Supercritical Fluid Extraction and Micronization of Ginkgo Flavonoids

Apr 30, 2010 - Supercritical Fluid Extraction and Micronization of Ginkgo Flavonoids from. Ginkgo Biloba Leaves. Shi-Feng Miao, Jin-Peng Yu, Zhe Du, Y...
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Ind. Eng. Chem. Res. 2010, 49, 5461–5466

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Supercritical Fluid Extraction and Micronization of Ginkgo Flavonoids from Ginkgo Biloba Leaves Shi-Feng Miao, Jin-Peng Yu, Zhe Du, Yi-Xin Guan,* Shan-Jing Yao, and Zi-Qiang Zhu Department of Chemical and Biological Engineering, Zhejiang UniVersity, Hangzhou 310027, China

Ginkgo flavonoids were extracted from Ginkgo biloba leaves using supercritical carbon dioxide extraction technology (SFE) with ethanol as the entrainer. The effects of important process parameters on SFE were examined, and the crude extracts were further refined. At optimized experimental conditions, i.e., 20 MPa of extraction pressure, a 40 °C extraction temperature, a 10 g/min CO2 flow rate, and a 6 mL/min ethanol flow rate, the extraction yield could reach 0.36%. Supercritical fluid assisted atomization introduced by a hydrodynamic cavitation mixer (SAA-HCM) was used to obtain microparticles of Ginkgo flavonoids. The influences of the solute concentration as well as the mass flow ratio of the CO2/liquid solution on particle size (PS) and particle size distribution (PSD) were investigated. Well-defined, spherical, and separated particles with a controlled particle size ranging between 0.2 and 3.0 µm were successfully obtained in this work. The micronization of Ginkgo flavonoids can improve the drug absorption in the body and has great potential for use in drug delivery systems. 1. Introduction Ginkgo biloba has been used in traditional Chinese medicine for thousands of years, and it is helpful in inhibiting the onset of dementia, slowing down cognitive decline and functional disability, and reducing the incidence of cardiovascular disease due to its ability to prevent free radical damage, improve brain function, and support microcirculation.1,2 Ginkgo biloba leaves contain various species of active ingredients,3 such as ginkgolides, bilobalide, flavonoids, proanthocyanidins, alkylphenols, simple phenolic acids, and so on. Ginkgo flavonoids are an important class of compounds in Ginkgo biloba leaves and usually act as scavengers of different oxidizing species,4 i.e., superoxide anions, hydroxyl radicals, or peroxy radicals. Ginkgo flavonoids can be obtained using supercritical carbon dioxide extraction technology (SFE), as extensively reported.5 In contrast with conventional extraction processes using liquid solvents, SFE is an ideal alternative to deal with heat-sensitive or easily oxidizable material. Without the addition of any toxic organic solvents, the application of SFE in the isolation and separation of natural products is becoming very popular. However, Ginkgo flavonoids cannot be absorbed completely in the alimentary canal,6 partially because some of the flavonoids in Ginkgo biloba leaves are poorly water-soluble substances. Micronization techniques are usually used to improve dissolution rates of poorly water-soluble drugs into biological environments.7 Recent research8 indicates that the absorption of micronized flavonoids is much better than that of nonmicronized ones when flavonoids are taken orally. Supercritical fluid assisted atomization introduced by a hydrodynamic cavitation mixer (SAA-HCM) has been proposed by Cai et. al,9 where a hydrodynamic cavitation mixer is used to replace the usual packed bed saturator in the supercritical fluid assisted atomization (SAA) process to intensify the mass transfer between the solution and CO2 and improve the mixing. Just like the SAA process, the SAA-HCM process is based on the dissolution of supercritical CO2 (SC-CO2) in a liquid solution containing the drug molecules and the atomization of the mixing solution through a nozzle. Therefore, SC-CO2 behaves both * To whom correspndence should be addressed. Tel.: +86-57187951982. Fax: +86-571-87951982. E-mail: [email protected].

