Screening of industrial enzymes for deproteinization of shrimp head for chitin recovery

Food grade proteolytic enzymes were examined for deproteinization of shrimp head. Shrimp head was easily deproteinized by Alcalase ® and trypsin at a pH of 8.0. Alcalase was chosen as the most efficient commercial enzyme for deproteinization of shrimp head. Alcalase treatment of shrimp head recorded 61% of weight loss on dry basis and a residual protein of 275 mg/g dried shrimp head. The enzymatically deproteinized shrimp head was later demineralized with lactic acid using microwave radiation at 400W. The combination of enzymatic and physicochemical treatments promoted the chitin recovery from dried shrimp head under eco-friendly conditions.     

酶法从虾壳中提取甲壳素工艺优化的研究

利用蛋白酶水解虾壳,以水解率为指标,研究了各种条件对水解效果的影响。通过比较不同蛋白酶的水解效果,发现碱性蛋白酶的水解效果最好,最适的水解条件为:碱性蛋白酶添加量5000U/g,pH8.5,水解温度60℃,水解时间6h,水解pH为8.5,底物质量浓度2g/100mL,在此条件下,水解率达到78.3%。最后通过扫描电镜(SEM)观察了采用不同脱除蛋白方法处理后的甲壳素表面状态,酶法与碱法脱蛋白对甲壳素表面微观状态影响不同,酶法脱蛋白后甲壳素表面较光洁,证实本方法分离效果合理。      

Production,properties, and some new applications of chitin and its derivatives

Chitin is a polysaccharide composed from N-acetyl-D-glucosamine units. It is the second most abundant biopolymer on Earth and found mainly in invertebrates, insects, marine diatoms, algae, fungi, and yeasts. Recent investigations confirm the suitability of chitin and its derivatives in chemistry, biotechnology, medicine, veterinary, dentistry, agriculture, food processing, environmental protection, and textile production. The development of technologies based on the utilization of chitin derivatives is caused by their polyelectrolite properties, the presence of reactive functional groups, gel-forming ability, high adsorption capacity, biodegradability and bacteriostatic, and fungistatic and antitumour influence. Resources of chitin for industrial processing are crustacean shells and fungal mycelia. Fungi contain also chitosan, the product of N-deacetylation of chitin. Traditionally, chitin is isolated from crustacean shells by demineralization with diluted acid and deproteinization in a hot base solution. Furthermore, chitin is converted to chitosan by deacetylation in concentrated NaOH solution. It causes changes in molecular weight and a degree of deacetylation of the product and degradation of nutritionally valuable proteins. Thus, enzymatic procedures for deproteinization of the shells or mold mycelia and for chitin deacetylation were investigated. These studies show that chitin is resistant to enzymatic deacetylation. However, chitin deacetylated partially by chemical treatment can be processed further by deacetylase. Efficiency of enzymatic deproteinization depends on the source of crustacean offal and the process conditions. Mild enzymatic treatment removes about 90% of the protein and carotenoids from shrimp-processing waste, and the carotenoprotein produced is useful for feed supplementation. In contrast, deproteinization of shrimp shells by Alcalase led to the isolation of chitin containing about 4.5% of protein impurities and recovery of protein hydrolysate.     

Proteolysis characteristics of sarcoplasmic, myofibrillar, and stromal proteins separated from grass carp and antioxidant properties of their hydrolysates

Proteolysis of grass carp sarcoplasmic, myofibrillar, and stromal proteins by 5 commercial proteases were studied. Sarcoplasmic and myofibrillar protein could be well hydrolyzed by Alcalase 2.4 L to reach high protein recoveries (PR) (71.86±2.46 and 80.77±3.05%, respectively), while the maximum PR for stromal protein was only 42.83±2.84%. However, stromal hydrolysates, containing mostly 6–10 kDa fraction, exhibited higher ·OH scavenging activities due to its high content of antioxidant-assisting amino acids. Alcalase 2.4 L and pancreatin 6.0, which produced hydrolysates with relative high degree of hydrolysis (DH), were used for further hydrolysis of whole grass carp protein with the assistance of response surface methodology (RSM). The results showed that serine proteases (Alcalase 2.4 L and pancreatin 6.0) could produce sarcoplasmic, myofibrillar, or stromal hydrolysates with relatively high PR, DH, and strong ·OH scavenging activity, which may be used to prepare antioxidant hydrolysates from grass carp.     

