Supercritical Carbon Dioxide (SC-CO ) Extraction of Catechin, Epicatechin,. 2. Rutin and Luteolin from Spearmint (Mentha spicata L.) Leaves. M. Bimakr, R.A. ...
World Applied Sciences Journal 5 (4): 410-417, 2008 ISSN 1818-4952 © IDOSI Publications, 2008
Supercritical Carbon Dioxide (SC-CO2) Extraction of Catechin, Epicatechin, Rutin and Luteolin from Spearmint (Mentha spicata L.) Leaves 1
M. Bimakr, 1,2R.A. Rahman, 1F.S. Taip, 1L.T. Chuan, 2A. Ganjloo, 1 L. Md Salleh, 3J. Selamat and 3A. Hamid
Department of Process and Food Engineering, Faculty of Engineering, University Putra Malaysia 43400 Serdang, Selangor, Malaysia 2 Department of Food Technolgy, Faculty of Food Science and Technology, University Putra Malaysia 43400 Serdang, Selangor, Malaysia 3 Department of Food Science, Faculty of Food Science and Technology, University Putra Malaysia 43400 Serdang, Selangor, Malaysia 1
Abstract: In this study, supercritical carbon dioxide (SC-CO2) extraction conditions were optimized for the simultaneous separation of four bioactive flavonoids (catechin, CA; epicatechin, EP; rutin, RU; luteolin, LU) contained in spearmint (Mentha spicata L.) leaves. SC-CO2 extraction parameters such as pressure, temperature and dynamic extraction time were optimized by Complete Randomize Design (CRD) full factorial. The optimum condition of SC-CO2 extraction was achieved at 200 bar, 60°C and 60 min (with 60.566 mg/g extraction yield). Extracted yield at optimum condition was then analyzed by high performance liquid chromatography (HPLC) for quantifying bioactive flavonoid compounds. At optimum conditions, four bioactive flavonoids including (+)-catechin, (-)-epicatechin, rutin and luteolin were detected at high concentration. Luteolin (0.657 mg/g) had the highest concentration among the other detected flavonoids. The results indicated that SC-CO 2 extraction is a promising and alternative process for recovering the bioactive compounds from spearmint leaves. Key words: Spearmint (Mentha spicata L.) Bioactive Flavonoid (+)-Catechin Luteolin Supercritical Carbon dioxide (SC-CO2) Extraction INTRODUCTION
Rutin
cancer, heart disease, dermal disorders and aging [3-5]. Occurrence of flavonoids in common foods are shown in Table 1 [6]. Spearmint belongs to the genus Mentha in the family Labiateae (Lamiaceae). Genus Mentha consisting about 25–30 species including spearmint, peppermint, wild mint, corn mint, curled mint, bergamot, American mint, Korean mint, etc. of which spearmint is the most common of all. Mint has post digestive effect and hot potency and its taste is pungent [7, 8]. Traditionally, spearmint (Mentha spicata L.) which is common plant in Britain and other European countries, has been used in folk remedies for exhaustion, weakness, depression, memory enhancement, circulation improvement and strengthening fragile blood vessels [9]. Researchers have found that Labiatae family plants are a source of compounds possessing high antioxidant [10], anti-inflammatory [11], anti-allergy [12] and antidepression [13] activity.