as a cosolute and as a pneumatic agent. A hydrodynamic cavitation mixer allows for the generation of cavity collapse similar to acoustic cavitation, i.e., the formation of cavitation bubbles and cavities within a liquid stream or at the boundary of the streamlined body resulting from a localized pressure drop in the liquid flow. Thus, complete solubilization is achieved in the hydrodynamic cavitation mixer, and the solution is then sent to a thin wall injector and sprayed into the precipitator. A two-step atomization is obtained: the first step is pneumatic atomization including the process where the primary droplets are produced at the outlet of the injector; the second step is decompressive atomization including the process where the primary droplets are further divided into secondary droplets by CO2 expansion from the inside of the primary ones.10,11 After evaporating the droplets with heated nitrogen, drug microparticles are obtained. The employment of a hydrodynamic cavitation mixer was successful in micronizing roxithromycin9 and levofloxacin hydrochloride12 with the SAA-HCM process, and well-defined microspheres were obtained at optimized operating parameters. The SAAHCM process can be applied to produce microparticles of both water-soluble and non-water-soluble compounds, and its application is extending from antibiotics to polymers and proteins rapidly. Until now, no application in the micronization of traditional Chinese medicines is reported using this promising technology. In this work, Ginkgo leaves were used as the raw materials. In order to obtain Ginkgo flavonoid particles with a controlled size, we coupled the two techniques of SFE and SAA-HCM. The crude extracts of Ginkgo flavonoids were first extracted from Ginkgo biloba leaves using the SFE method, and the effects of important parameters on extraction yield were investigated. After the crude extracts were refined, the samples were treated with the techniques of SAA-HCM. The produced powders were characterized with respect to morphologies, particle size, and particle size distribution. Moreover, the influences of some process parameters on the particle size distribution were studied in detail.

10.1021/ie902001x  2010 American Chemical Society Published on Web 04/30/2010

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Figure 1. Schematic representation of supercritical fluid assisted atomization introduced by the hydrodynamic cavitation mixer process. H1, H2, H3: heating jacket; H4: nitrogen heater; C1, C2: check valve; Re: cooling bath; P1, P2: pumps; P3: vacuum pump; V1, V2, V3: valve; V: CO2 reservior; Mi: hydrodynamic cavitation mixer; N: nozzle; Pr: precipitator; Co: collector

2. Experimental Section 2.1. Materials. Ginkgo biloba leaves were obtained from Zhejiang Zhenyuan Share Co., Ltd. (Zhejiang, China), further ground by a disintegrator, and screened through an 80 mesh standard sieve. The total amount of flavonoids in 100 g of dried Ginkgo biloba leaves was 0.96 g. Rutin (98%) was purchased from Nanjing Tcm Institute of Chinese Materia Medica (Jiangsu, China). Carbon dioxide (99.9%) was obtained from Hangzhou Jingong Gas Co., Ltd. (Zhejiang, China). Ethanol, NaOH, NaNO2, and Al(NO3)3 were analytical reagents. All other chemicals used were of analytical grade without further purification. 2.2. Experimental Apparatus. SFE Apparatus. The SFE500R-2-FMC10 system (Thar, USA) had two feed lines: one for CO2 and the other for the entrainer (ethanol). CO2 was stored in a cylinder container and released into the system through a valve. The CO2 first entered into the cooling bath and was maintained at a certain temperature to facilitate a constant feeding rate of the pump. Then, CO2 was mixed with the entrainer in the mixer. The temperature of the compound was controlled by a heating jacket. Subsequently, the mixture of SCCO2 and the entrainer contacted the Ginkgo biloba leaves in the extraction vessel (500 mL), which was maintained at a constant temperature and pressure. The crude extracts were finally gathered in separation vessel (500 mL). All of the parameters were controlled with a programmed system. SAA-HCM Apparatus. The SAA-HCM experimental system9 is shown in Figure 1. It consisted of three feed lines to deliver supercritical CO2, the solution, and hot N2. The CO2 feed line was nearly the same as described above. The liquid solution was stored in a graduated cylinder, and the flow rate was regulated by a plunger pump (P2). The liquid solution was mixed with CO2 in the hydrodynamic cavitation mixer (Mi). The temperature in the Mi was controlled by a heating jacket (H2), and its pressure was maintained by P1. The mixture was then sprayed out through a nozzle with an inner diameter of 200 µm, and the spray pattern was stabilized by adjusting the valve (V3). The droplets formed were then sent to the precipitator (Pr). Meanwhile, N2 was taken from a cylinder container, heated in an electric heat exchanger (H4), and sent to the cylindrical precipitator. In the precipitator, the heating N2 dried

