Aqueous Enzymatic Extraction Process Research Articles

Improving enzyme accessibility in the aqueous enzymatic extraction process by microwave-induced porous cell walls to increase oil body and protein yields

Aqueous enzymatic extraction (AEE) is a green and sustainable extraction method. However, the intact cell wall hinders the accessibility of enzymes, severely limiting the efficiency and application of the AEE method. In this study, microwave-assisted aqueous enzymatic extraction (MAAEE) was used to degrade the peanut cell wall and improve the yield of oil and protein. Transmission electron microscopy, scanning electron microscopy, BET-specific surface area, Fourier transform infrared spectroscopy, X-ray diffraction, and chemical composition analysis were used to characterize the structure and chemical changes of the peanut cell wall. The results showed that microwave treatment could break the structure of the peanut cell wall, increase the porosity of raw materials, accelerate the efficiency of enzymatic hydrolysis, and promote the extraction of oil and protein. Compared with raw peanut powder (MC-0), microwaved peanut powder (MC-4) had a higher oil body yield (>44%) and protein release rate (>76%). At the microscopic level, cell wall rupture, oil accumulation, and cell contents scattered outside the cell were observed. The mean pore size, pore volume, and specific surface area increased by 54.1%, 149.4%, and 63.5%, respectively, while the crystallinity decreased by 4.4%. The content of hemicellulose and CDTA-soluble pectin (CSP) decreased by 18.6% and 32.9%, respectively. The degradation rates of cellulase and pectinase on cell wall components were significantly increased by 8.5%–43.2% (p < 0.05). In summary, microwave-induced porous cell walls could significantly improve enzyme accessibility, accelerate enzymatic hydrolysis efficiency, and promote oil and protein extraction.

Demulsification of Emulsion Using Heptanoic Acid during Aqueous Enzymatic Extraction and the Characterization of Peanut Oil and Proteins Extracted.

Peanut oil body emulsion occurs during the process of aqueous enzymatic extraction (AEE). The free oil is difficult to release and extract because its structure is stable and not easily destroyed. Demulsification can release free oil in an oil body emulsion, so various fatty acids were selected for the demulsification. Changes in the amount of heptanoic acid added, solid-liquid ratio, reaction temperature, and reaction time were adopted to investigate demulsification, and the technological conditions of demulsification were optimized. While the optimal conditions were the addition of 1.26% of heptanoic acid, solid-liquid ratio of 1:3.25, reaction temperature of 72.7 °C, and reaction time of 55 min, the maximum free oil yield was (95.84 ± 0.19)%. The analysis of the fatty acid composition and physicochemical characterization of peanut oils extracted using four methods were studied during the AEE process. Compared with the amount of oil extracted via other methods, the unsaturated fatty acids of oils extracted from demulsification with heptanoic acid contained 78.81%, which was significantly higher than the other three methods. The results of physicochemical characterization indicated that the oil obtained by demulsification with heptanoic acid had a higher quality. According to the analysis of the amino acid composition, the protein obtained using AEE was similar to that of commercial peanut protein powder (CPPP). However, the essential amino acid content of proteins extracted via AEE was significantly higher than that of CPPP. The capacity of water (oil) holding, emulsifying activity, and foaming properties of protein obtained via AEE were better than those for CPPP. Overall, heptanoic acid demulsification is a potential demulsification method, thus, this work provides a new idea for the industrial application of simultaneous separation of oil and proteins via AEE.

