Saccharification of chitin using solid-state culture of Aspergillus sp. S1-13 with shellfish waste as a substrate

Saccharification of chitin was performed in a suspension (mash) of a solid-state culture of chitinase-producing Aspergillus sp. Sl-13 with acid-treated shellfish waste as a substrate. The conditions for the saccharifying reaction and the solid-state cultivation were examined from the viewpoint of saccharification in the mash. Optimum cultivation conditions were defined: a solid-state medium consisting of 5 g of 10% lactic acid-treated crab shells (0.50-2.36 mm in size) and 3 ml of a basal medium (0.028% KH2PO4 0.007% CaCl2.2H2O, and 0.025% MgSO4.7H2O) supplemented with 0.3% peptone was inoculated with 4 ml of spore suspension (1 x 10(7) spores/ml), and the water content of the medium was adjusted to 75%; static cultivation at 37 degrees C for 7 d. When a culture obtained under the optimum conditions was suspended in 70 ml of 50 mM sodium phosphate-citrate buffer (pH 4.0) and incubated at 45 degrees C for 11-13 d, 55 mM N-acetylglucosamine (GlcNAc) was formed in the solid-state culture mash, indicating that at least 33% of the initial chitin in the solid material was hydrolyzed. Through the experiments, the amounts of G1cNAc formed in the solid-state culture mash varied in a way similar to that of the water-extractable pnitrophenyl beta-D-N-acetylglucosaminide-hydrolyzing enzyme in the culture, but not to that of the colloidal chitin-hydrolyzing enzyme. G1cNAc-assimilating lactic acid bacteria, which were inoculated into the mash after or at the start of the saccharification, formed lactic acid with decreasing GlcNAc.     

Solid substrate production of Epicoccum nigrum conidia for biological control of brown rot on stone fruits

Production of conidia of Epicoccum nigrum, a biocontrol agent of the fungal pathogen Monilinia laxa, was tested in liquid- and solid-state fermentation. Liquid fermentation was conducted in 250 ml Erlenmeyer flasks containing 50 ml of a mineral medium (containing per litre: 20 g lactose, 10 g NO3K, 1 g K2HPO4, 0.5 g MgSO4.7H2O, and 1 ml of a minor-element solution), inoculated with 2 x 10(5) E. nigrum conidia ml(-1), and incubated at 20-25 degrees C and 150 rpm for 7 days. Solid-state fermentation was carried out in specially designed plastic bags (600 cm3) (VALMIC) containing either 50 g of peat/vermiculite (1:1, w/w), or 50 g of peat/vermiculite/lentil meal (1:1:1, w/w/w) with 40% (v/w) initial moisture content. Substrate was inoculated with a conidial suspension of E. nigrum to give 10(5) conidia g(-1) substrate, and bags were incubated at 20-25 degrees C for 7 days in darkness. The amount of conidia of E. nigrum obtained in solid-state fermentation with substrate based on peat/vermiculite/lentil meal was 10-fold higher than with substrate based on peat/vermiculite or in liquid fermentation. Conidial production under these conditions was maintained in the range of 10(8) conidia g(-1) substrate from 10 to 150 days after inoculation. Germinability of these conidia was >90%. Addition of other nutrients than lentil meal to peat/vermiculite did not enhance production of conidia. Presence of peat in the substrate was necessary for good conidia production, but change in the kind of peat or vermiculite did not improve conidial production. Conidial production was similar when the substrate was inoculated with 10(5), 10(6) or 10(7) conidia g(-1) dry substrate. Incubation of bags in light conditions did not enhance conidial production. Fresh conidia produced in this solid-state fermentation system reduced the incidence and lesion diameter induced by M. laxa on peaches.     

Polyol production by culture of methanol-utilizing yeast

Four methanol-utilizing yeasts, Candida boidinii, Hansenula polymorpha, Hansenula ofunaensis, and Pichia pinus, produced polyols from corresponding sugars in a methanol medium. H. polymorpha produced larger amounts of xylitol than the other yeasts. Productivity was the highest at pH 8 when 5 g (dry)/l cultured cells were incubated with 2.5 g/l urea as the nitrogen source in a medium containing 1% (v/v) methanol and 1 g/l MgSO4.7H2O. Under these conditions, 57 g/l xylitol was obtained from 110 g/l D-xylose after 3 d of cultivation. The largest amount of xylitol (58 g/l; yield, 0.62 g/g) was produced from 125 g/l, D-xylose and 5% (w/v) glycerol instead of methanol after 4 d of cultivation.     

