Effect of Flow Rate on the Extraction of Volatile Concentrates and Resinoid Compounds from Origanum vulgare L. ssp. virens (Hoffm. et Link) letswaart using Compressed CO2
Extraction using compressed fluids, particularly carbon dioxide, has become an established method for separating volatile concentrates from plants. It is often regarded as an alternative to the more widely used and traditional techniques of steam distillation and organic solvent extraction, though the initial high capital costs often place it beyond the financial means of most processors. A supercritical extract from an aromatic herb is comprised of the volatile concentrate (similar in composition to the essential oil obtained from distillation methods) together with other co-extracted resinoid materials (often referred to as cuticular waxes). This extract generally depends on the herb material and on the process parameters, such as applied pressure and temperature. The attractiveness of a supercritical fluid as a solvent lies in its unique properties: liquid-like densities, high rates of mass transfer and a tunable selectivity, which arises from varying the density by changes in temperature and pressure. In comparison to the traditional techniques, a supercritical extract has a fingerprint closer to that of the natural source (1,2) and is not contaminated with solvent residue.
The effect of the supercritical fluid flow rate on the extraction rates from a plant material can be used to determine whether an extraction is limited primarily by solubility and chromatographic retention of the solute or by the rate of transport of solutes from the matrix to the bulk of the extraction fluid (3). In the first case, the increase of flow rate would result in a clear increase of the extraction rate. Moreover, if the extraction rate is controlled only by solubility, the extraction rate is directly proportional to the solvent flow rate. On the other hand, if intraparticle resistances are the only restriction, the extraction rate is controlled by the kinetics of the intraparticle transport, and consequently, not dependent on the solvent flow rate. Supercritical fluid extraction of volatile concentrates from herbaceous matrices have been reported as being limited by intraparticle resistances [see, for example, Naik et al. (4), Goto et al. (5), Reverchon et al. (6), Reverchon (7), Catchpole and Grey (8)]. However, an efficient pre-treatment of herbaceous matrices is required if acceptable yields and rates of extraction of the volatile concentrates are to be obtained. Communition of an herbaceous matrix prior to extraction has been shown to reduce dramatically the internal resistances; this affects the extraction of volatile concentrates by increasing the amount of solutes accessible to the solvent (9). A significant effect of solvent flow rate on the extraction of the resinoid compounds present within the herb is also to be expected. Resinoid components are present at the surface of the herb matrix, and therefore, their extraction is likely to be controlled by solubility and/or film resistance.
The effect of solvent flow rate on the extraction of volatile concentrates and resinoid compounds from a selected herb was studied under two different conditions, using liquid CO2 at 70 bar and 300 K and supercritical CO2 at 100 bar and 310 K. These two selected conditions also presented an opportunity to illustrate the effect of CO2 state at similar densities (712 and 689 kg/m3 for liquid CO2 and supercritical CO2, respectively) on the extraction rates and yields. The herb used in the present work was Origanum vulgare L. ssp.virens (Hoffm. et Link) letsioaart. Essential oils produced by several species of the Labiatae (L.) family constitute an important group of economic plant products, though this actual species has not achieved economic importance. The herb was selected on grounds of its supply cost and the importance of its family as a whole. Experimental tests were undertaken on a communited sample of the herb with an average bract size of 0.36 mm.
Preparation of the herb matrix: The wild herb was collected after the flowering period (late July) from Serra d’Arrabida region (central west Portugal). The sun-dried herbs arrived as intact plants, and a batch was prepared by removing the bracts by hand. Care was taken to discard any stalk-like material from the sample batch. Typically, 500 g of material was prepared for a batch and, after sieving, a mean bract size of 1.55 mm was obtained. The bracts were then comminuted to 0.36 mm in a commercial blender. A prepared batch was kept in an air-tight re-sealable polypropylene bag and stored at 4°C. The essential oil content of a batch was determined prior to compressed CO2 extraction by hydrodistillation, and the composition of the essential oil was analyzed by GC. The hydrodistillation equipment and procedure were based on those detailed in the European Pharmacopoeia (10). An oil yield of 0.67% (w/w) with a standard deviation of 0.04% was obtained. (Main essential oil content: γ-terpinene 20.5%, thymol 12.2%, carvacrol 10.3%, p-cymene 9.3%, linalool 7.2%, α-teqjineol 6.4%, α-pinene 3.8%, β-caryophyllene 4.4%, carophyllene oxide 2.8%.)