Pumpkin seed coat pigments affected aqueous enzymatic extraction processing through interaction with its interfacial protein

In this study, the oil recovery from pumpkin seed with or without the presence of pigment was evaluated during the aqueous enzymatic extraction process (AEEP). Additionally, the interaction of pumpkin seed coat pigment—protochlorophyll (Pchl) with interfacial proteins (globulin-O/W, globulin-W, and albumin-W) during the aqueous extraction process was investigated. When Pchl was removed from pumpkin seeds, the yield of free oil was significantly increased by 50.71% after the first centrifugation. And the total oil recovery increased from 68.59% to 89.91% using enzymatic demulsification. The β-sheet percentage of globulin-O/W decreased by 5.6% with the presence of Pchl based on the circular dichroism spectra analysis. The particle size of globulin-W and albumin-W significantly increased in contrast to globulin-O/W. In addition, the increase of surface hydrophobicity and surface sulfhydryl group showed a conformational change of three protein extracts. Fluorescence spectra analyses showed that Pchl quenched the intrinsic fluorescence of globulin-W, albumin-W, and globulin-O/W in a static mode. Thermodynamic analyses indicated that the main force between Pchl and proteins was hydrophobic interactions. The hydrolysis efficiency of Papain was reduced during demulsification in AEEP with the presence of Pchl. Therefore, the removal of pumpkin seed coat pigments facilitated the aqueous extraction of seed oil by getting rid of the interaction of Pchl with its interfacial protein.

Study on Aqueous Enzymatic Extraction Process of Common Pistache Oil

The aqueous enzymatic method was applied to extract the common pistache oil and the optimal extraction process conditions were identified. By observing the effect of enzymatic hydrolysis time, pH value, temperature on aqueous enzymatic extraction process and performing the orthogonal experiment based on the single factor test, the optimal process parameters were obtained, namely, the optimal time, temperature, and pH value were respectively 3 h, 50℃, and 7 with a final extraction rate of 25.38 %.

EXTRACTION AND EVALUATION OF EDIBLE OIL FROM SCHIZOCHYTRIUM SP. USING AN AQUEOUS ENZYMATIC METHOD

<List> <ListItem><ItemContent> • Aqueous enzymatic extraction (AEE) is performed for oil extraction from <i>Schizochytrium</i> sp. </ItemContent></ListItem> <ListItem><ItemContent> • AEE process is optimized by response surface methodology. </ItemContent></ListItem> <ListItem><ItemContent> • Microalgal oil extracted by AEE has high contents of PUFA, tocopherols and phenolics. </ItemContent></ListItem> <ListItem><ItemContent> • AEEO exhibits considerable antioxidant activity as compared with SEO. </ItemContent></ListItem></List> <i>Schizochytrium</i> sp., a marine microalga, is a potential source of edible oil due to its short growth cycle and rapid lipid accumulation, especially of docosahexaenoic acid. An approach to isolate edible microalgal oil from <i>Schizochytrium</i> sp. using aqueous enzymatic extraction (AEE) was developed. Parameters were optimized by single-factor experiments followed by Box-Behnken design. Proteases were effective in extracting oil. The maximum free oil recovery (49.7%±0.58%) and total oil recovery (68.1%±0.94%) were obtained under optimum conditions of liquid-to-solid ratio of 4.8:1, a 2.5% enzyme concentration of papain and an extraction time of 2.2 h. There was a significant difference (<i>P</i><0.05) in polyunsaturated fatty acid composition between microalgal oil obtained by AEE and by Soxhlet extraction, with the former having superior physiochemical properties and higher concentrations of bioactive components including total phenolic compounds and total tocopherols. These findings indicate a potential application of AEE for extraction of oil from <i>Schizochytrium</i> sp.

Aqueous enzymatic extraction of polyunsaturated fatty acid–rich sacha inchi (Plukenetia volubilis L.) seed oil: An eco-friendly approach

Sacha inchi seeds are a potential source of natural oil rich in essential polyunsaturated fatty acids and have various nutraceutical and health benefits. This study developed an aqueous enzymatic extraction process for eco-friendly extracting sacha inchi seed oil. The activities of different enzymes (proteases and cellulase) in the extraction process were evaluated. Among the tested enzymes, papain exhibited the highest extraction activity. The papain-assisted extraction was subsequently optimized through response surface methodology to yield maximum oil level. A maximum oil yield of 28.45% was obtained under the following extraction conditions: enzyme loading, 4.46%; water-to-sample ratio, 4.45 mL/g; extraction time, 4.95 h; and temperature, 38.9 °C. Under optimal extraction conditions, papain could be repeatedly used for extraction. This study indicates that papain is a potential protease and that aqueous enzymatic extraction is an eco-friendly and efficient process for extracting beneficial oil from sacha inchi seeds.

