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

Downstream 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.     

水酶法提取大豆油

The procedure of enzymatic aqueous extraction of soybean oil was assessed when two-step controlled enzymatic hydrolysis was applied. With aqueous extraction of soybean oil-containing protein, the highest yield of oil was 96.1% at the optimized conditions studied. Soybean oil-containing protein was hydrolyzed and resulted in releasing part of oil. The separated protein that contained 40% oil was enriched due to its adsorption capacity of released oil, the average oil extraction yeild reached 93.5%. Then the high oil content protein was hydrolyzed again to release oil by enzyme, the oil extraction yeild was 80.4%. As a result, high quality of soybean oil was obtained and the content of total oil vield was 74.4%.     

Optimization of the Aqueous Enzymatic Extraction of Rapeseed Oil and Protein Hydrolysates

An aqueous enzymatic extraction method was developed to obtain free oil and protein hydrolysates from dehulled rapeseeds. The rapeseed slurry was treated by the chosen combination of pectinase, cellulase, and β-glucanase (4:1:1, v/v/v) at concentration of 2.5% (v/w) for 4 h. This was followed by sequential treatments consisting of alkaline extraction and an alkaline protease (Alcalase 2.4L) hydrolysis to both produce a protein hydrolysate product and demulsify the oil. Response surface methodology (RSM) was used to study and optimize the effects of the pH of the alkaline extraction (9.0, 10.0 and 11.0), the concentration of the Alcalase 2.4L (0.5, 1.0 and 1.5%, v/w), and the duration of the hydrolysis (60, 120, and 180 min). Increasing the concentration of Alcalase 2.4L and the duration of the hydrolysis time significantly increased the yields of free oil and protein hydrolysates and the degree of protein hydrolysis (DH), while the alkaline extraction pH had a significant effect only on the yield of the protein hydrolysates. Following an alkaline extraction at pH 10 for 30 min, we defined a practical optimum protocol consisting of a concentration of 1.25–1.5% Alcalase 2.4L and a hydrolysis time between 150 and 180 min. Under these conditions, the yields of free oil and protein hydrolysates were 73–76% and 80–83%, respectively. The hydrolysates consisted of approximately 96% of peptides with a MW less than 1500, of which about 81% had a MW less than 600 Da.     

Enzymatic Demulsification of the Oil-Rich Emulsion Obtained by Aqueous Extraction of Peanut Seeds

This study details the enzymatic destabilization of the emulsion formed during aqueous extraction of peanut seeds and the quality of the resulting oil. The emulsion was exposed to enzymatic treatment and pH adjustment. The experimental results suggest that the alkaline endopeptidase Mifong^®2709 was the most effective demulsifier, while Phospholipase A2 and pH adjustment had little effect on emulsion stability. The demulsifying conditions of Mifong^®2709 were optimized by response surface methodology (RSM). The optimal conditions which produced a free oil yield of ~94 % were: 1:1 water-to-emulsion ratio, enzyme concentration of 1,600 IU/g of emulsion and 70 min hydrolysis time at 50 °C. We found that these conditions resulted in a positive relationship (R ² = 0.9671) between free oil yield and the degree of protein hydrolysis. Increased protease treatment produced a smaller number of oil droplets, but the size of these droplets increased significantly. When compared to demulsified oil products obtained by using thermal treatment, the oil obtained by Mifong^®2709 exhibited lower acid and peroxide values, contained more tocopherols and had a longer induction time as determined in the Rancimat test. The high yield and quality of peanut oil obtained by enzymatic treatment makes enzyme demulsification a promising approach to recovering free oil in aqueous extractions of peanuts.     

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

This 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.     

Destabilization of the Emulsion Formed during Aqueous Extraction of Soybean Oil

Characterization and destabilization of the emulsion formed during aqueous extraction of oil from soybean flour were investigated. This emulsion was collected as a cream layer and was subjected to various single and combined treatments, including thermal treatments and enzymatic treatments, aimed at recovery of free oil. The soybean oil emulsion formed during the aqueous extraction processing of full fat flour contains high molecular weight glycinin and β-conglycinin proteins and smaller oleosin proteins, which form a multilayer interface. Heat treatment alone did not modify the free oil recovery but freeze–thaw treatment increased the oil yield from 3 to 22%. After enzymatic treatment of the emulsion, its mean droplet size changed from 5 to 14 μm and the oil recovery increased to 23%. This increase could be attributed to the removal (due to enzymatic hydrolysis) of large molecular weight polypeptides from the emulsion interface, resulting in partial emulsion destabilization. When enzymatic treatment was followed by a freeze–thaw step, the oil recovery increased to 46%. This result can be attributed to the thinner interfacial membrane after enzymatic hydrolysis, partial coalescence during freeze–thaw, and coalescence during centrifugation. Despite the reduction in emulsion stability achieved, additional demulsification approaches need to be pursued to obtain an acceptably high conversion to free oil.     