Since 1949, a number of synthetic antioxidants have been used to prevent oxidation of lipids during processing and storage of foods. By considering to possible adverse effects of some synthetic food additives on human health there is a great interest on natural antioxidants [1]. Flavonoid compounds, abundant in fruits, vegetables, teas, medicinal plants, are a kind of highly effective natural antioxidants and less toxic than synthetic antioxidants [2]. The consumption of fruits and vegetables plays a key role as health protecting factor. This beneficial effect is mainly associated with the antioxidant activity of the flavonoid compounds. Antioxidants act as reducing agents by donating hydrogen, by quenching singlet oxygen, by acting as chelators and by trapping free radicals. These highly reactive molecules are present in biological systems and can reduce the risk of degenerative diseases such as Coressponding Author:
(-)-Epicatechin
R.A. Rahman, Department of Process and Food Engineering, Faculty of Engineering, University Putra Malaysia 43400 Serdang, Selangor, Malaysia
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In recent years, a great deal of study has been done to use supercritical fluid extraction (SFE) with carbon dioxide (CO2) as a solvent for extraction of natural compounds from different raw materials [9]. Supercritical fluid extraction was developed in 1960. In this technique, supercritical carbon dioxide is widely used as an extraction solvent. Carbon dioxide has the following advantages: chemically inert, low toxicity, no pollution and shorter concentration time. These advantages have attracted increasing interest from researchers. Nowadays, the application of supercritical fluid extraction (SFE) includes the industries of food, pharmacy and environmental engineering etc [14]. The special note of this process for selective extraction of soluble compounds from a raw material is usage of gases above their critical points [15-18]. Several researchers studied the extraction of natural compounds from plant matrix by using supercritical carbon dioxide (SC-CO2) [19, 20]. For example, Leal et al. [21] determined the antioxidant activity of various SFE extracts, among them rosemary extract. Reverchon and Sanatore [22] reported that the content of 1,8-cineol in rosemary volatile oil and rosemary SFE extract was approximately constant. The camphor content of the volatile oil was 10.26% and that of the SFE extract 15.33%. The SFE of Mentha spicata L. essential oil has been studied previously [23, 24]. Also, there are a few reports on the antioxidant property of Mentha [25-27] but to the best of our knowledge, there is not any report about the supercritical fluids extraction of Malaysian Mentha spicata L. flavonoid compounds. This study was undertaken to approach the following goals: (i) finding the best SC-CO2 extraction condition to obtain the highest crude extraction yield (ii) determination the quality and quantity of the catechin, epicatechin, rutin and luteolin in the extract obtained with best and control SC-CO2 extraction conditions.
0.53 mm. Carbone dioxide (purity 99.99%), contained in a diptube cylinder, was purchased from MOX Company in Malaysia. Ethanol (EtOH, 99.5%, analytical grade) and methanol (MeOH, HPLC grade) were purchased from Fisher Scientific Chemical (Loughborough, England). TFA (tri-flouro-acetic acid, 98%) and all flavonoid standards including (+)-catechin, (-)-epicatechin, apigenin, rutin, luteolin, kaempferol, myricetin and naringenin (purity 99.99%), were purchased from Aldrich Chemical Co. (Gillingham-Dorset, England). Supercritical Carbon Dioxide (SC-CO2) Extraction: Supercritical carbon dioxide (SC-CO2) extraction experiments were carried out using a supercritical fluid extractor (ABRP200, Pittsburgh, PA, USA) with the extractor vessel volume 500 ml. Figure 1shows schematic diagram of supercritical fluid extractor. The flow rate of CO2, extraction temperature and pressure were adjusted by using ICE software and the extraction time was measured. Liquid CO2 was supplied from a gas cylinder. Before the liquid CO 2 passed into the extraction vessel, it was pressurized to the desired pressure and heated to the specified temperature by the means of a pump (P-50, Pittsburg, PA, USA) to reach the supercritical state. Absolute ethanol (EtOH) acted as the modifier. The supercritical CO2 flow rate was maintained at 15 g/min and also, the duration of static extraction time was fixed to 30 min. The powdered spearmint leaves (30 g) was mixed with 90 g glass beads (2.0 mm in diameter) and placed into the extractor vessel. The extraction was then performed under various experimental conditions in accordance with the full factorial with complete randomize design (CRD). All obtained extracts were collected with modifier. In order to remove ethanol extracts were vacuum evaporated using a rotary evaporator (Eyela, A-1000S, Japan) at 40°C. The extract then placed in the oven at 40°C for 30 minutes before transferred into a desiccator for final constant weight. Then a gravimetric measurement was used to obtain the amount of total extract weight. Finally, the obtained extracts under the optimum and control SFE conditions were transferred into brown glass bottles and stored in a freezer until HPLC analysis. The extractions were performed in duplicate.