Physicochemical Properties and Stabilities of Crude and Purified Oil Bodies Extracted from High Oleic Peanuts

AbstractPhysicochemical properties and emulsion stabilities of two types of oil bodies—crude oil bodies (COB) and purified oil bodies (OB)—recovered from high oleic peanuts via an aqueous enzymatic extraction process are investigated. RGB color analysis and flavor volatiles identification reveal that both COB and OB present desirable color and acceptable flavor. OB tends to have stronger antioxidant capacity and higher nutritional value compared to COB according to results from fatty acids, amino acids, and tocopherol analyses. The emulsification of oil bodies shows a slightly lower level than lecithin, but excellent thermal stability and storage stability (especially for OB), which relates greatly to small particle size, high zeta‐potential, and high content of hydrophobic amino acids. The freeze–thaw stability of OB is poorer than that of COB, which may relate to the presence of impurities in oil bodies. Thus, the peanut oil bodies, especially purified oil bodies, are proven to be advantageous emulsifying agents and additional valuable sources of antioxidants.Practical Applications: The results in this work clarify the differences in physiochemical composition and the fundamental emulsifying properties between COB and OB, and demonstrate the possibility of replacing traditional emulsifiers by natural green emulsifiers (e.g., oil bodies) in the food industry. Moreover, oil bodies extracted from high oleic peanuts, which contain plenty of unsaturated fatty acids and tocopherols, are a potential source for food supplements. In the long term, this work will increase the industrial conversion rate of oil bodies, enhance the processing and utilization level of high oleic peanuts, and promote the development of new foods based on peanut resources.

Study on Extraction of Peanut Protein and Oil Bodies by Aqueous Enzymatic Extraction and Characterization of Protein

Cell wall degrading enzymes break down the cell wall by degrading the main cell wall components and destroying structure of the cell wall without influencing the protein. Effects of various enzymes (Viscozyme® L, cellulase, hemicellulase, and pectinase) on the molecular weight distribution of peanut protein and yield of peanut protein and oil bodies during an aqueous enzymatic extraction process were investigated in this study. The molecular weight distribution of peanut protein was not changed, and Viscozyme® L was selected to assist peanut protein and oil bodies extraction by the aqueous extraction process. The aqueous enzymatic extraction process was optimized by a signal factor experiment and response surface methodology, and the optimal condition was enzyme hydrolysis temperature of 52°C, solid-liquid ratio of 1 : 4, enzyme concentration of 1.35%, and enzyme hydrolysis time of 90 min. A peanut protein yield of 78.60% and oil bodies yield of 48.44% were achieved under the optimal condition. Compared with commercial peanut protein powder (CPPP), the solubility and foaming properties of peanut protein powder obtained by aqueous enzymatic extraction (AEEPPP) were a little lower. However, the functional properties of foam stability, emulsifying activity, emulsifying stability, water holding capacity, and oil holding capacity of AEEPPP were better than that of CPPP.

The Mechanism of Extraction of Peanut Protein and Oil Bodies by Enzymatic Hydrolysis of the Cell Wall.

Degradation of the peanut cell wall is a critical step in the aqueous enzymatic extraction process to extract proteins and oil bodies. Viscozyme® L, a compound cell wall degrading enzyme, has been applied as an alternative to protease in the process of aqueous enzymatic extraction, but the mechanism of cell wall enzymolysis remains unclear. The present study aims to investigate the changes in cellulose, hemicellulose, and pectin content of the peanut cell wall hydrolyzed by Viscozyme® L. The degree to which the main components of the peanut cell wall, such as trans-1, 2-cyclohexanediamine-N,N,N',N'-acetic acid-soluble pectin (CDTA-soluble pectin), Na2CO3-soluble pectin, cellulose, and hemicellulose, are degraded is closely related to the extraction of oil bodies and peanut protein at different solid-liquid ratio of powered peanut seed in distilled water, enzyme concentration, enzyme hydrolysis temperature, and enzyme hydrolysis time. The key sites of Viscozyme® L activity on cell wall polysaccharides were explored by comparing the changes in chemical bonds under different extraction conditions using Fourier-transform infrared spectroscopy (FT-IR) absorption bands and principal component analysis (PCA). Viscozyme® L acted on the C-O stretching, C-C stretching, and CH2 symmetrical bending of cellulose, the C-O stretching and O-C-O asymmetrical bending of hemicellulose, and the C-O stretching and C-C stretching of pectin.