the fine droplets promptly to form dry particles. A vacuum pump (P3) is connected to the bottom of the precipitator in order to maintain a constant pressure. The dry particles were collected on a 0.5 µm pore size stainless steel membrane filter situated between the vacuum pump and the precipitator. 2.3. Purification of the Crude Extracts. The crude extracts obtained by SFE contained a large amount of impurities, such as chlorophyll, proteins, polysaccharide, etc. A primary purification had to be carried out before the extracts were micronized using the SAA-HCM process. The purification procedure was as follows: The crude extracts in the ethanol were first concentrated by vacuum distillation and then dissolved into a 20% (v/v) ethanol aqueous solution and filtered. The Ginkgo flavonoids in the filtrate were adsorbed by D101 resin and eluted with a 70% (v/v) ethanol aqueous solution. Then, elution was distilled under reduced pressure, and the purified extracts were obtained. 2.4. Assay of Ginkgo Flavonoids. The concentration of Ginkgo flavonoids in the extracts was measured by UV-vis spectrophotometry with a chromogenic system of NaNO2-Al(NO3)3-NaOH.13 Rutin was taken as the reference sample, and a standard curve was drawn: A510 ) 11.75C, where A510 denoted the absorbance at 510 nm and C was the concentration of rutin (g/L). The concentration of Ginkgo flavonoids can be determined by this curve. The extraction yield was estimated by the total amount of flavonoids in the crude extracts divided by the weight of dried Ginkgo biloba leaves. 2.5. Particle Characterization. The morphologies of Ginkgo flavonoid particles prepared by the SAA-HCM process were characterized with a scanning electron microscope (JSM-6390A, JEOL, Japan). Samples were deposited on carbon sticky tabs and coated with gold prior to analysis. The particle sizes and the size distribution were analyzed with the Nano Measurer 1.2 software, and more than 1000 particles were counted per calculation of particle size distribution. 3. Results and Discussion 3.1. Optimization of Ginkgo Flavonoids Extraction by SCF. Previous work5 indicated that it was hard to extract flavonoids from dried Ginkgo biloba leaves using SC-CO2 as the sole solvent, because the polarities of Ginkgo flavonoids were far different from that of SC-CO2, which resulted in a low solubility of Ginkgo flavonoids in SC-CO2. Usually, ethanol was added as an entrainer in the SFE process to enhance the solubility of polar Ginkgo flavonoids in SC-CO2. The effects of important parameters in supercritical fluid extraction including extraction pressure, extraction temperature, CO2 flow rate, and ethanol flow rate were examined in this experiment. The effects of extraction pressure on the Ginkgo flavonoid yield are shown in Figure 2. It can be seen that the extraction yield of Ginkgo flavonoids increased with an extraction pressure under 20 MPa. However, when the pressure was above 20 MPa, the extraction yield began to decrease. This result is in agreement with those of other researchers14,15 for the SC-CO2 extraction of natural products. Ginkgo flavonoid solubility depends on a complex interaction between supercritical fluid density and solute (extracted material) vapor pressure, both controlled by the temperature and pressure of the SC-CO2 and ethanol mixture. At a constant temperature, the density of the solvent increased with an increase in pressure, but the vapor pressure of the solute decreased with an increase in pressure. When the pressure elevated from 10 to 20 MPa, the density of SC-CO2 was dominant in this extraction process, resulting in a favorable extraction at lower than 20 MPa of pressure. However, when

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Figure 2. Effects of pressure on the extraction yield of Ginkgo flavonoids. (temperature 40 °C, CO2 flow rate 10 g/min, ethanol flow rate 2 mL/min)

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Figure 4. Effects of CO2 flow rate on extraction yield of ginkgo flavonoids (pressure 20 MPa, temperature 40 °C).

Figure 5. Effects of ethanol flow rate on extraction yield of Ginkgo flavonoids (pressure 20 MPa, temperature 40 °C, CO2 flow rate 10 g/min). Figure 3. Effects of temperature on extraction yield of Ginkgo flavonoids (pressure 20 MPa, CO2 flow rate 10 g/min, ethanol flow rate 2 mL/min).

