cn
R. Li  W. Chen  W. Tian  X. Zhang
Key Laboratory of Medicine and Clinical Immunology of Jiangsu Province, Suzhou 215006, China
W. Wang
College of Pharmacy, Soochow University, Suzhou 215007, China
X. Zhang
Institute of Medical Biotechnology, Soochow University, Suzhou 215007, China

Although people conventionally think that low yield. and hence platelet aggregation. 2007. and dimethyl disulfide.. 2009.. especially in the Nordic countries Sweden and Norway. 2004).. 2007). gastric cancer. diallyl trisulfide. 2007). loss of volatile compounds. In this article.. Chyau et al. The major components of garlic essential oil are 3-vinyl-4H-1. The essential oils of the native spices have been isolated using steam distillation and/or SE (Xu et al... Bordia et al.. Shenoy and Choughuley (1992) further indicated that garlic (Allium sativum L.Med Chem Res (2010) 19:1092–1105 1093 concluded. We still .. Chemical compositions of the SFE extract and essential oil obtained by SE were analyzed by GC and GC/MS methods. the results indicated that garlic essential oil may be useful for treatment of patients with inflammatory disease. degradation of unsaturated compounds. diallyl sulfide. dially disulfide. Eidi et al. (1998) reported that diallyl disulfide and diallyl trisulfide. Keywords Box–Behnken design  Garlic  Essential oils  Extraction  GC–MS  Supercritical fluid extraction  CD4?/CD8?  Gastric cancer  CD40 Introduction Garlic (Allium sativum) is widely used for flavoring purposes in food. SFE method also has some disadvantages that prohibit its industrial uses because of high equipment investment and small extraction scale. A four-factor. Khajeh et al. Moreover. Garlic (Allium sativum) has played an important dietary as well as medicinal role for centuries (Chauhan 2006. Quynh et al. propyl allyl disulfide. the two major organosulfur compounds derived from garlic essential oil. thereby preserving the original extract composition and properties. 2006. The yields and the composition of each sample of essential oils resulting from the experiments of the BBD have been analyzed and compared. Franco et al. that the SE method offers obvious advantages over SFE. 2008) since it has certain advantages over steam distillation and solvent extraction (SE). and in Asia. there have been a few successful reports using such techniques (Cho et al. toxic solvent residue. the SE method offers obvious advantages over SFE. This study still describes a comparative study of classical SE and SFE of garlic essential oils.g. Khajeh et al. and production of undesirable off-flavors (due to heat) are often involved in using such techniques. threelevel Box–Behnken design (BBD) has been developed to specify the importance of the four major factors affecting the SE. Supercritical fluid extraction (SFE) of active compounds from plant material is a promising field for the industrial application of SFE (Reverchon 1997. Kitzberger et al. e. and suggested that diallyl sulfur compounds in garlic essential oil might also act as nitrite scavengers.. 2004. could inhibit platelet thromboxane formation. Unfortunately..) essential oil were effective in reducing the chemical formation of N-nitrosamines (NAs). the SE of garlic essential oil has been studied to understand in depth and to optimize this classical technique of extraction. To date..2-dithiin. SFE can be performed at lower temperatures. the United Kingdom. 2007. long extraction times.

and CD8? and the ratio of CD4?/CD8? in gastric cancer rats. 5) of organic solvent (ethanol. 50. 2003) was applied to determine the best combination of extraction variable parameters for the production of essential oil from garlic. ligarine. with both coded values and natural values investigated in this study. 35. 25°C). Experimental design A four-factor. Extraction of essential oil The ground garlic samples (50 g) were weighed and quantitatively transferred into a 500-ml flask. where xi is the coded value of an independent variable. 50 g of garlic was blended with 20-ml deionized water for 2 min in a commercial blender. 40. or n-pentane) for a given time (10. 3.1094 Med Chem Res (2010) 19:1092–1105 investigated the role of garlic essential oil in inflammation and immune response by measuring the mean (?SE) percentage of CD3?. Materials and methods Materials Garlic was purchased from local market (Suzhou City. CD4?. 30. ethyl acetate. 60. In each batch. After extraction. and 50°C. 45. The combined filtrate was concentrated using rotary vacuum evaporator at 40°C and weighed on an analytical balance to give the extraction rate of essential oil. Table 2 lists the actual experimental parameters corresponding to the . and ethyl acetate. respectively set at 20. The variables Xi were coded as xi according to the following Equation: xi ¼ ðXi  Xi=2Þ=DXi. During the experiment. three-level Box–Behnken design (Yin and Dang 2008. 30. then extracted with different volume (1. 25. are represented in Table 1. 2. The variables and their levels. Xi is the real value of an independent variable. The water layer was extracted with another equal volume of the solvent. the organic layer was filtered and collected. Muthukumar et al. all the samples were maintained in our lab at appropriate conditions (dark. The remaining solvent in the essential oil extract was determined by GC analysis. The factorial design consisted of three center points and six BBD Blocks. n-pentane.. Xi/2 is the real value of an independent variable at the central point. China). and 70 min) when extraction temperatures were. 40. 4. Four different solvents were used for the distillations and the extractions of garlic samples: pure ethanol. The extraction procedure was repeated several times (1–5). ligarine. leading to 27 sets of experiments. 20. and DXi is the step change value.