Rapid Salt‐Assisted Microwave Demulsification of Oil‐Rich Emulsion Obtained by Aqueous Enzymatic Extraction of Peanut Seeds

AbstractAqueous enzymatic extraction (AEE) is an environmentally friendly edible‐oil‐extraction process that can also provide edible protein. However, the AEE process may form a stable emulsion in most cases, which seriously limits the large‐scale industry applications for producing vegetable oils. In this study, the salt‐assisted microwave radiation demulsification of the oil‐rich emulsion prepared with AEE from peanuts is investigated. The microwave demulsification method is compared with other conventional demulsification methods, including heating, and freezing–thawing. The salt‐assisted microwave demulsification of the emulsions shows a greater free oil yield than conventional heating demulsification. Moreover, the microwave demulsification shows a similar free oil yield in less time than freezing–thawing method. Under the optimal operating conditions of demulsification, the free oil yield can reach 92.3% with CaCl2‐assisted microwave demulsification for only 2 min. In addition, the oxidative properties and the fatty acid compositions of the demulsified peanut oil are investigated. No significant difference in the fatty acid composition is observed among salt‐assisted microwave, freezing–thawing, and heating demulsified oil. The oxidative properties of the salt‐assisted microwave demulsified peanut oil is better than the conventional heating demulsified oil. Thus, salt‐assisted microwave demulsification provides a quick and effective demulsification method to obtain vegetable oils with high quality.Practical Applications: Aqueous enzymatic extraction (AEE) is an environmentally friendly edible‐oil‐extraction process. To solve the problem of stable emulsion formed during AEE process, the salt‐assisted microwave demulsification of the oil‐rich emulsion prepared with AEE is developed with high efficiency (demulsification for 2 min). In addition, the oxidative properties of the microwave demulsified oil is better than the conventional heating demulsified oil.

Efficient and Response Surface Optimized Aqueous Enzymatic Extraction of Camellia oleifera (Tea Seed) Oil Facilitated by Concurrent Calcium Chloride Addition

AbstractThis paper reports an efficient aqueous enzymatic extraction (AEE) method for Camellia oleifera seed oil with the aid of response surface analysis. A maximum oil recovery of ~93.5% was obtained when a 2‐step AEE process was performed using 0.80% cellulase (v/w) solution at pH 6.0 maintained at 50 °C for 1 h followed by a solution of 0.70% Alcalase® with pH 9.2 at 57 °C for 4.1 h. It was found that the addition of Ca2+ during the proteolysis stage improved the free oil yield from ~62.1 to ~86.6%. This was attributed to the removal of tea saponins, cross‐linkage of anionic polysaccharides, and destabilization of cream emulsion by Ca2+. This was verified by decreased tea saponin and polysaccharide levels in the cream emulsion and bulk solution as well as lowering of the emulsion fraction. It was determined that addition of CaCl2 solution in continuous flow to the proteolysate is superior to one‐time or batch addition in inhibiting emulsion formation. The addition of CaCl2 may provide a means of replacing the more laborious, time‐consuming demulsification process otherwise required.