GC analysis: The oils and extracts collected from both extraction techniques were analyzed using an Ai Cambridge GC94 gas Chromatograph equipped with an FID detector. A DB-5 capillary column (30 m × 0.32 mm (5%-phenyl)-methylpolysiloxane, film thickness 0.25 µm), supplied by J & W Scientific, was used. The GC conditions were as follows: injection and detector temperature 270°C; oven temperature 65°C for 10 min, then 65°-150°C at 5°C/min ramp; carrier gas: helium at 1.7 mL/min; make-up gas: nitrogen at 12.3 mL/min; split flow at 11 mL/min. The oil and volatile concentrates extracted were identified by comparing their relative retention times with those of pure components obtained from Fluka. GC injection errors were minimized by comparing the peak area of an internal standard (limonene) to a predetermined set value and applying any necessary correction.
Results and Discussion
Effect of solvent flow rate on the extraction of the volatile concentrate: The extraction curves of the total volatile concentrates for both CO2 extraction states are shown in Figures 1 and 2 in terms of extraction time and mass of CO2 passed, respectively (L = liquid CO2, SC = supercritical CO2 and barg indicates bar gage pressure). Their extraction was mostly time dependent at the conditions tested, which can be attributed to the extraction still being limited by intraparticle resistances. In Figure 1 it can be seen that only in the earlier stages of extraction was there an increase of the extraction yields with the increase of flow rate. With liquid CO2, the extraction curves virtually converged after about 100 min, while with supercritical CO2 the convergence appeared earlier (at around 50 min). The slight discrepancies observed in the earlier stages of extraction are likely to have resulted from fluid phase retention in the bed (18.1 min for SC-CO2 and 18.7 min for L-CO2, at the lowest flow rate) and, in addition, from axial dispersion of the extracts throughout the bed, rather than from solubility effects.
The extraction degrees of the total volatile concentrates at 200 min ranged from 91-92% with liquid CO2 and from 92-93% with supercritical CO2. The effect of solvent state at similar densities was therefore negligible. Any difference in favor of the supercritical state was likely to have resulted from the slightly greater extent of the co-extraction of resinoid compounds, which would have made the volatile concentrates more accessible to extraction. The extraction of the last fraction of volatile concentrates was likely to have occurred in parallel to that of the resinoid compounds (12).
Effect of solvent flow rate on the extraction of resinoid materials: The resinoid extraction curves for both CO2 conditions are shown in Figures 3 and 4 in terms of extraction time and mass of CO2 passed, respectively. Unlike the volatile concentrates, their extraction was mostly dependent on the amount of CO2 used and supports the hypothesis that their extraction was limited by their solubility in compressed CO2. When comparing the yields obtained at the two CO2 conditions, it can be noted that slightly higher yields and extraction rates were obtained with supercritical CO2. The enhanced volatility of the resinoid components at 310 K was likely to be responsible for their increased solubility at supercritical conditions, despite the slightly lower density of the CO2 solvent (689 vs. 712kg/m^sup 3^).
Effect of solvent flow rate on the composition of the extract: The effect of solvent flow rate on the evolution of the extract composition for the two CO2 conditions is shown in Figure 5. It can be seen that as the extractions preceded and when the flow rate was increased, the concentrations of the volatile concentrate in the extracts decreased. An extract with slightly higher volatiles was obtained with liquid CO2 and was due to a decrease in the co-extraction of the resinoid components. In light of this, the use of liquid CO2 at low flow rates seems favorable; however, at very low flow rates, higher retention times and a flow condition far away from plug flow (the ideal extraction situation) would exist, both of which are detrimental for the rate of removal of volatile concentrates from the extraction vessel.
In conclusion, a higher degree of extraction of volatile concentrates was obtained at the lower flow rates. In spite of the comminuted sample, the extraction of the volatiles was still controlled by intraparticle resistances. An extract slightly richer in volatile concentrates was collected using liquid CO2. The use of liquid CO2 therefore seems more economically favorable in obtaining a product rich in volatile concentrates. The extraction of resinoid compounds was mostly dependent on the mass of CO2 used, which is strong evidence of mass transfer affected by solubility.
The authors gratefully thank thefundingprovided by the PRAXIS XXI Program and the European Union Commission Directorate- General for Agriculture (Project No. AIR-CT-0818).
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