46 10 -1 0 0 1 0.73 13 0 -1 -1 0 0. .67 27 0 0 0 0 0.34 18 -1 0 1 0 0.47 14 0 -1 1 0 0.71 17 -1 0 -1 0 0.63 19 1 0 -1 0 0.69 9 -1 0 0 -1 0.53 3 1 -1 0 0 0. Y.79 21 0 -1 0 -1 0. Each experiment was performed in duplicate and the average of extraction yield of essential oil was taken as the response.65 25 0 0 0 0 0.67 26 0 0 0 0 0.72 0.59 23 0 1 0 -1 0.38 2 -1 1 0 0 0.53 22 0 -1 0 1 0.Med Chem Res (2010) 19:1092–1105 Table 1 The level of variables chosen for the Box–Behnken design 1095 Variable Coded levels -1 0 ?1 Extraction temperature X1 (°C) 35 40 45 Extraction time X2 (min) 30 40 50 Ratio of liquid to raw material X3 2 3 4 Extraction number X4 3 4 5 Natural levels Table 2 Box–Behnken design with coded values for four extraction parameters and results Run X1 X2 X3 X4 Y 1 -1 -1 0 0 0.67 designed levels that were carried out for developing the model.66 20 1 0 1 0 0.59 7 0 0 1 -1 0.46 5 0 0 -1 -1 6 0 0 -1 1 0.62 24 0 1 0 1 0.51 11 1 0 0 -1 0.67 8 0 0 1 1 0.57 4 1 1 0 0 0.68 15 0 1 -1 0 0.65 12 1 0 0 1 0.55 16 0 1 1 0 0.

Column temperature was initially kept at 50°C for 3 min. The supercritical CO2 with dissolved compounds was passed through a heated micrometer valve. passed through a coil immersed in a water bath at a temperature above the supercritical one.. the extractor was charged with about 50 g of ground garlic. film thickness 0. GC–MS) analysis The essential oil was analyzed using a Hewlett Packard 5890 II GC equipped with a FID detector and HP-5ms capillary column (30 m 9 0.0 (SAS Institute.. The quality of the fit of the polynomial model equation was expressed by the coefficient of determination R2. (iii) equilibrium time: 5 min. Supercritical fluid extraction of garlic essential oil Supercritical fluid extraction of garlic essential oil was performed according to the method described by Gouveia et al. . France). and the total volume of gas was measured with a wet test meter. The experiment was carried on a HP 7680 A supercritical fluid Extraction Unit (Hewlett Packard. For each extraction test. Les Ulis. (iv) experiment temperature: 35°C.25 lm). and subsequently expanded to ambient pressure. (v) trap temperature: 0°C. The liquid CO2 from a cylinder was compressed to the working pressure at the temperature of a water bath. and its statistical significance was checked by an F-test. before reaching the extractor. and the fluid. Quantitative data were obtained electronically from FID area percent data without the use of correction factors. Gas chromatography. For GC/MS detection. and the solutes were collected in cooled glass U-tubes filled with glass wool. (2007) and Wang et al. an electron ionization system with ionization energy of 70 eV was used. The experimental conditions were : (i) density of CO2: 0. (ii) flow of CO2: 1 ml/min.1096 Med Chem Res (2010) 19:1092–1105 The responses obtained from each set of experimental design (Table 2) were subjected to multiple nonlinear regression using the software SAS 8. and (vii) extraction pressures: 10 MPa.6 g/ml. respectively. (2006). gas chromatography–mass spectrometry (GC. Tentative identification of the compounds was based on the comparison of their mass spectra with those of NIST 98 and Wiley 275 library data of the GC/MS system. Helium was the carrier gas. Gas flow rate was monitored with a rotameter. Samples of 1 ll were injected manually and in the splitless mode. Inc) to obtain the coefficients of the second polynomial model. The oil weight was measured by precision balance until no oil was extracted out from the ground garlic. (vi) extraction time: 130 min. The significances of the regression coefficient were tested by a t-test.25 mm. then gradually increased to 240°C with a rate of 3°C/min. The pressure was controlled by a back-pressure regulator. at a flow rate of 1 ml/min. Injector and detector temperatures were set at 220 and 290°C. The extraction column was packed with powdered raw materials and glass beads.