High pressure pre-treatment of Moringa oleifera seed kernels prior to aqueous enzymatic oil extraction

Abstract High pressure processing (HPP) was applied as a pre-treatment on Moringa oleifera (MO) kernels, for the first time, prior to aqueous enzymatic extraction (AEE) of the MO oil, and the effect of this pre-treatment is reported in terms of the free oil recovery and the nature of the cream emulsions formed. The HPP pre-treatments (50–250 MPa, 20–60 °C, 10–60 min) generally resulted in higher free oil recoveries and thinner emulsion layers from ground-sieved kernels than the whole kernels. Optimization of the HPP parameters indicated linear increment in free oil recovery with increase in temperature and time, but not the pressure level. Without the pre-boiling step in the AEE process, the use of HPP pre-treatment at 50 MPa and 60 °C for 35 min resulted in approximately 73% (w/w) free oil recovery with thinner emulsion layer than the use of AEE alone. These findings highlighted the ability of HPP in altering the MO protein structure into a form of less emulsifying functional properties, thus further de-emulsification method may not be necessary. Industrial relevance High pressure processing (HPP) applications are increasing in the food industry. High pressure application allows the use of relatively lower temperatures in processes, in order to achieve comparable outcomes of higher temperature processing. This study explores the application of HPP on Moringa oleifera (MO) kernels prior to aqueous enzymatic extraction (AEE) of oil. In general, the main disadvantage of AEE is its lower oil recovery in comparison with solvent extraction, which is attributed to the formation of a relatively stable cream emulsion after extraction. This study shows that the thickness of the creamy emulsion can be significantly reduced, and oil recoveries improved, by subjecting the kernels to HPP prior to extraction. The improvement in recoveries did not depend strongly on the level of high pressure applied, but downstream processes for free oil separation and recovery were considerably simplified.

Aqueous enzymatic extraction of Moringa oleifera oil

This paper reports on the extraction of Moringa oleifera (MO) oil by using aqueous enzymatic extraction (AEE) method. The effect of different process parameters on the oil recovery was discovered by using statistical optimization, besides the effect of selected parameters on the formation of its oil-in-water cream emulsions. Within the pre-determined ranges, the use of pH 4.5, moisture/kernel ratio of 8:1 (w/w), and 300stroke/min shaking speed at 40°C for 1h incubation time resulted in highest oil recovery of approximately 70% (goil/g solvent-extracted oil). These optimized parameters also result in a very thin emulsion layer, indicating minute amount of emulsion formed. Zero oil recovery with thick emulsion were observed when the used aqueous phase was re-utilized for another AEE process. The findings suggest that the critical selection of AEE parameters is key to high oil recovery with minimum emulsion formation thereby lowering the load on the de-emulsification step.

Optimization of Viscozyme-L assisted extraction of coconut milk and virgin coconut oil

The present work aimed to optimize the extraction of milk and oil from coconut by an aqueous enzymatic extraction process using the enzyme Viscozyme-L. The combine effect of grinding time (24min), amount of Viscozyme-L (60-180FBGU) and incubation time (2-7h), were optimized using Response Surface Methodology. The selected method provides the best yield of coconut milk (73.88± 0.5%, w/w) and Virgin Coconut Oil (21.57± 0.4%, w/w) with percentage recovery of oil was 86.14% of fat present in the fresh coconut kernel at grinding time (3min), the amount of enzyme (120FBGU) and incubation time (4.5h). The experimental results were significantly (P< 0.05) comparable to the predicted yield of coconut milk (73.33%) and oil (21.12%). A validation model that performed within the selected range of variation confirmed these results, microscopic image of cell wall and particle distribution in coconut milk, suggested that the use of the Viscozyme-L treatment could enhance the extraction of oil.