MATERIALS AND METHODS Raw Materials: The leaves of spearmint (Mentha spicata L.) were obtained from Cameron Highlands, Pahang, Malaysia. After harvesting, the leaves were separated and washed under tap water. Leaves were dried at 40°C in a ventilated drying oven (1350FX, USA) for 24 h and then stored at ambient temperature in the dark place. To avoid degradation, the dried plant material was ground just before extraction. The dried plant leaves were ground in a blender (MX-335, Panasonic, Malaysia) for 10 s to produce powder with an approximate size of
High Performance Liquid Chromatography (HPLC) Analysis: The extracts that were obtained from the optimum and control SFE conditions were analyzed using a high-performance liquid chromatography (HPLC) which was consist of a Water 600 pump Controller, 9486 tunable 411
World Appl. Sci. J., 5 (4): 410-417, 2008
Fig. 1: Schematic diagram of supercritical fluid extractor rate was set at 1.0 ml/min. Quantity calculations were made according to the linear calibration curves of standards.
Table 1: Occurrence of flavonoids in common foods Flavonoids subgroup
Major foods
Examples of foods
Flavones
Herbs
Parsley, Thyme
Flavonol
Vegetable Fruits Beverages
Onion, kale, broccoli Apples, cherries, berries Tea, red wine
Flavanones
Fruits
Citrus
Catechins
Fruits Beverages
Apples Tea
Anthocyanidins
Fruits
Cherries, grapes
Isoflavones
Vegetable
Soya beans, legumes
Statistical Analysis: SFE operations always involve many variables that may affect the efficiency of the extraction. Selection of these variables and their levels is critical. Several statistical techniques such as simplex optimization and factorial design have been employed for the optimization of analytical methods [28, 29]. Factorial design has some advantages over simplex optimization in that global optimum can be provided, large amounts of quantitative information can be extracted and both discrete and continuous factors can be estimated. However, the number of the experiments can be considerably reduced by the use of orthogonal array design [30, 31]. In this work, the SFE of spearmint leaves were planned according to the full factorial with complete randomize design (CRD) for the highest crude extraction yield. The factors and levels investigated were reported in Table 2. Data were subjected to analyses of variance (ANOVA) and multiple comparison tests were performed using a least significant difference (LSD) at 95% of confidence level. All the analyses were carried out using the statistical software, MINITAB release 14. A probability value of P< 0.05 was considered significant.
Table 2: Experimental levels of the factors used in Complete Randomized Design (CRD) full factorial
Factors
Levels ----------------------------------------------------1 2 3
Pressure (bar)
100
200
300
Temperature (°C)
40
50
60
Dynamic time (min)
30
60
90
absorbance UV detector and equipped with an Eclipes XDR- C18 reversed-phase column (25 cm× 4.6 mm × 5 µm, Supelco, USA). The volume of the injection loop was 20 µl. Classic Millenium 2010 software was used for manipulation of data processing. The mobile phase used for analysis was solvent A: TFA (triflouroacetic acid) 2.5 pH in deionized water and solvent B consists of 100% Methanol (HPLC grade). Also, 70% methanol was required for washing of the system. The flavonoids were detected at 280 nm. All of the main flavonoid compounds were identified by matching their retention time against those of standard compounds. The temperature was set to room temperature and flow
RESULTS AND DISCUSSION Effect of the experimental conditions on the SC-CO2 extraction yield: Generally, extraction pressure, 412
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Fig. 2: The yields of crude extract under Complete Randomize Design (CRD) full factorial
Fig. 3:
HPLC chromatograms. (A): mix standard, (B): SC-CO2 extraction with 100 bar, 40°C and 30 min (minimum condition as control run), (C) SC-CO2 extraction with 200 bar, 60°C and 60 min (optimum condition), (D): SC-CO2 extraction with 300 bar, 60°C and 90 min (maximum condition as control run)