Four months after essential oil treatment. . The unbound cells are washed using a manual washing manifold connected to an electric vacuum pump while keeping the plate on a magnetic frame. respectively. Rats in group I orally received an equal amount of vehicle.Med Chem Res (2010) 19:1092–1105 1097 Animals and treatment Male Wistar rats (245 ± 10 g) were housed in steel cages. respectively. The T cell subsets are estimated using peroxidase labeled monoclonal antibodies specific to CD4 and CD8 cells. USA) at a wavelength of 450 nm with a reference wavelength of 630 nm. (France) was used to measure CD4 and CD8 counts. Rat gastric cancer model induced by 1-methyl-3nitro-1-nitrosoguanidine was performed in all animals. The calibrated reader provided the numbers of cells corresponding to the OD values. A few modifications were introduced such as manual rotation for 18 min during the 20-min incubation after the addition of the conjugate and gentle manual rotation of the plate for 15 min after the addition of substrate. The data presented in this report are of tests done within 2 h of collection of blood samples. Four cell standards and a blank each for CD4 and CD8 were included as mentioned in the protocol for the validation and interpretation of the results. Whenever a given standard was not close to the expected value. rats were euthanized and bled from the abdominal aorta. The Capcellia CD4/CD8 whole blood kit from Sanofi DIAGNOSTICS Pasteur. that particular standard was omitted and reading was taken by feeding the other three standard values and the blank. All the animal experiments were performed under the guidelines of the Laboratory Animal Experiment Committee of China. This is an immuno-capture ELISA-based assay in which paramagnetic particles coated with anti-pan-T cell antibodies capture T-lymphocytes. Rats were allotted into three groups: normal rats (group I) and essential oil-fed rats (groups II and III). Tetra methyl benzidine (TMB) is the substrate used in this system. all the animals were sacrificed. The readings were taken using a ELx800 Automated Microplate ELISA reader (Bio-Tek. After 4 weeks. Rats of groups II and III orally received essential oil at a single dose of 50 mg/kg and 100 mg/kg. CD4/CD8 cell estimation Two milliliters of blood was drawn from these individuals between 08:00 and 10:00 h into 5-ml collection tubes containing 60 ll of 5% EDTA. Plasma were stored at -70°C until use. They were allowed free access to drinking water and normal diet during the experiment. Blood samples were collected in heparinized tubes and centrifuged (4°C) at 15009g for 15 min. Though the kit manufacturer recommended each sample to be tested as singletons. The cell counts specified for each of the four standards and the blank (considered to have ‘‘0’’ cells) as recommended by the kit manufacturer were fed to the reader prior to taking reading thus enabling the software to draw a line of best fit. we tested all the samples in duplicate wells. The rats were acclimatized for 7 days and randomly allotted into four groups.

Effects of the ratio of solvent to raw material on extraction yield of essential oil The effect of the ratio of solvent to raw material on extraction yield of essential oil is shown in Fig. Therefore. optimum extraction temperature was chosen as 45°C. 35. As expected.74. This increase results from the decrease of the concentration of the extract Table 3 Effect of different solvents on extraction yield of essential oil Ethanol Extraction yield (%) Residual solvent (mg/kg) 0. 1c. There existed maximum extraction yield of essential oil when extraction time was 60 min. 1b. Therefore.78% of total oil at extraction temperature of 20. 30. respectively. and 50°C. Extraction time of 60 min is recommended based on the experimental results. yields increased with the increase in the ratio of solvent to raw material at a given condition. Results and discussion Effect of different solvents on extraction yield of essential oil Table 3 showed that maximum extraction yield of garlic essential oil was. 0.78.75% (ligarine). 0. This indicated that the increase in extraction temperature. 25. 0. and 0. in general. Results showed that yield of the extract was the highest when ligarine was used as a solvent.69% (ethyl acetate). we used ligarine as the solvent to extract garlic essential oil in this work. Moreover. 0. 0. The average essential oil yields from garlic in 50 min were 0. 2009).69.71 53 n-Pentane 0. in fact due to the increase of oil solubility. within the range investigated. 45..75 35 Ethyl acetate 0.71% (ethanol).69 57 . Effect of extraction time on extraction yield of essential oil The effect of extraction time on extraction yield of essential oil was shown in Fig. 0. the quality of the essential oil extract was the best. The extraction yield of essential oil increased with the increase of extraction time in the range of 10–60 min.34.52. and 0. increased the mass transfer coefficient resulting in higher extraction rates from garlic.43. Effect of extraction temperature on extraction yield of essential oil Increasing the extraction temperature increased the essential oil extraction (Fig. 0.1098 Med Chem Res (2010) 19:1092–1105 CD40 analysis CD40 expression rate was measured according to our past method (Li et al.76% (n-pentane).76 40 Ligarine 0. in turn. 0. 40. 1a).