Aqueous Enzymatic Extraction of Rapeseed Oil and Protein from Dehulled Cold-pressed Double-low Rapeseed Cake

Abstract The oil and protein of dehulled cold-pressed double-low rapeseed cake was extracted by an aqueous enzymatic process. The rapeseed cake was treated by the chosen combined enzymes of Viscozyme L and Alcalase 2.4L (VLA,1:1,w/w). Preliminary experiments and Response Surface Methodology (RSM) were used to study the effects of enzyme concentration, incubation time and water-to-cake ratio on the extraction yield of oil and protein. This is how the desirable conditions were obtained. Transmissive electron microscope photo showed that after cold-pressing the cell structure of rapeseed was partly damaged while dehulling had little effect on the cell structure of rapeseed. In RSM experiments water-to-cake ratio showed significant effects on the extraction of oil and protein (P&lt;0.05),while incubation time only showed significant effects on protein yield (P&lt;0.05).The desirable conditions were as follows: 1.0% concentration (w/w) of VLA; water-to-cake ratio(w/w),6:1; 80 min incubation time. Under this condition, the extraction yield of protein and oil were 82.10% and 71.89%, respectively. Through combining both the cold-press and the aqueous enzymatic processes together, the total oil yield reached 91.6%, which is higher than the normal cold-press process or the aqueous enzymatic extraction process alone.

Aqueous Enzymatic Extraction of Oil and Protein Hydrolysates from Peanut

Aqueous enzymatic extraction (AEE) is a safe and efficient vegetable-oil extraction process that may also result in edible protein hydrolysates. In this study, an AEE process was developed to recover oil and protein hydrolysates from blanched peanut. The enzyme type and enzyme concentration for the most efficient extraction were selected. Under the condition of pH8.50, 60°C, an enzyme level of 1.5% and 8 h incubation, peanut protein hydrolysates (PPH) yield of 82.5% and oil yield of 92.2% were achieved when using alcalase2.4L. The fatty acid composition of oil obtained by AEE vs. hexane-extraction was very similar. The good oil quality could save the refine cost for edible. The functional properties of PPH were first investigated and they showed good effects of scavenging DPPH (α- dipheny -β- picrylhydrazyl) free radical and inhibiting the angiotensin-I-convertine enzyme (ACE).

Downstream Processes for Aqueous Enzymatic Extraction of Rapeseed Oil and Protein Hydrolysates

AbstractDownstream processes following aqueous enzymatic extraction (AEE) of rapeseed oil and protein hydrolysates were developed to enhance the oil and protein yields as well as to purify the protein hydrolysates. The wet precipitate (meal residue) from the AEE was washed with twofold water at 60 °C, pH 11 for 1 h. Emulsions from the AEE and the washing step were pooled and submitted to a stepwise demulsification procedure consisting of storage‐centrifugation and freezing–thawing followed by centrifugation. Aqueous phases were pooled and adsorbed onto macroporous adsorption resins (MAR) to remove salts and sugars. Following extensive rinsing with deionized water (pH 4), desorption was achieved by washing with 85% ethanol (v/v) to obtain crude rapeseed peptides (CRPs). In a separate experiment, stepwise desorption was carried out with 25, 55, and 85% ethanol to separate the bitter peptides from the other peptides. Using a combination of the AEE process, washing and demulsification steps, the yields of the total free oil and protein hydrolysates were 88–90% and 94–97%, respectively. The protein recovery was 66.7% and the protein content was enriched from 47.04 to 73.51% in the CRPs. No glucosinolates and phytic acid were detected in the CRPs. From the stepwise desorption, a non‐bitter fraction RP25 (containing 64–66% of total desorbed protein) had a bland color and significantly higher protein content (81.04%) and hence was the more desirable product.

Enzymatic treatment of rice bran to improve processing

AbstractA modification of the process of oil extraction from rice bran is proposed, introducing one or two enzymatic reactions previous to solvent extraction. Although a total aqueous enzymatic extraction process did not result in reasonable oil extraction yields, an interesting alternative results from enzymatic reactions previous to solvent extraction or pressing. A thermal treatment of rice bran is first applied to deactivate lipase, but also to gelatinize starch previous to reaction with α‐amylase. This is followed by a saccharifying step with glucoamylase to produce glucose (28 g/100 g of rice bran treated), while the residual paste, 66.7% of the original bran, may be subjected to a proteolytic process for protein extraction or directly treated with the solvent to obtain bran oil. Finally, under the defined extraction conditions using hexane, yields of oil are 5% higher when rice bran has been previously treated with α‐amylase.