Efficient production of a heterologous peroxidase, DyP from Geotrichum candidum Dec 1, on solid-state culture of Aspergillus oryzae RD005

The productivity of a peroxidase (DyP) originating from Geotrichum candidum Dec 1 was enhanced in the solid-state culture using Aspergillus oryzae RD005. When the humidity, water content, and temperature were adjusted to 60%, 50% and 27°C, respectively, the productivity of DyP reached 5.3 g per kilogram wheat bran, which was used as the solid medium. The yield of 5.3 g per kg wheat bran corresponded to the yield of a 56 kg submerged culture. The productivity per gram carbon of the medium in the solid-state culture was 4.1-fold that in the submerged culture.     

pH值及乳酸菌对米曲霉固态制曲过程的影响

考察了在固态培养条件下,不同的初始pH值及乳酸菌对米曲霉固态制曲过程的影响。以豆粕和麸皮(质量比为55∶45)为原料,1∶0.6的料水比,0.3%～0.5%的接种量接入沪酿3.042米曲霉种曲,33℃下制曲,定时取样测定固体曲的蛋白酶活力及孢子数。结果表明:适当偏酸性的环境有利于成曲蛋白酶的提高,培养基初始pH调节为6.5时米曲霉中性蛋白酶活力获得最大值(4 080±61)U/g干基,较对照组提高33%～37%;添加1.0%～2.0%乳酸菌,也同样起到改善米曲霉制曲过程的效果。     

Rennin-like milk coagulant enzyme produced by a local isolate of Mucor

Among 20 isolates of Mucor isolated from various environments in Jordan and found to produce a rennin-like acid protease, known as Mucor rennin-like enzyme (MRE), Mucor J20 was found to produce the highest level of MRE. The optimum incubation conditions for enzyme production in a fortified wheat bran mixture using solid-state fermentation were 3–4 days at 30°C. The highest MRE activity (185–200 rennin units or RU) was produced in a medium containing wheat bran and lentil straw (1 : 1 w/w) moistened with whey, and incubated in clay pots at 30°C for 4 days. A slightly lower activity value (178 RU) was found when using a mineral salt solution or distilled water instead of whey, or when using wheat bran alone with whey. At pH 4, the MRE retained its complete activity (100%) for 6 weeks at 5°C and 10°C, and for 3 and 2 weeks at 20°C and 30°C, respectively. After heating at 60°C for 10 min, the enzyme lost its activity at all pH levels used (pH 2–8). The crude extract of MRE was successfully applied in the manufacture of a cheese curd.     

Stimulation of polygalacturonase production in an immobilized system by <Emphasis Type="Italic">Aspergillus</Emphasis> sp.: effect of pectin and glucose

The synthesis of polygalacturonases (PG) is known to be influenced by Aspergillus growth conditions, namely, environmental factors and pectin content in the cultivation medium containing a mixed carbon source. Optimal conditions were attained at a temperature of 30 °C and an initial pH of 4.5. PG activity (3.29 and 2.48 U/mL) was determined after a two-day culture of Aspergillus sp. HC1 and Aspergillus sp. CC1, respectively, in a basic medium containing 2% citrus pectin as the sole carbon source. The addition of glucose (2% w/v) to the basic medium led to a 2-fold increase in PG production. However, enzyme synthesis was repressed when a higher concentration of glucose was used in the medium containing the mixed carbon source. Spores from the two fungi were immobilized in a 3% Ca–alginate system and the mechanical strength of the gel beads allowed the use of this process system 6-fold longer (288 h) than the free culture. In the Aspergillus sp. CC1 immobilized system, PG production increased nearly 10-fold in the medium with 2% glucose added (5.95 U/mL) in comparison to the medium without sugar (0.55 U/mL). The results demonstrate that a different response in activity was produced by free and entrapped spore systems. PG production remained approximately constant throughout the six 48 h cycles in the medium containing citrus pectin (2% w/v) as the sole carbon source.     