Optimization of extraction conditions A three-factor three-coded level Box–Behnken design was used to determine the response value (Y) for different extraction parameters. the extraction yield of essential oil rapidly increased up to 0. 1d. Independent variables and their levels for the Box–Behnken design used in this study are shown in Table 1. the recommended ratio of solvent to raw material is between 3 and 5. 1c. Maximum extraction yield of essential oil (0. Using the relationships in Table 1.79%) was recorded under the experimental conditions of extraction . extraction time (b). and extraction number (X4) were independent variables studied to predict the response (Y). the actual levels of the variables for each of the experiments in the design matrix were calculated. The extraction yield of essential oil rapidly increased up to 0. and then no longer increased. extraction time (X2). As is shown in Fig.Med Chem Res (2010) 19:1092–1105 A C 1099 B D Fig. ratio of solvent to raw material (c) extraction number (d) extraction yield of essential oil solution. and the experimental results obtained are as given in Table 2. Effects of number of extractions on extraction yield of essential oil The effects of varying extraction number over a range of 1–5 on the production of essential oil were shown in Fig. Table 2 shows the experimental conditions and the results of essential oil production according to the factorial design. Extraction temperature (X1). ratio of solvent to raw material (X3). 1 Effect of extraction temperature on (a). leading to the increase of its solvent power.78% when the ratio of solvent to raw material varied from 1 to 4. This indicates that essential oil in garlic sample has been completely extracted when extraction number is 4. Therefore.77% when extraction number varied from 1 to 4 and then no longer increased.

1100 Med Chem Res (2010) 19:1092–1105 temperature 45°C.59963 R-square 96. 20. The figures show good results both near center of extraction Table 4 Fit statistics for Y Master model Mean 0.05).52% 96.59963 Predictive model 0.001) for the essential oil production were extraction temperature and ratio of solvent to raw material. ratio of solvent to raw material 4. The value of the adjusted determination coefficient (Adj. and the coefficients of the model were evaluated for significance with the Student t-test.054935 . Effect of variables on response value (Y) In order to gain a better understanding of the results. 2a–f. and Fig. and extraction number) except the ratio of solvent to raw material were significant (P \ 0.05%) indicated a better precision and reliability of the experiments carried out. respectively.5% of the total variations were not explained by the model. The four linear coefficients. the final predictive equation obtained is as given below: Y ¼ 0:67 þ 0:105833 X1 þ 0:046667 X2 þ 0:091667 X3 þ 0:030833 X4  0:054583 X1 X1  0:04 X1 X3  0:050833 X2 X2  0:032083 X4 X4 Statistical analysis revealed that the most relevant variables (P \ 0. In this case. the predicted models with one variable kept at central level and the other two varying within the experimental ranges are presented in Fig. a relatively lower value of the coefficient of variation (CV = 5.47% 92. The goodness of fit of the model was checked by determination coefficient (R2). 2a–f as the 3D response surface plots and Fig.52% Adj.965) indicated that only 3. The predicted maximum extraction yield (0.030311 CV 5. extraction time 40 min. extraction time.782%) derived from RSM regression was obtained when extraction temperature is 45°C.05). and extraction number 4. Neglecting the insignificant terms. R2 = 0.925) was also very high to advocate for a high significance of the model. The obvious trough in the response surfaces indicates that the optimal conditions were exactly located inside the design boundary (Fig. R-square 92. ratio of solvent to raw material 4. the value of the determination coefficient (R2 = 0. All the cross-product coefficients except X1X3 were eliminated in the refined equation as the P values of these coefficients are very insignificant (P [ 0. extraction time 42 min.47% RMSE 0. Multiple regression analysis was performed on the experimental data. and extraction number 4 in the test set No.030311 0.054935 5. and three quadratic terms (extraction temperature. The analysis of fit Statistics for Y is shown in Table 4. Figures 2a and 3a show the effect of extraction temperature and extraction time on extraction yield. At the same time. 3a–f as the 2D contour plots for extraction yield. 3a–f).