the pressure increased further from 20 to 30 MPa, the vapor pressure of the solute overcame the relatively small change of solvent density, resulting in an unfavorable extraction at higher pressure.16 The effects of extraction temperature on Ginkgo flavonoid yield are presented in Figure 3. We can see that temperature significantly increased the extraction yield up to 40 °C, while a further increase in temperature above 40 °C makes the variation of yield complex. It was believed that increasing the temperature could intensify the mass transfer between SC-CO2 and powders of Ginkgo biloba leaves, but at the same time, it will also reduce the density of SC-CO2. It can be found in Figure 3 that the mass transfer between CO2 and Ginkgo leaves was a leading factor at a temperature range of 40 to 55 °C, while the density of SC-CO2 was more important in the ranges of 35 to 40 °C and 55 to 60 °C. The two factors interacted oppositely to make the influences of temperature on the extraction yield complicated. Figure 4 indicated the influence of the CO2 flow rate on the Ginkgo flavonoid extraction yield. When the extraction time

was fixed, the amount of extraction solvent volume increased with the CO2 flow rate, which resulted in a higher extraction yield. The optimal CO2 flow rate was 10 g/min, where the flow rate ratio of CO2 and ethanol was fixed at 5:3. As an entrainer of the SFE process, the ethanol flow rate had a remarkable effect on the extraction yield, as shown in Figure 5. On the basis of the principle of “similarity and intermiscibility”, an increase in the ethanol flow rate would improve the solubility of Ginkgo flavonoids in SC-CO2, since Ginkgo flavonoids were polar mixtures, while SC-CO2 was a kind of nonpolar solvent. When the ethanol flow rate reached 6 mL/ min, it seemed that the polarity between extraction solvent and Ginkgo flavonoids was very near. From the above experiments, the optimal experimental conditions were determined as follows: At a CO2 flow rate of 10 g/min, an ethanol flow rate of 6 mL/min, 20 MPa of extraction pressure, an extraction temperature of 40 °C, and after 90 min of extraction, the extraction yield of Ginkgo flavonoids could reach 0.36%. This result was comparable to that of Chiu et al.5 Since 100 g of dried Ginkgo biloba leaves contains 0.96 g of total flavonoids in the raw material, about 37.5% of the

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Figure 7. PSDs of micronized Ginkgo flavonoids at different R values based on particle number.

Figure 6. SEM images of Ginkgo flavonoids precipitated by SAA-HCM from ethanol with different R values (w/w) (A, 2.3; B, 1.8; C, 1.5).

Ginkgo flavonoids were extracted in one run using SCF from Ginkgo biloba leaves. After the crude extracts were refined, the purity of Ginkgo flavonoids in the extracts increased from 2.1% to 19.3%. 3.2. Preparation of Ginkgo Flavonoids Microparticles by SAA-HCM Process. In the SAA-HCM experiment, ethanol was used as a liquid solvent for Ginkgo flavonoid extracts. Operating parameters in the mixer were first optimized; two important process parameters, i.e., the concentration of the solution and the mass flow ratio CO2/liquid solution, were then explored, and their effects on particle morphologies, PS, and PSD were investigated. Selection of Mixer Operating Pressure and Temperature. Reverchon et al.17,18 proposed that the solubilization of SCCO2 in the liquid solution inside the mixer, which was determined by the high-pressure vapor liquid equilibria (VLEs)

of the system of liquid solvent/CO2, played a key role in the efficiency of the SAA-HCM. Hence, the temperature, the pressure in the mixer, and the mass flow ratio CO2/liquid solution are three important parameters that actually determined the operating point with respect to the VLE of the system liquid solvent/CO2. Although data on high-pressure VLEs for the binary systems of ethanol/CO2 were already available,19,20 the influence of Ginkgo flavonoids on the binary VLE was not easy to determine, as Ginkgo flavonoids were a mixture composed of three main aglycone derivatives (isorhamnetin, kaempferol, quercetin) and six biflavonones. On the basis of the above consideration, a semiempirical approach was carried out to find the suitable operating pressure and temperature in the mixer, in a range between 8 and 12 MPa and between 40 and 80 °C. The optimal SAA-HCM operating conditions were ascertained on the basis of spherical particle morphology, smaller particle size, narrower PSD, as well as the stability of the process. What’s more, the antioxidant activity of Ginkgo flavonoids should also be taken into account. Since a high temperature may increase the degradation rates of flavonoids, which subsequently attenuates the antioxidant activity of flavonoids.21 According to the work done by Cai et al.9,12 and the preliminary experiments performed in our lab, the process parameters were set at 10 MPa and 40 °C in this work. Effect of Mass Flow Ratio CO2/Liquid Solution (R). When the pressure and temperature are fixed, the mass flow ratio (R) determines the operating point of the process in the ternary VLE diagram of the Ginkgo flavonoids/CO2/liquid solvent system. Systematic experiments were performed in order to investigate the effects of the flow ratio CO2/liquid solution on the SAAHCM processed Ginkgo flavonoids. Figure 6 shows SEM images of processed drugs at different R values ranging between 1.5 and 2.3, when the Ginkgo flavonoid concentration (Csol) was fixed at 10 mg/mL. All three images indicated that the Ginkgo flavonoid microparticles were produced with the morphology of well-defined separated spheres. There was an increase in diameter of the particles when R values decreased. PSDs at different R values were calculated from SEM images and reported in Figure 7, in terms of the particle number. It can be seen from Figure 7 that the mode (the most frequent particle size) varied from 0.45 to 0.58 µm, and an enlargement of the distributions was also observed when the R value decreased.