超临界CO2萃取玉米胚芽油工艺的研究

本文研究采用超临界ＣＯ２从玉米胚芽中萃取油品的工艺条件，运用响应面法探讨了萃取压力、萃取温度和物料粒径在油品产率为９０％时对ＣＯ２消耗量的影响，确定了最佳工艺参数，分析比较了超临界ＣＯ２萃取、正己烷索氏萃取和压榨三种方法对油品质量和脂肪酸组成的影响。     

水酶法提取肉桂油的工艺研究

探索了肉桂油的水酶法提油工艺.研究了不同酶系以及酶解条件对肉桂油得率的影响.结果表明,果胶酶的作用效果好;酶解条件中,加酶量、酶解时间都对肉桂油得率有显著的影响,通过单因素分析及正交试验的结果,得出在固液比1∶6、酶解pH=3.5、酶解温度40℃、加酶量1.5％、酶解时间2h条件下,肉桂油得率能达到1.72％.经检测得到处理得到的肉桂油中桂皮醛含量达到83.6％.     

Ethanol-Assisted Aqueous Enzymatic Extraction of Peony Seed Oil

An ethanol-assisted aqueous enzymatic extraction was performed for peony seed oil (content of 30%). This method included cooking pretreatment, pectinase hydrolysis, and aqueous ethanol extraction, and the corresponding variables in each step were investigated. The changes in viscosity and dextrose equivalent values of the reaction medium as a function of changing enzymatic hydrolysis time were compared to the oil yield. The microstructures of peony seeds were analyzed using confocal laser scanning microscopy to understand the process of oil release as a result of cooking and grinding. The highest oil yield of 92.06% was obtained when peony seeds were cooked in deionized water with a solid–liquid ratio of 1:5 (w/v) at 110°C for 1 hour, ground to 31.29 μm particle size, treated with 0.15% (w/w) pectinase (temperature 50°C, pH 4.5, time 1 hour), and then extracted with 30% (v/v) aqueous ethanol (temperature 60°C, pH 9.0, time 1 hour). After processing with pectinase followed by ethanol extraction, the residual oil content in water and sediment phase decreased to 5% and 3%, respectively. The quality of the oil obtained by ethanol-assisted aqueous enzymatic extraction was good, complying with the Chinese standard.     

Bioactive Components of Commercial and Supercritical Carbon Dioxide Processed Wheat Germ Oil

Wheat germ oil (WGO) is a specialty product with a very high nutritional value. The chemical composition of both commercial and pilot scale supercritical carbon dioxide (SC-CO₂) processed WGO was examined. This study showed that methods used for oil extraction and refining did not have a significant effect on the fatty acid composition of the oil. SC-CO₂ extracted oil had a higher tocopherol content than that of commercially hexane extracted oil. The phospholipid content of the SC-CO₂ extracted oil was very low indicating that the SC-CO₂ extraction method could eliminate the degumming step from edible oil refining processes. Although the conventional chemical oil refining technique reduced the tocopherol content of the WGO, it was possible to concentrate tocopherols in WGO by using physical refining methods such as molecular distillation. Published with approval of the Director, Oklahoma Agricultural Experiment Station.     

Optimization of the Aqueous Enzymatic Extraction of Wheat Germ Oil Using Response Surface Methodology

A process of aqueous enzymatic extraction of wheat germ was carried out by a multi-enzyme preparation consisting of cellulase, pentosanase, neutrase and fungal amylase (CPNF, 2:1:2:1 w/w/w/w). Hydro-thermal heating (at 112 °C for 60 min) was more effective than oven-drying regarding emulsified oil yield. Wheat germ was ground with a rate of 10,000 rpm for 90 s. The adding level (w/w) of multi-enzyme preparation of CPNF was 1.6%. Response surface methodology was used to obtain the desired data in the process optimization. The optimal set of variables was water to wheat germ ratio (v/wt, mL/g) of 3.46, pH of 5.24, temperature of 48.49 °C and time of 6 h. The emulsified oil yield was 86.74% at the optimal levels of the tested factors. Compared with organic solvent extracted oil, the content of free fatty acid of AEE extracted oil was higher and the color was slightly darker, while the peroxide value was lower and the oxidative stability was higher owing to high content of α-tocopherol. This technique for recovering oil from fresh wheat germ with enzymes is a significant improvement in both oil yield and quality over the traditional organic solvent process.     