That is. The chosen conditions for extraction temperature. ratio of solvent to raw material. along with the predicted and measured results. and extraction number are all listed in Table 5. a slight change in extraction temperature or in ratio of solvent to raw material could lead to a greater extraction yield. In the contour plots for extraction yield. d X2–X3. extraction time. According to the principle. c X1–X4. 2c and 3c show the effect of extraction temperature and extraction number on extraction yield. and f X3–X4 .Med Chem Res (2010) 19:1092–1105 1101 time and high extraction temperature level studied. The figures show good results both near center of extraction number and high ratio of solvent to raw material level studied. As shown in Table 5. Confirmation experiments In order to confirm the validity of the statistical experimental strategies. one run of additional confirmation experiments was conducted. extraction temperature and ratio of solvent to raw material were two significant factors on extraction yield. b X1–X3. 2 Surface graphs of extraction yield showing the effect of variables: a X1–X2. It shows the maximum extraction yield near center of extraction number and high ratio of solvent to raw material. The figures show good results both near center of extraction number and high extraction temperature level studied. Figures 2e and 3e show the effect of extraction time and extraction number on extraction yield. the measured extraction yield were close to those Fig. the more steep the curve was the more significant the factors on the response value were. Figures 2f and 3f show the effect of extraction number and ratio of solvent to raw material on extraction yield. Figures 2d and 3d show the effect of extraction time and ratio of solvent to raw material on extraction yield. Figures 2b and 3b show the effect of extraction temperature and ratio of solvent to raw material on extraction yield and Figs. e X2–X4.

This result is slightly different from the other authors’ findings (Qiao et al. 2001.776 4 4 0. d X2–X3. 2005). We postulate that different solvents used in the SE method is responsible for the difference. Lin et al. 3 Contour plots of extraction yield showing the effect of variables: a X1–X2.782 . diallyl trisulfide. b X1–X3. and dimethyl disulfide were the main components of garlic essential oil. c X1–X4. Composition of garlic oil obtained by different methods Table 6 listed the composition of garlic essential oil obtained by two different methods according to the results of GC–MS.. and f X3–X4 estimated using RSM. e X2–X4. and relative concentrations of the identified main compounds were slightly different. This also testifies that the RSM approach was appropriate for optimizing the operational conditions of the extraction process. The composition of the garlic essential oil extracted by two different methods was mostly similar. It could be seen that 23 compounds in the garlic essential oil had been identified.. predicted. in which 3-vinyl-4H-1. propyl allyl disulfide.2-dithiin. and experimental value of response at that condition Optimum condition Extraction yield (%) Extraction temperature (°C) Extraction time Ratio of solvent to raw Extraction (min) material number Experimental Predicted 45 42 0. Table 5 Optimum conditions.1102 Med Chem Res (2010) 19:1092–1105 Fig.