羊肚菌液体发酵培养基成分及培养条件的优化

采用单因素和正交试验设计法对羊肚菌液体发酵培养基成分及培养条件的优化进行了研究。结果表明,羊肚菌液体发酵培养基最佳配方为可溶性淀粉2.5%,酵母膏2.5%,MgSO4 0.1%, KH2PO4 0.2%, VB1 0.2%;最适培养条件：pH值为7,发酵温度28℃、转速140r/min,装液量100mL/250mL,摇床振荡培养7d,在最佳条件下所生产的菌丝生物量为3.438g/200mL。羊肚菌液体发酵培养基成分和发酵条件的优化工艺能高效生产菌丝体,为以后开发羊肚菌真菌资源产品奠定了基础。     

Development of a solid-state fermentation process for production of an alpha amylase with potentially interesting properties

Ca-independency with potential activity and stability at low pH are among the most interesting characteristics of α-amylase in starch industry. In this attempt the synergetic effect of low pH on activity of crude Ca-independent α-amylase isolated from a native Bacillus sp. KR-8104 in solid-state fermentation (SSF) was studied using wheat bran (WB) as a substrate. The effects of different parameters including moisturizing agents, solid substrate to moisture ratio, particle size, incubation temperature and period, inoculum (v/w) and supplementation with 1% (w/w) different carbon and nitrogen sources on enzyme production were investigated. Maximum enzyme production of 140 U/g dry fermented substrate was obtained from wheat bran moistened with tap water at a ratio of 1:1.5 and supplemented with 1% (w/w) NH₄NO₃ and 1% (w/w) lactose after 48 h incubation at 37 °C. Even though the production of α-amylase was lower at 40 and 45 °C, the viable cell count was higher. In addition response surface methodology (RSM) was applied to find optimum conditions of temperature and pH on crude amylase activity. Using central composite design (CCD) a quadratic mathematical model equation was derived for the prediction of enzyme activity. The results showed that the model was in good agreement with experimental results, with R² = 0.90 (p < 0.0001) and the low pH has a synergetic effect on enzyme activity at higher temperature.     

球孢链霉菌溶菌酶的产生条件研究

研究了球孢链霉菌S186（Streptomycesglobisporus）溶菌酶的产生条件，球孢链霉菌在由糊精2．0％，大豆蛋白0．4％，多聚股0．2％，NaCl0．7％，KH2PO40．05％，Na2HPO40．1％，CaCl20．01％（起始pH7．5）组成的培养基中，30℃，240r／min振荡培养96h，酶活力达到30．1U／ml。     

红曲霉Monascus sanguineus X1处理黄水的培养基优化及酯化液制备

黄水是白酒酿造的主要副产物之一,富含多种有机质,可用于制备酯化液促进白酒增香,提高其利用率。研究了一株高产酯化酶的红曲霉菌株Monascus sanguineus X1,以酯化酶活力为指标,从碳源、碳氮比、接种量和pH值4个方面对X1菌株的培养基进行单因素实验及正交试验优化;采用酯化酶液处理黄水,制备酯化液,研究了黄水、乙醇、己酸等酯化前体物质添加量对酯化液制备的影响。结果表明,该红曲霉优化发酵培养基组分(以100 mL计):大米粉7.0 g,大豆蛋白胨2.0 g,NaNO30.2 g,KH2PO40.15 g,MgSO4·7H2O 0.1 g,培养基初始pH值5.0,接种量10%(体积分数)。在此条件下,添加体积分数为10%的黄水和体积分数为4%的乙醇,酶活力可达745.80 U/mL。向酯化酶液中加入体积分数为0.8%的己酸和体积分数为20%的乙醇继续培养1 d后,酯化液中己酸乙酯的体积分数可达0.121%;而向酯化酶液中补加体积分数为10%的黄水后,己酸乙酯和乳酸乙酯的体积分数分别可达0.055%和0.032%。该结果旨在为将黄水资源用于白酒增香提供理论参考。     

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.     

Optimization of growth conditions of Lentinus edodes mycelium on corn processing waste using response surface analysis

This research was conducted to evaluate the use of corn processing waste as an alternative growth medium for the cultivation of Lentinus edodes mycelium and to determine the optimum growth conditions under solid-state cultivation. The substrate concentration, pH, and temperature for maximizing the growth rate of L. edodes mycelium, 9.3+/-0.6 mm/d, were 44.3 g/l, 4.7, and 24.7 degrees C, respectively. Therefore, the results suggest that corn processing waste can be utilized as a growth substrate for cultivating L. edodes mycelium.     