The effect of enzyme systems and processing on the hydrolysis of peanut (Arachis hypogaea L.) protein

In vitro protein digestion studies were carried out on raw and roasted peanut flour as the starting material in the production of peanut protein hydrolysate. Peanut flour was hydrolyzed with alcalase and alternately in a sequential digestion with pepsin-pancreatin, both for up to 24 h. The degree of hydrolysis (DH) at different times of hydrolysis was determined using the trinitrobenzenesulfonic acid (TNBS) method. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was used to indicate destruction of native protein units in the enzymatic digests.Hydrolysis with alcalase was very rapid for the first 6 h after which a plateau was reached, whereas that with pepsin–pancreatin was more gradual reaching a plateau after 12 h of hydrolysis. Raw peanut hydrolyzed with alcalase and pepsin–pancreatin had 23% and 21% DH after 24 h respectively, whilst roasted peanut hydrolyzed with alcalase had 21% DH, with the pepsin–pancreatin hydrolysate recording the highest value of 25% after 24 h of hydrolysis.SDS-PAGE results showed that raw peanut samples behaved differently from the roasted samples; increasing hydrolysis time reduced larger peanut protein subunits, with only peptides of <20 kDa visible after hydrolysis for raw peanut, and virtually no distinct visible bands for the roasted peanut after 3 h of hydrolysis.     

EXTRACTION OF CAROTENOPROTEIN FROM BLACK TIGER SHRIMP SHELLS WITH THE AID OF BLUEFISH TRYPSIN

The effect of bluefish (Pomatomus saltatrix) trypsin on the recovery and characteristics of carotenoprotein from black tiger shrimp (Penaeus monodon) shells was investigated. Trypsin concentration and reaction time both affected the hydrolysis and the recovery of carotenoproteins ( P <  0.05). The recovery of carotenoproteins from shrimp shells was maximized by the hydrolysis of shrimp shells using 1.2 trypsin units/g shrimp shells for 1 h at 25C. Freeze-dried carotenoprotein recovered contained 70.20% protein, 19.76% lipid, 6.57% ash, 1.50% chitin, and 87.91 µg total astaxanthin/g sample, indicating a substantial reduction in the levels of antinutrients associated with shrimp waste, while enriching the product in carotenoid pigments and valuable essential nutrients (proteins and lipids). Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) of the recovered carotenoprotein revealed that protein with molecular weight of 45 kDa was the major constituent. When hydrolytic activities of bluefish and bovine trypsins toward carotenoproteins in black tiger shrimp shells were compared, the recovery efficacy of protein and pigment by bluefish trypsin was similar to that achieved by trypsin from bovine pancreas. Therefore, bluefish trypsin could be used as an alternative cheap proteinase for extraction of carotenoproteins from black tiger shrimp shells.

PRACTICAL APPLICATIONS

Carotenoproteins from black tiger shells, the byproduct of shrimp processing, can be recovered with the aid of fish trypsin. This product can be used for both food and feed applications. Additionally, the fish trypsin can be used instead of bovine trypsin. As a whole, the utilization of fish and shellfish processing wastes can be maximized.     

Extraction of Carotenoprotein from Shrimp Process Wastes with the Aid of Trypsin from Atlantic Cod

Atlantic cod trypsin or bovine trypsin were used to aid the extraction of carotenoprotein from shrimp wastes at 4°C. When 25 mg% cod trypsin was added to extraction medium containing 0.5N ethylene diaminetetraacetic acid (EDTA) 64% of the astaxanthin and 81% of the protein of shrimp waste was recovered as carotenoprotein in 24 hr. With 25 mg% bovine trypsin, under otherwise identical conditions, the carotenoprotein recovered represented 49% of the astaxanthin and 65% of the protein of the waste. Semi-purified cod trypsin was not as effective as pure trypsin in facilitating recovery of carotenoprotein from shrimp waste. The recovery of carotenoprotein from shrimp waste, during extraction at 4°C with or without trypsin, was facilitated by EDTA.     