Optimization of an Aqueous Extraction Process for Pomegranate Seed Oil

Response surface methodology employing a five-level, four-variable central composite rotatable design was applied to study the effects of extraction time, extraction temperature, pH and water/solid ratio on the extraction yield of pomegranate seed oil using an aqueous extraction approach. In addition, quality indices, fatty acid composition and antioxidant activity of the obtained oil were studied and compared with those of typical hexane-, cold press- and hot press-extracted oil. Aqueous extraction resulted in the maximum oil recovery of 19.3% (w/w), obtained under the following critical values: water/solid ratio (2.2:1.0, mL/g), pH 5.0, extraction temperature = 63 °C and extraction time = 375 min. This yield is lower than that obtained via hexane extraction (26.8%, w/w) and higher than the yields from cold press (7.0%, w/w) and hot press (8.6%, w/w) extraction. A comparison of the characteristics of the oils based on extraction method revealed that the unsaturated fatty acid content was highest for the oil obtained by aqueous extraction. In addition, higher levels of iodine and peroxide and lower levels of acid, p-anisidine and unsaponifiable matter were observed. The oil obtained with aqueous extraction also exhibited higher antioxidant activity than oils obtained by hexane or hot press extraction.     

Simplex-Centroid Mixture Design Applied to the Aqueous Enzymatic Extraction of Fatty Acid-Balanced Oil from Mixed Seeds

A one-step method was developed to extract oil from a mixture of soybeans, peanuts, linseeds, and tea seeds using an aqueous enzymatic method. The proportion of the four seeds was targeted in accordance with a fatty acid ratio of 0.27 (SFA, saturated fatty acid(s)): 1 (MUFA, monounsaturated fatty acid(s)): 1 (PUFA, polyunsaturated fatty acid(s)), and the oil extraction yield was maximized by applying the simplex-centroid mixture design method. Three models were developed for describing the relationship between the proportion of the individual seeds in the mixture, the fatty acid ratio in the extracted oil, and the oil extraction yield, respectively. The developed models were then analyzed using an ANOVA and were found to fit the data quite well, with R ² values of 0.98, 0.93, and 0.93, respectively. The three models were validated experimentally. The results indicated that the ratio of fatty acids in the oil ranged between 0.98 and 1.12 (MUFA:PUFA) and between 0.26 and 0.28 (SFA:MUFA), which were quite close to the target values of 1 and 0.27, respectively. The oil extraction yield of 62.13 % was slightly higher than the predicted value (60.32 %).     

Foam Separation of Oil from Enzymatically Treated Wet-Milled Corn Germ Dispersions

More than 9 billion gallons of ethanol were produced in 2008, mostly from dry grind corn fermentation plants. These plants are a potential source of substantial amounts of corn oil, if an economical method of separating it can be developed. In this work, oil was separated from corn germ by aqueous enzymatic extraction (AEE). Batches of wet-milled corn germ in water were preheated in a pressure cooker, ground in a colloid mill, and churned in a vertical column/mixing vessel system, after the addition of enzyme. Nitrogen gas was then bubbled through the column removing an overflowing foam fraction which was subsequently centrifuged to separate free oil. Using a newly commercialized enzyme complex it was found that 80% of the oil could be recovered using a w/w ratio of enzyme solution to germ of 1:80. The low dose and low price of the enzyme complex leads to a cost estimate of AEE of corn oil from germ, similar to the wet-milled germ extracted, cost competitive with expelled oil (with the separation and drying of the foam protein), and feasible for commercialization in a dry grind plant retrofitted to separate germ.     

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

Aqueous 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.     

Destabilization of Emulsion Formed During Aqueous Extraction of Peanut Oil: Synergistic Effect of Tween 20 and pH

To destabilize the emulsion formed during aqueous extraction processing (AEP) of peanuts, Tween and Span series surfactants (Tween 20, Tween 80, Span 20, and Span 80) were used alone or in combination to break the emulsion. Results indicate that only Tween surfactants had a pronounced demulsifying effect that was dependent on Tween concentration and system pH. When 1.2 wt% Tween 20 aqueous solution was used for oil extraction at pH 10.0, the highest free oil yield was achieved at 76.1 %, which was similar to the oil recovery of using proteases as a destabilization agent. The results obtained using a model emulsion system containing peanut oil and Tween 20/peanut protein isolates (PPI) showed that when Tween 20 and PPI coexisted in extraction medium at pH 10.0, the dynamic interfacial tension and droplet size distribution curves were very similar to those when Tween 20 was used alone, suggesting that Tween 20 dominated at the interface, instead of PPI. Destabilization of the model emulsions relied on three important factors: inclusion of Tween 20 at the initial mixing stage, high pH, and a gentle mixing speed. A synergistic destabilization mechanism of using Tween 20 at high pH during AEP was proposed. The discovery of Tween 20 as an effective demulsifier significantly contributes to the development of AEP of oilseeds.     