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World Appl. Sci. J., 5 (4): 410-417, 2008 Table 3: Results obtained at the experimental condition using Complete Randomized Design (CRD) full factorial Parameter
Yield (mg/g) L1 a
Yield (mg/g) L2 a
Yield (mg/g) L3 a
Kb
Pressure
40.69
51.07
45.13
10.38
Temperature
40.48
45.36
51.06
10.58
Time
40.28
48.21
48.40
8.12
Average responses of each level about extraction yield
a
K value means range between three average responses of each level about extraction yield
b
Table 4: Identification and quantification of the compounds extracted by SFE under different conditions Extraction Mode
Yield (mg/g)
Flavonoid Content (mg/g) (+)-Catechin
(-)-Epicatechin
Rutin
Luteolin
---------------------------------------------------------------------------------------------------------------------------Type 1 a
30.133
0.120
0.100
0.105
0.060
Type 2 b
60.566
0.140
0.156
0.148
0.657
Type 3c
52.900
0.103
0.128
-
0.101
a
Minimum level of each studied parameter (40°C, 100 bar and 30 min)
b
Optimum level of each studied parameter (60°C, 200 bar and 60 min)
c
Maximum level of each studied parameter (60°C, 300 bar and 90 min)
temperature and dynamic time are considered as the most important factors. In this study, full factorial with completely randomized design (CRD) was used to obtain the highest crude extraction yield and determine the best and optimum SC-CO2 extraction condition. After that, the product of best extraction condition with another two SFE condition as control runs are analysed by HPLC to detect the major bioactive flavonoid compounds. A full evaluation of the effect of three selected factors from three levels (Table 2) and duplicate experiments on the extraction yield needed 54 (3×3×3×2) experiments. The results of extraction yield were expressed as the yield of crude extract as follows:
long extraction time, 60 min dynamic extraction time (with 60.57 mg/g extraction yield) was preferable to other tested levels of dynamic time for SC-CO 2 extraction of flavonoids from spearmint leaves. The average response of each level about extraction yield is presented in Table 3. The K value which is mentioned in Table 3 represents the range between three average responses of each level about extraction yield. From the K value it can be concluded that temperature (with K value of 10.58) had a dominant effect on the crude extract yield after which followed by the pressure (with K value of 10.38) and dynamic time (with K value of 8.12). Identification and Quantification of Catechin, Epicatechin, Rutin and Luteolin: The optimum and best SC-CO2 extraction condition for obtaining the highest crude extract yield from spearmint (Mentha spicata L.) leaves was: pressure, 200 bar; extraction temperature, 60°C; dynamic time, 60 min. The product of best condition was analyzed by HPLC in order to determine the contents of CA, EP, RU and LU in the spearmint (Mentha spicata L.) extracts. Also, the product of another two SFE conditions which included minimum (100 bar, 40°C, 30 min) and maximum (300 bar, 60°C, 90 min) level of each studied parameters as control runs were analyzed by HPLC. All flavonoid compounds were identified by matching the retention time and their spectral characteristics against those of standards. Quantification of the mentioned flavonoid contents in various crude extracts of spearmint (Mentha spicata L.) obtained under optimum and control SC-CO2 extraction conditions were
m mg Yextract ( ) = 1000 x extract g m herb
Where; m extract is the crude extract mass (g) and m herb is the extracted herb mass (g). Figure 2 shows the variation of the extraction yields obtained under different experimental conditions. The mean values of the extraction yields for the corresponding parameters at each level were calculated according to the assignment of the experiment. The mean values of the three levels of each parameter showed how the extraction yield changes when the level of that parameter is changed. The highest mean value of extraction yield was obtained at 200 bar, 60°C and 90 min and the difference of obtained extraction yield between 60 and 90 min was not significant according to the t-test analysis (P value > 0.05). Therefore by considering to economic costs and degradation of sensitive compounds during 414