Bioconversion of Shellfish Chitin Waste: Waste Pretreatment, Enzyme Production, Process Design, and Economic Analysis

Study of pretreatment of shrimp processing waste for a chitin bioconversion scheme to produce yeast single-cell protein established conditions for size reduction, deproteination, and demineralization. Enzymatic hydrolysis of pretreated chitin waste achieved 80% conversion in 24 hr. Optimum temperature and pH were determined for maximum chitinase production in submerged culture, using pretreated chitin waste as substrate. An integrated process scheme for conversion of shrimp shell chitin waste to yeast single-cell protein based on these and previous results was designed and analyzed economically, giving a negative after-tax cash flow of $0.06 per kg of wet waste.     

Functional Properties and Bioactivities of Pine Nut (Pinus gerardiana) Protein Isolates and Its Enzymatic Hydrolysates

The functional properties and bioactivities of the pine nut protein isolates (PPI) and its enzymatic hydrolysates (PPH) prepared with Alcalase at 5 %, 10 %, 15 % and 25 % degree of hydrolysis (DH) were studied. The solubility of PPH significantly increased (p?<?0.05) with the increase of the DH, while the foaming capacity of PPH was only improved at a low DH. However, enzymatic hydrolysis reduced the emulsifying capacity of PPH. The DPPH radical scavenging and inhibition of linoleic acid oxidation activities of PPH were significantly improved by a low DH (5 %) compared with those of PPH with a higher DH and the original PPI (p?<?0.05). The reducing power of PPH at all DH decreased in comparison to that of the original PPI. Potent angiotensin-converting enzyme (ACE) inhibitory peptides could be generated by hydrolysis with Alcalase, and the ACE inhibitory activity of PPH increased (p?<?0.05) with the DH. These results revealed that a low degree of enzymatic hydrolysis was appropriate to obtain PPH with improved functional properties and good antioxidant activities, while a high degree of hydrolysis was essential to obtain highly potent ACE inhibitory peptides from PPI. These results suggest that the control of the DH may be an effective strategy to modify specific functional and bioactive properties of PPH, and PPH has potential as a functional food ingredient for related functional and health benefits.     

花生蛋白水解产物ACE抑制活性的研究

采用胰蛋白酶、碱性蛋白酶、中性蛋白酶水解花生蛋白,研究了水解过程中水解度的变化,并对水解产物的ACE抑制活性进行了探讨。得出三种酶对花生蛋白的水解作用:碱性蛋白酶>胰蛋白酶>中性蛋白酶。碱性蛋白酶水解产物ACE抑制活性明显高于胰蛋白酶和中性蛋白酶,水解产物的ACE抑制活性高达89.73%,中性蛋白酶水解产物ACE抑制率仅为27.24%。     

双酶法制备马鹿茸降血糖肽工艺优化及其对α-葡萄糖苷酶的抑制效果

为探索制备马鹿茸降血糖肽的最佳工艺条件,以α-葡萄糖苷酶抑制率为指标,从碱性蛋白酶、风味蛋白酶、中性蛋白酶和胰蛋白酶中筛选出两种酶,根据其体外降血糖效果确定酶的作用顺序,再以水解度、α-葡萄糖苷酶抑制率和蛋白质回收率为指标进行单因素试验和正交试验,优化降血糖肽制备工艺条件。结果表明:碱性蛋白酶和风味蛋白酶比中性蛋白酶和胰蛋白酶更适合用于制备马鹿茸降血糖肽。采用碱性蛋白酶-风味蛋白酶顺序对马鹿茸进行水解,所得酶解产物的α-葡萄糖苷酶抑制率、蛋白质回收率和水解度较高,分别为21.11%、39.12%、19.88%。通过单因素试验和正交试验确定双酶酶解最佳工艺条件为先用碱性蛋白酶在p H 8.0、60℃、底物质量分数12%、加酶量5 000 U/g条件下酶解3 h,再用风味蛋白酶于p H 6.5、45℃、底物质量分数5%、加酶量6 000 U/g条件下酶解1 h。双酶分步水解终产物的α-葡萄糖苷酶抑制率受质量浓度的影响,当质量浓度为3 mg/m L时,α-葡萄糖苷酶抑制率可达94.09%,IC50值为1.82 mg/m L。碱性蛋白酶-风味蛋白酶双酶分步水解马鹿茸可获得高α-葡萄糖苷酶抑制率的降血糖肽。     