product quality. the process cost and scale of SE in the extracted oil was by far lower and larger than that of SFE.22 4 Methyl propyl disulfide 1.3 2.02 6.06 5 3-methyl-2-cyclopentene-1-thione 0.87 8 Dimethyl disulfide 7.3-dithiin 1.17 31.2-dithiin 32.02 14 Diallyl trisulfide 12. Therefore. However.01) lower than that in group I rats (control).31 15 3.78 130 min 42 min SFE supercritical fluid extraction.56 16 Methyl 2-propenyl disulfide 0.10 0.67 5. and CD8?.89 11 Dially disulfide 8.81%) obtained by SFE.52 6 Vinylfuran 0.44 7.3 2 1.Med Chem Res (2010) 19:1092–1105 1103 Table 6 Chemical composition of garlic essential oil NO. process cost. However.2-epithiopropane 0.87 12 3-vinyl-4H-1.92 7 Methyl allyl disulfide 1.78%) of garlic by employing SE method was slightly lower than that (0.4-trithiolane 2.92 1. the extract time (42 min) by SE method was shorter than that (130 min) obtained by SFE.62 2.98 0. Compounds 1 3-methylthio propanal 0. essential oil-treated rats dose-dependently and significantly (P \ 0.2. extraction yield and product quality obtained by SE was basically similar to the ones obtained by SFE. CD4?.78 Extraction yield (%) = (essential oil/garlic sample) 9 100 Extraction time (min) SFE (%) SE (%) 0. SE solvent extraction Extraction yield.5-diethyl-1. . ratio of CD4?/CD8? and CD40 expression in gastric cancer rats In contrast to control rats (group I).34 3 Diallyl sulfide 2. In addition.12 1.01) increase in the CD4?/CD8? ratio in groups II and III rats after the essential oil treatment as compared to the group I rats (Table 7).94 13.34 17 2-vinyl-4H-1. Furthermore. Effect of garlic oil on mean (?SE) percentage of CD3?. the percent of CD8? leukocytes and CD40 expression in groups II and III was significantly (P \ 0.81 0. respectively.89 13 Methyl allyl trisulfide 4.14 1. comprehensive comparisons of the garlic essential oils obtained by two different methods were further done.05 9 Methyl butyl trisulfide 10 Propyl allyl disulfide 13. One consequence of this bidirectional effect was a significant (P \ 0. and scale are the important factors for the industrialization. extraction time.01) increased the percent of CD3? and CD4? leukocytes in groups II and III.04 0.95 13. It could be seen from Table 6 that the extraction yield (0.

02a Group (III) High dose a 57. the result.1 ± 2.73 ± 0.09 P \ 0.3 ± 1. ratio of solvent to raw material. Han C-M. Shih C-C. ratio of solvent to raw material 4. Weng C-C. blood sugar. e.1 ± 2. Although main compositions of the essential oils obtained by SE and SFE are basically similar.8 ± 4.1 ± 1. compared with control rats Conclusions The objectives of this study were to evaluate the effect of four major variables (extraction temperature. Abd El-Aty AM.4 32. Choi J-H. their minor compositions do differ quantitatively.03a a a a a 1. Results of optimal experiment indicate that both extraction temperature and ratio of solvent to raw material had the most apparent effect on the yield of essential oil. J Ethnopharm 108:385–394 Cho S-K. that is. extraction time 42 min.87 ± 0. and extraction number 4. Tsai S-Y. CD4? and CD8?.49 ± 0. and the number of extraction) on the performance of the SE of garlic essential oil.5 22.8 ± 3. the detection of a significant increment in ratio of CD4?/CD8? in serum of rats after administration of garlic essential oil.8a 0. In addition.9 0. Prostaglandins Leukot Essent Fatty Acids 58:257–263 Chauhan NB (2006) Effect of aged garlic extract on APP processing and tau phosphorylation in Alzheimer’s transgenic model Tg2576. ratio of CD4?/CD8? and CD40 expression in gastric cancer rats CD3? CD4? CD8? CD4?/CD8? CD40 (I) Control 40. Therefore.05 1. Food Chem 100:808–812 .9 ± 1. References Bordia A. SS0711).g. According to extraction yield. comprehensively considering various factors. J Pharm Biomed Anal 44:23–29 Chyau C-C. Acknowledgment The work was supported by the National Natural Science Foundation of China (No. Kim MR. However. SE is considered as the optimum process for obtaining garlic essential oil with high quality. Srivastava KC (1998) Effect of garlic (Allium Sativum) on blood lipids.5 ± 2. Verma SK.78%. it can be concluded that the SE method offers obvious advantages over SFE.01 (II) Low dose 48.16 ± 0. Shim JH (2007) Optimized conditions for the extraction of secondary volatile metabolites in Angelica roots by accelerated solvent extraction.5 1.61 ± 0. fibrinogen and fibrinolytic activity in patients with coronary artery disease. indicated that garlic essential oil may be useful for treatment of patients with inflammatory disease. Yang J-H. the best optimal extraction conditions were extraction temperature 45°C.2a 29..68 ± 0. extraction yield of SE was slightly higher than that obtained by SFE. when the yield was 0.01.4 37. gastric cancer.1104 Med Chem Res (2010) 19:1092–1105 Table 7 Effect of garlic essential oil on mean (?SE) percentage of CD3?.04a 1.3 45.7 ± 1. Mau J-L (2007) The essential oil of Glossogyne tenuifolia. In addition.2a 39. extraction time. 30872943) and the social development projects of Suzhou (No. The compositions of garlic essential oil obtained by SE and SFE methods were compared.

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