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Figure 8. SEM images of Ginkgo flavonoids precipitated by SAA-HCM from ethanol with different solute concentrations (A, 5 mg/mL; B, 10 mg/mL; C, 20 mg/mL; D, 40 mg/mL).

(Csol) between 5 and 40 mg/mL, when R was 2.3. SEM images of the particles collected at different Csol’s are shown in Figure 8. It can be seen from Figure 8 that the morphologies of particles in the images were all well-defined, spherical, and separated. An increase of particle size with the increase of Csol can be observed. SEM images were analyzed to determine the PSDs and are reported in Figure 9. It can be seen from Figure 9 that the mode varied from 0.45 to 0.75 µm, and an enlargement of the distributions was also observed when the Csol was increased. The solute concentration had a large effect on particle size because the viscosity and surface tension of the liquid solution were increased with Csol, resulting in larger primary droplets and larger solid microparticles. 4. Conclusions

Figure 9. PSDs of micronized Ginkgo flavonoids at different Csol values based on particle number.

The effect of R values on PS and PSD can be explained in terms of the molar fraction of CO2 dissolved in the liquid solution in the mixer. During the SAA-HCM process, all feed CO2 was supposed to dissolve in the liquid solution. When R values increased, the quantities of CO2 dissolved in the liquid phase became greater, which resulted in a stronger decompressive atomization, and the microparticles produced become smaller.17 Effect of Solute Concentration. A group of experiments were performed with different Ginkgo flavonoid concentrations

Ginkgo flavonoids were extracted from Ginkgo biloba leaves by SCF, and the influences of process parameters on the extraction yield were investigated. The extraction pressure and temperature had great impacts on the extraction yield. The optimal experimental conditions were found: at a CO2 flow rate of 10 g/min, an ethanol flow rate of 6 mL/min, a pressure of 20 MPa, a temperature of 40 °C, and 90 min of extraction, the yield could reach 0.36%. The SAA-HCM process is very promising in producing micronic and submicronic particles of controlled diameters. Ginko flavonoid particles were successfully produced by SAAHCM with a controlled PS and PSD ranging between 0.2 and 3 µm. The morphologies, PS, and PSD of Ginkgo flavonoid microparticles showed a dependence on the solute concentration as well as the mass flow ratio R. Thus, tailoring of particle size and particle size distribution can be easily obtained varying the

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process parameters of solution concentration and mass flow ratio, to produce micro- or nano- sized particles. The Ginkgo flavonoid microparticles obtained by the SAA-HCM process have the potential to intensify the dissolution rates into biological environments and improve the absorption of Ginkgo flavonoids into the body. Acknowledgment The authors are grateful for financial support provided by the National Natural Science Foundation of China (20676118). Literature Cited (1) Maurer, K.; Ihl, R.; Dierks, T.; Fro¨lich, L. Clinical efficacy of Ginkgo biloba special extract EGb 761 in dementia of the Alzheimer type. J. Psychiatr. Res. 1997, 31, 645–655. (2) Oyama, Y.; Chikahisa, L.; Ueha, T.; Kanemaru, K.; Noda, K. Ginkgo biloba extract protects brain neurons against oxidative stress induced by hydrogen peroxide. Brain Res. 1996, 712, 349–352. (3) Van Beek, T. A. Chemical analysis of Ginkgo biloba leaves and extracts. J. Chromatogr. A 2002, 967, 21–55. (4) Harborne, J. B.; Williams, C. A. Advances in flavonoid research since 1992. Phytochemistry 2000, 55, 481–504. (5) Chiu, K. L.; Cheng, Y. C.; Chen, J. H.; Chang, C. J.; Yang, P. W. Supercritical fluids extraction of Ginkgo Ginkgolides and flavonoids. J. Supercrit. Fluids 2002, 24, 77–87. (6) Hollman, P.; Katan, M. Absorption, metabolism and health effects of dietary flavonoids in man. Biomed. Pharmacother. 1997, 51, 305–310. (7) Charoenchaitrakool, M.; Dehghani, F.; Foster, N. R. Micronization by rapid expansion of supercritical solutions to enhance the dissolution rates of poorly water-soluble pharmaceuticals. Ind. Eng. Chem. Res. 2000, 39, 4794–4802. (8) Garner, R. C.; Garner, J. V.; Gregory, S.; Whattam, M.; Calam, A.; Leong, D. Comparison of the absorption of micronized (Daflon 500 mg) and nonmicronized 14C-Diosmin tablets after oral administration to healthy volunteers by accelerator mass spectrometry and liquid scintillation counting. J. Pharm. Sci. 2002, 91, 32–40. (9) Cai, M. Q.; Guan, Y. X.; Yao, S. J.; Zhu, Z. Q. Supercritical fluid assisted atomization introduced by hydrodynamic cavitation mixer (SAAHCM) for micronization of levofloxacin hydrochloride. J. Supercrit. Fluids 2008, 43, 524–534.