菜籽分离蛋白分子质量分布及酶解条件的研究

以脱脂"双低"油菜籽为原料,利用碱溶解酸沉淀法提取菜籽分离蛋白;用SDS-PAGE凝胶电泳研究菜籽蛋白的分子质量的组成;以水解度和氮回收率为考察指标,用响应面分析法拟合了Alcalase 2.4L酶解菜籽蛋白成菜籽肽的二次多项数学模型,优化了酶解菜籽分离蛋白的工艺参数。SDS-PAGE凝胶电泳研究表明利用碱溶解酸沉淀提取的菜籽分离蛋白主要是2S清蛋白。响应面分析法研究的试验结果表明,酶的使用量、pH、酶解温度、酶的使用量与pH的交互作用对Alcalase 2.4L酶解菜籽蛋白的水解度和氮回收率的影响均显著(P0.05)。通过求解菜籽肽的二次多项数学模型的逆矩阵方程,可得Alcalase 2.4L水解菜籽分离蛋白的最佳条件为:酶的使用量0.05 Au/g,pH 9.0,酶解温度54℃;在此酶解条件下,菜籽分离蛋白浓度为5%时,水解5 h,所得的水解度和氮回收率分别为35.12%及52.96%。     

贻贝蛋白的酶解及其酶解物的抗氧化活性研究

比较了6种不同的蛋白酶(胰蛋白酶、风味蛋白酶、中性蛋白酶、木瓜蛋白酶、复合蛋白酶、碱性蛋白酶)对贻贝粗蛋白的酶解效果,确定碱性蛋白酶为最适用酶。用此酶制备不同水解度(DH 6%、DH 11.5%、DH16%、DH 20%、DH 25%)的贻贝蛋白酶解物,考察不同DH酶解产物的抗氧化活性。试验结果表明:贻贝酶解产物具有较强的抗氧化活性,并呈一定的量效关系;当DH为25%时,贻贝酶解物对DPPH自由基、超氧自由基、羟基自由基的清除率最高,分别为77.4%、75.2%、43.4%,同时具有最强的金属螯合率(64.7%);而DH为16%时,酶解物对亚油酸的过氧化抑制作用较还原型谷胱甘肽强,达65.6%。DH对酶解产物的抗氧化活性有一定的影响,但在不同的抗氧化体系中,影响趋势不一致。     

Carotenoproteins from lobster waste as a potential feed supplement for cultured salmonids

Lobster waste (including the head and hard carapace, viscera, mandibles and gills) contains approximately 54 μg/g total astaxanthin, 29% protein, 23% chitin, 34% ash and 2.2% crude fat on a dry weight basis. Trypsin from bovine pancreas was applied to facilitate the recovery of carotenoid pigments and protein as carotenoprotein complex, which was subsequently air‐dried to a stable powder form at 45°C and 15% relative humidity. The product obtained was found to contain 60% protein, 15% crude fat, 6% ash, 8% chitin and 295 μg/g total astaxanthin. Thus, the process achieved a substantial reduction in the levels of anti‐nutrients associated with lobster waste (i.e., ash and chitin) while elevating the levels of carotenoid pigments and essential nutrients such as protein and fat in the recovered product These characteristics of the final product suggest that it could be used as an inexpensive source of pigment and protein in diets of cultured salmonid species.     

蚕蛹多肽制备工艺及其体外抗氧化活性

采用碱性蛋白酶、风味蛋白酶、复合蛋白酶、木瓜蛋白酶、胰蛋白酶水解蚕蛹蛋白,以水解度和清除DPPH·能力为指标对酶解过程进行分析,并研究水解产物多肽的体外抗氧化活性。结果表明,碱性蛋白酶对蚕蛹蛋白具有较好的水解效果,其水解产物有较高抗氧化活性,对DPPH·、超氧阴离子自由基(O2·)和羟自由基(·OH)都具有较强的清除能力。     

中国对虾调味料风味前体物质酶解制备工艺研究

以中国对虾下脚料为原料,利用酶解技术获得中国对虾调味料风味前体物质——酶解液,并通过响应面分析方法优化酶解工艺。研究结果表明,最佳酶解工艺条件为料水比1/9,酶解时间3h,酶用量2.4%,酶解温度68℃,初始pH6.5。在此条件下,中国对虾下脚料水解度为16.58%。水解液中氨基酸含量为23.2028mg/mL,其中必需氨基酸含量为10.5312mg/mL。所得酶解液具有诱人、浓郁的虾风味,可以作为调味基料。     