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calculated from the calibration curves of standard compounds. Detailed identification and quantification of the compounds extracted by SFE under different conditions were presented in Table 4. By considering to the mentioned HPLC chromatographs in Figure 3 A-D, the variation in the concentration of flavonoid compounds was obvious. Optimum condition was more efficient than minimum and maximum levels of studied parameters for obtaining bioactive flavonoid compounds from spearmint leaves. At minimum level of each studied parameters (40°C, 100 bar and 30 min) four flavonoid compounds, CA, EP, RU and LU, were extracted at lower concentration and many residues were existed. However, at the best SC-CO 2 extraction condition (60°C, 200 bar and 60 min) bioactive flavonoids including CA, EP, RU and LU were detected with good separation and high concentration. In the product obtained under the best condition luteolin (0.657 mg/g) had the highest concentration among the other flavonoids. In the extract obtained under maximum levels of each parameter (60°C, 300 bar and 90 min) only three bioactive flavonoids including CA, EP and LU were identified. With using this condition the highest concentration was belonged to (-)-epicatechin (0.128 mg/g). For a volatile solute, there is a competition between its solubility in supercritical carbon dioxide and its volatility [32]. Therefore, it is difficult to predict the effect of temperature. In the present study, the concentration of bioactive compounds increased with temperature (40 to 60°C) and the highest concentrations of flavonoids were obtained at 60°C. In this manner, the solute vapour pressure played a key role leading to increase the bioactive concentration. According to the obtained results, the bioactive concentration increased with pressure from 100 to 200 bar, that was due to increase of SC-CO2 density at higher pressures. However, an increase in the pressure level above 200 bar led to an unexpected reduction in the bioactive concentration. This unexpected result can probably be related to the reduced diffusion rates of the extracted compounds from the plant matrix to the supercritical fluid medium [33]. Therefore, in this study, the highest bioactive concentration were achieved at 200 bar and at pressure above 200 bar, less extraction efficiency than that expected by the solubility enhancements at higher pressures (above 200 bar) was obtained in result of major role of diffusion in the mass transfer rates of the extractable materials from the sample matrix into the supercritical fluid. Therefore, it is interesting to control the composition of the extract using
pressure. In this study, the flavonoid yield increased with increasing pressure to a certain value (200 bar). Over this range of pressure, increasing fluid density is presumably the main mechanism leading to a higher flavonoid yield. Above this range of pressure, a decreasing flavonoid yield with increasing pressure was observed. The volatility and polarity of extracted analytes might be responsible for the result [34, 35]. By considering to the flavonoid concentration the preferable dynamic extraction time was 60 min. By using the shortest dynamic extraction time (30 min) the lower concentration of flavonoids was extracted. In contrast, by using the longest dynamic extraction time (90 min) rutin was disappeared and the concentrations of the other flavonoids were reduced. Bioactive flavonoid compounds are thermolabile compounds [24] so, it is necessary to reduce the time of extraction as it is possible. Therefore, 60 min dynamic extraction time was more suitable and efficient for isolation of CA, EP, RU and LU from spearmint leaves. Finally, it can be concluded that, using the best SC-CO2 extraction condition (pressure, 200 bar; extraction temperature, 60°C; dynamic time, 60 min) is more efficient for extraction of CA, EP, RU and LU from spearmint leaves. However, there are may be some novel