(10) Reverchon, E.; Antonacci, A. Polymer microparticles production by supercritical assisted atomization. J. Supercrit. Fluids 2007, 39, 444– 452. (11) Porta, G. D.; Vittori, C. D.; Reverchon, E. Supercritical assisted atomization: A novel technology for microparticles preparation of an asthmacontrolling drug. AAPS PharmSciTech 2005, 6, 421–428. (12) Cai, M. Q.; Guan, Y. X.; Yao, S. J.; Zhu, Z. Q. Supercritical fluid assisted atomization introduced by hydrodynamic cavitation mixer for micronization of roxithromycin. CIESC J. 2008, 59, 293–300. (13) Zhu, H. B.; Wang, Y. Z.; Liu, Y. X.; Xia, Y. L. Analysis of flavonoids in Portulaca oleracea L. by UV-Vis spectrophotometry with comparative study on different extraction technologies. Food Anal. Meth. 2009, DOI: 10.1007/s12161-009-9091-2. ¨ .; C¸alımlı, A. Determi(14) S¸anal, I˙.S.; Bayraktar, E.; Mehmetog˘lu, U nation of optimum conditions for SC-(CO2 + ethanol) extraction of β-carotene from apricot pomace using response surface methodology. J. Supercrit. Fluids 2005, 34, 331–338. (15) Machmudah, S.; Shotipruk, A.; Goto, M.; Sasaki, M.; Hirose, T. Extraction of Astaxanthin from Haematococcus pluvialis using supercritical CO2 and ethanol as entrainer. Ind. Eng. Chem. Res. 2006, 45, 3652–3657. (16) Roy, B. C.; Goto, M.; Hirose, T. Extraction of ginger oil with supercritical carbon dioxide: experiments and modeling. Ind. Eng. Chem. Res. 1996, 35, 607–612. (17) Reverchon, E.; Antonacci, A. Polymer microparticles production by supercritical assisted atomization. J. Supercrit. Fluids 2007, 39, 444– 452. (18) Reverchon, E.; Porta, G. D. Micronization of antibiotics by supercritical assisted atomization. J. Supercrit. Fluids 2003, 26, 243–252. (19) Jennlngs, D. W.; Lee, R. J.; Teja, A. S. Vapor-liquid equilibria in the carbon dioxide + ethanol and carbon dioxide + l-butanol systems. J. Chem. Eng. Data 1991, 36, 303–307. (20) Suzuki, K.; Sue, H. Isothermal vapor-liquid equilibrium data for binary systems at high pressures: carbon dioxide-methanol, carbon dioxideethanol, carbon dioxide-1-propanol, methane-ethanol, methane-1-propanol, ethane-ethanol, and ethane-1-propanol systems. J. Chem. Eng. Data 1990, 35, 63–66. (21) Runha, F. P.; Cordeiro, D. S.; Pereira, C. A. M.; Vilegas, J.; Oliveira, W. P. Production of dry extracts of medicinal brazilian plants by apouted bed process: Development of the process and evaluation of thermal degradation during the drying operation. Trans. IchemE. 2001, 79, 160– 167.

ReceiVed for reView December 17, 2009 ReVised manuscript receiVed April 16, 2010 Accepted April 21, 2010 IE902001X