Separating Oil from Aqueous Extraction Fractions of Soybean

Previous research has shown that enzyme-assisted aqueous extraction processing (EAEP) extracts 88–90% of the total soybean oil from extruded full-fat soy flakes into the aqueous media, which is distributed as cream (oil-in-water emulsion), skim, and free oil. In the present work, a simple separatory funnel procedure was effective in separating aqueous skim, cream and free oil fractions allowing mass balances and extraction and recovery efficiencies to be determined. The procedure was used to separate and compare liquid fractions extracted from full-fat soy flour and extruded full-fat soy flakes. EAEP extracted more oil from the extruded full-fat soy flakes, and yielded more free oil from the resulting cream compared to unextruded full-fat soy flour. Dry matter partitioning between fractions was similar for the two procedures. Mean oil droplet sizes in the cream and skim fractions were larger for EAEP of extruded flakes compared to non-enzymatic AEP of unextruded flour (45 vs. 20 μm for cream; 13 vs. 5 μm for skim) making the emulsions from EAEP of extruded flakes less stable. All major soy protein subunits were present in the cream fractions, as well as other fractions, from both processes. The cream could be broken using phospholipase treatments and 70–80% of total oil in the extruded full-fat flakes was recovered using EAEP and a phospholipase de-emulsification procedure.     

Effect of Subcritical Carbon Dioxide Extraction and Bran Stabilization Methods on Rice Bran Oil

The extraction of rice bran oil using the conventional organic solvent‐based Soxhlet method involves hazardous chemicals, whereas supercritical fluid extraction is a costly high‐temperature operating system. The subcritical carbon dioxide Soxhlet (SCDS) system, which operates at a low temperature, was evaluated for the extraction of rice bran oil in this study. In addition, rice bran that had been subjected to steam or hot‐air stabilization were compared with unstabilized rice bran (control). The yields; contents of tocopherols, tocotrienols and oryzanol; fatty acid profiles; and the oxidative stabilities of the extracted rice bran oils were analyzed. The yields using hexane and SCDS extraction were approximately 22 and 13–14.5 %, respectively. However, oil extracted using the SCDS system contained approximately 10 times more oryzanol and tocol compounds and had lower free fatty acid levels and peroxide values compared with hexane‐extracted oil. Overall, SCDS extraction of steamed rice bran represents a promising method to produce premium‐quality rice bran oil.     

Optimization of Aqueous Enzymatic Microwave Assisted Extraction of Macadamia Oil And Evaluation of Its Chemical Composition,Physicochemical Properties,and Antioxidant Activities

This study presents the green and effective aqueous enzymatic process assisted microwave extraction (AEME) to preparate macadamia nut oil (MNO). The conditions of the extraction process are optimized (extraction temperature 50 °C, extraction time 64 min, enzyme concentration 1.60% (w/w), and irradiation power 450 W). An oil yield of 58.09 ± 0.63% is achieved under these optimal conditions. The scanning electron micrograph (SEM) analysis of nuts before and after extraction illustrates that AEME promotes the emancipation of oil stored within the organelles. Gas chromatography-flame ionization detector (GC-FID) analysis reveals the fatty acid compositions of MNOs obtained by AEME and the Soxhlet extraction (SE) are similar and mainly dominated by monounsaturated fatty acids beneficial to human health which is higher in MNO than in any other known food. Moreover, gas chromatography-mass spectrometry (GC-MS) analysis reveals higher amounts of more odoriferous oxygenated terpenes is present in the MNO extracted by AEME in comparison with SE. The physicochemical properties of AEME oil are more excellent than those of SE oil. Moreover, AEME oil exhibits superior antioxidant capacities. In conclusion, green AEME gives relatively satisfactory yield and better retains the fragrance and functionality of MNO. Practical Applications: The present study provides a green extraction method and valuable data for the process design as well as industrial scale-up applications. In addition, compared to the nonsustainable and environmentally nonfriendly traditional method, AEME preserves the initial composition of the flavor substances and enhances the extraction of healthy beneficial compounds in MNO. Therefore, AEME oil can be used to develop functional edible oils or even in medicinal, cosmetic, and pharmaceutical preparations.