南美白对虾虾头制备鲜味水解物的研究

本文以25 ℃下自溶6 h后的南美白对虾虾头为原料,采用酶水解法进行鲜味水解物的制备。以鲜味滋味、蛋白质回收率、肽得率和水解度为评价指标,进行外源酶的选择和酶解工艺条件的逐步优化研究。研究表明,经四种单酶酶解,均明显提升酶解效率,其中菠萝蛋白酶(Bromelain)和风味蛋白酶(Flavorzyme)能有效提升酶解液的鲜味、降低苦涩味。制备具有良好鲜味特征水解物的条件为:自溶后的虾头,加入菠萝蛋白酶(420 U/g样品)和风味蛋白酶(12 U/g样品)两种酶(料液比1:1.5 (g/mL)),在pH7.5、50 ℃下酶解3 h,该工艺下蛋白质回收率、肽得率和水解度分别可达79.75%、71.71%和18.28%。获得水解物氨基酸组成中ΣEAA/ΣAA与ΣEAA:ΣNEAA分别为37%和1:1.7。65%的总蛋白氨基酸释放为游离氨基酸,其中,59%的总蛋白鲜甜味氨基酸释放为游离氨基酸,水解物营养价值高且具有良好鲜味。     

牡蛎复合酶解提高蚝油风味的工艺探讨

采用多种酶复配的方法对牡蛎进行酶解处理,以获得风味良好、且蛋白质回收率和水解程度都比较高的产品.研究结果表明采用含0.05%中性蛋白酶+0.1%碱性蛋白酶+0.1%风味蛋白酶+0.1%复合蛋白酶的复合酶,可以使蛋白质回收率达到85%、水解度达到43.5%,酶解后的游离氨基酸含量比酶解前提高122.2%,酶解液无腥味,且...     

Combined effects of high-pressure and enzymatic treatments on the hydrolysis of chickpea protein isolates and antioxidant activity of the hydrolysates

A chickpea protein isolate (CPI) was pretreated before hydrolysis under a pressure that varied between 100 and 600MPa. The hydrolysis rate increased significantly with pressure above 300MPa. At 40min, the DH of the control was 15.3%, while the DH of the CPI treated at 300MPa was 18.5%, which reached 23.74% post treatment at 400MPa. The pretreatment of CPI above 300MPa enhanced the superoxide anion capturing rate of enzymatic hydrolysis. Pretreatment at 400MPa significantly reduced the hydrolysis time with the release of antioxidant peptides. While hydrolysis by Alcalase during treatment at high pressure (100-300MPa) significantly increased the degree of hydrolysis (DH), its maximum value peaked after hydrolysis at 200MPa for 30min. In addition, hydrolysates obtained at high pressure (100-300MPa) had a higher superoxide anion capturing rate. High-pressure treatment at 200MPa for 20min resulted in products with high antioxidative activity. The molecular-weight (MW) determination of the enzymatic hydrolysates indicated that hydrolysis at high pressure could significantly increase the amount of low-molecular-weight peptides.     

酶法制备不同水解度高温豆粕水解产物的理化特性研究

采用Alcalase酶和木瓜蛋白酶分别对高温大豆粕进行酶解,通过控制酶解反应得到水解度为5%、10%和15%的6种水解产物,研究两种酶对不同水解度的水解产物理化特性的影响。结果表明,Alcalase酶和木瓜蛋白酶均可产生6种不同分子量范围的水解产物,但各部分比例具有显著差异(P0.5),其平均分子量随水解度的增加逐渐减少,Alcalase酶的水解产物中小于2562 Da小分子量肽所占比例更高。豆粕蛋白的疏水基团在酶解反应中发生暴露与断裂的数量差,导致其表面疏水性随水解度增加呈现先下降再上升的变化,即水解度为10%的表面疏水性最低。zeta电势的绝对值随水解度不断上升,分子间的斥力增大,相同水解度下两种酶对zeta电势的影响并不显著。此外,在pH值为3、5、7和9时,水解产物的溶解性随着水解度的增加而逐渐增高,乳化活性和乳化稳定性则逐渐降低。