flavonoids compound of spearmint with using higher pressure or longer extraction time. Using only HPLC, it has not possible to identify all of the flavonoid compounds in the spearmint extract. CONCLUSION In summary, it may be possible to extract the bioactive flavonoid compounds (catechin, epicatechin, rutin and luteolin) from the spearmint leaves by manipulating extraction condition of SC-CO 2 extraction. Among several studied parameters which affected on the SC-CO2 efficiency temperature had the dominant effect on the crude extract yield after that followed by pressure and dynamic time. At the best SC-CO 2 extraction condition (pressure, 200 bar; extraction temperature, 60°C; dynamic time, 60 min) CA, EP, RU and LU were detected at high concentration and luteolin had the highest concentration (0.657 mg/g) among the other studied flavonoids. Finally it can be concluded that supercritical carbon dioxide extraction is a promising method for the extraction of bioactive flavonoid compounds (catechin, epicatechin, rutin and luteolin) from spearmint leaves. Also based on the obtained HPLC chromatograms spearmint is a natural source of bioactive flavonoid compounds such as catechin, epicatechin, rutin and luteolin. 415
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ACKNOWLEDGMENT
12. Ito, H., T. Miyazaki, M. Ono and H. Sakurai, 1998. Antiallergic activities of Rabdosiin and its related compounds: chemical and biochemical evaluations. Bioorganic and Medicinal Chem., 6: 1051-1056. 13. Takeda, H., M. Tsuji, T. Matsumiya and M. Kubo, 2002. Identification of rosmarinic acid as a novel antidepressive substance in the leaves of Perilla frutescens Britton var. acuta Kudo (Perillae Herba). Nihon Shinkei Seishin Yakurigaku Zasshi, 22(1): 15-22. 14. Mei-Chih L., M.J. Tsai and K.C. Wen, 1999. Supercritical fluid extraction of flavonoids from Scutellariae Radix. J. Chromatography A, 830: 387- 395. 15. Sofia, C., R.G.R. Monica, R.M. Francisco, J. Laura, S. Susana, F.J. Senorans, R. Guillermo and I. Elena, 2006. Supercritical fluid extraction of antioxidant compounds from oregano Chemical and functional characterization via LC-MS and in vitro assays. J. Supercritical Fluids, 38: 62-69. 16. Baysal, T., S. Ersus and D.A.J. Starmans, 2000. Supercritical CO2 extraction of B-carotene and lycopene from tomato paste waste. J. Agric. Food Chemical, 48: 5507-5514. 17. Diaz-Maroto, M.C., M.S. Perez-Coello and M.D. Cabezudo, 2002. Supercritical carbon dioxide extraction of volatiles from spices: comparison with simultaneous distillation-extraction. J. Chromatography A, 947: 23-33. 18. Elena, I., A. Oca, G. De-Murga, S. Lopez-Sebastian, J. Tabera and G. Reglero, 1999. Supercritical fluid extraction and fractionation of different preprocessed rosemary plants, J. Agric. Food Chem., 47: 1400-1409. 19. Povh, N.P., M.O.M. Marques and A.A. Meireles, 2001. Supercritical CO2 extraction of essential oil and oleoresin from chamomile (Chamomilla recutita L. Rauschert). J. Supercritical Fluids, 21: 245-256. 20. Ebrahimzadeh, H., Y. Yamini, F. Sefidkon, M. Chaloosi and S.M. Pourmortazavi, 2003. Chemical composition of the essential oil and supercritical CO2 extracts of Zataria multiflora Boiss. Food Chem., 83: 357-361. 21. Leal, P.F., M.E.M. Braga, D.N. Sato, J.E. Carvalho, M.O.M. Marques and M.A.A. Meireles, 2003. Functional properties of spice extracts obtained viasupercritical fluid extraction. J. Agric. Food Chem., 51: 2520-2532. 22. Reverchon, E. and F. Sanatore, 1992. Isolation of rosemary oil: comparasion between hydrodistillation and supercritical CO2 extraction. J. Flavonoid Fragrance, 7: 227-236.
The authors are grateful for the financial support received from the RMC, the Universiti Putra Malaysia for this project. REFERENCES 1.
Grigonis, D., P.R. Venskutonis, B. Sivik, M. Sandahl and C.S. Eskilsson, 2005. Comparison of different extraction techniques for isolation of antioxidants from sweet grass (Hierochloe odorata). J. Supercritical Fluids, 33: 223-233. 2. Benguo, L. and Z. Yongyi, 2007. Extraction of flavonoids from flavonoid-rich parts in tartary buckwheat and identification of the main flavonoids. J. Food Engineering, 78: 587-596. 3. Cook, N.C. and S. Samman, 1996. Flavonoids: chemistry, metabolism, cardioprotective effects and dietary sources. Nutritional Biochem., 7: 66-76. 4. Harborne, J.B. and C.A. Williams, 2000. Advances in flavonoids research since 1992. Phytochem., 55: 481-504. 5. Heim, K.E., A.R. Tagliaferro and D.J. Bobilya, 2002. Flavonoid antioxidants: chemistry, metabolism and structure-activity relationship. The J. Nutritional Biochem., 13: 572-584. 6. Hollman, P.C.H., M.G. Hetrog and M.B. Katan, 1996. Analysis and health effects of flavonoids. Food Chem., 57: 43-46. 7. Kivilompolo, M. and T. Hyotylainen, 2007. Comprehensive two-dimensional liquid chromatography in analysis of Lamiaceae herbs: Characterisation and quantification of antioxidantphenolicacids. J. Chromatography A, 1145: 155-164. 8. Choudhury, R.P., A. Kumar and A.N. Garg, 2006. Analysis of Indian mint (Mentha spicata) for essential, trace and toxic elements and its antioxidant behavior. J. Pharmaceutical and Biomedical Analysis, 41: 825-832. 9. Huafu, W., J.P. Gordon and H. Keith, 2004. Determination of rosmarinic acid and caffeic acid in aromatic herbs by HPLC. Food Chem., 87: 307-315. 10. Zheng, W. and S.Y. Wang, 2001. Antioxidant activity and phenolic compounds in selected herbs. J. Agric. Food Chem., 49(1): 5165-5170. 11. Al-Sereiti, M.R., K.M. Abu-Amer and P. Sen, 1999. Pharmacology of rosemary (Rosmarinus officinalis Linn.) and its therapeutic potentials. Indian J. Experimental Biol., 37: 124-130. 416
World Appl. Sci. J., 5 (4): 410-417, 2008
23. Barton, P., R.E. Hughes and M.M. Hossein, 1992. Supercritical carbon dioxide extraction of peppermint and spearmint. J. Supercritical Fluids, 15: 157-162. 24. Aghel, N., Y. Yamini, A. Hajiakhoondi and S.M. Pourmortazavi, 2004. Supercritical carbon dioxide extraction of Mentha pulgium L. essential oil. Talanta, 62: 407-411. 25. Sweetie R., R.C. Kanatt and A. Sharma, 2007. Antioxidant potential of mint (Mentha spicata L.) in radiation-processed lamb meat. Food Chemistry, 100: 451-458. 26. Dorman, H.J.D., M. Kosar, K. Kahlos, Y. Holm and R. Hiltunen, 2003. Antioxidant properties and composition of aqueous extracts from Mentha species, hybrids, varieties and cultivars. J. Agric. Food Chem., 51: 4563-4569. 27. Marinova, E.M. and N.V. Yanishlieva, 1997. Antioxidative activity of extracts from selected species of the family Lamiaceae in sunflower oil. Food Chem., 58: 245-248. 28. Roy, B.C., M. Goto and T. Hirose, 1996. Extraction of ginger oil with supercritical carbon dioxide: experiments and modelling. Industrial and Engineering Chem. Res., 35: 607-612. 29. Kazazi, H., K. Rezaei, S.J. Ghotb-Sharif, Z. EmamDjomeh and Y. Yamini, 2007. Supercriticial fluid extraction of flavors and fragrances from Hyssopus officinalis L. cultivated in Iran. Food Chem., 105: 805-811.
30. Lan, W.G., M.K. Wong, N. Chen and Y.M. Sin, 1994. Orthogonal array design as a chemometric method for the optimization of analytical procedures Part 1. Two-level design and its application in microwave dissolution of biological samples. Analyst, 119: 1659-1667. 31. Lan, W.G., K.K. Wong, H.K. Lee and Y.M. Sin, 1995. Orthogonal array design as a chemometric method for the optimization of analytical procedures Part 4. Mixed-level design and its application to the high-performance liquid-chromatographic determination of polycyclic aromatic hydrocarbons. Analyst, 120: 281-287. 32. Pourmortazavi, S.M. and S.S. Hajimirsadeghi, 2007. Supercritical fluid extraction in plant essential and volatile oil analysis. J. Chromatography A, 1162: 2-24. 33. Rezaei, K. and F. Temelli, 2000. Using supercritical fluid chromatography to determine the binary diffusion coefficient of lipids in supercritical CO 2. J. Supercritical Fluids, 17: 35-44. 34. Gomes, P.B., V.G. Mata and A.E. Rodrigues, 2007. Production of rose geranium oil using supercritical fluid extraction. J. Supercritical Fluids, 41, 50-60. 35. Wang, L., B. Yang, X. Du and C. Yi, 2008. Optimisation of supercritical extraction of flavonoids from Pueraria lobata. Food Chemistry, 108: 737- 741.
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