Glucose Oxidase/Catalase Solution for On-board Control of Shrimp Microbial Spoilage: Model Studies

The potential of an immersion system of glucose oxidase (GOX, 1 unit/ml) and catalase (CAT) added to 4% w/v glucose in artificial seawater was determined for on-board shrimp preservation. Fresh shrimp were frozen, radiation-sterilized, thawed by adding artificial seawater and inoculated with Pseudomonas fluorescens (lo⁴ CFU/g shrimp). Samples were stored at 1°C and treated after 24 hr or left as controls. Changes in shrimp and solution were monitored by total plate counts, ammonia and total volatile nitrogen. Solution discoloration due to shrimp melanosis was followed spectrophotometrically. Microbial lag phase was extended 44 5 d and after 14 d, GOX/CAT had reduced browing by ～80% and inhibited ammonia and total volatile nitrogen production. Due to increase in nitrogen compounds, the enzyme solution should be replaced after 14 days.     

枯草芽孢杆菌发酵麦糟制备蛋白肽培养基研究

以麦糟为原料,对枯草芽孢杆菌（Bacillus subtilis）固态发酵麦糟制备蛋白肽培养基条件进行研究。通过单因素试验考察碳源种类、碳源添加量、料液比、磷酸二氢钾添加量和拌料水pH值对肽得率的影响,通过Box-Behnken试验方案和响应面分析优化培养基。结果表明,枯草芽孢杆菌发酵生产麦糟蛋白肽的最优培养基为：pH 7.0,葡萄糖添加量4.2%,麦糟与水的料液比1.0∶2.1（g∶mL）,磷酸二氢钾添加量0.6%,在该条件下,发酵麦糟肽得率可达12.40%。     

紫色红曲霉FBKL3.0018液态发酵产红曲色素条件的优化研究

以分离自贵州某浓香型酒厂中温大曲的紫色红曲霉（Monascus purpureus）FBKL3.0018为研究对象,以发酵液中红曲色素色价为考察指标,对紫色红曲霉FBKL3.0018产红曲色素的发酵培养基配方进行优化。考察了碳源、氮源、无机盐、生长因子及初始pH值对红曲色素生产的影响,选取对红曲色素生产影响较显著的蛋白胨、FeSO4和初始pH进行响应面优化试验。得到最佳培养基配方为：葡萄糖60 g/L、蛋白胨26 g/L、FeSO4 0.9 g/L、L-谷氨酸 2 g/L和初始pH 4.5。在此优化条件下,红曲色素色价为105.22 U/mL,比优化之前（33.62 U/mL）提高了3.13倍。同时,通过验证试验,实际值105.22 U/mL与预测值108.82 U/mL相对误差为0.97%,说明所建立的回归模型可靠。     

黑曲霉发酵花生壳提取水溶性膳食纤维

以花生壳为原料,应用黑曲霉固态发酵制备花生壳水溶性膳食纤维。通过对黑曲霉菌株特性及其培养基优化的研究,确定培养温度、菌龄、接种量、培养时间的最佳水平,响应面辅助法获得花生壳水溶性膳食纤维的最佳提取工艺条件为：以8g 花生壳为原料,水1.88mL/g 原料,(NH4)2SO4 1.88g/100mL、KH2PO4 1.88g/100mL和MgSO4 5.63g/100mL 的优化培养基,培养温度27℃、黑曲霉菌龄2.9d、接种量16mL、培养时间9.1d,黑曲霉发酵液水解花生壳酶解率可以达到11.03%,水溶性膳食纤维中已糖的聚合度为152.71%,综合评分为105.48。     

Cultural condition affecting the growth and production of beta-galactosidase by Bifidobacterium longum CCRC 15708 in a jar fermenter

In the present study, the growth and production of beta-galactosidase by Bifidobacterium longum CCRC 15708 in a 5-L jar fermenter as influenced by cultivation temperature (27-42 degrees C), medium pH (4.5-7.5) and agitation speed (5-200 rpm) were evaluated. In general, it was found that a cultivation temperature of 37 degrees C proved optimal for both growth and beta-galactosidase production by the test organism. Although the growth of the test organism was the highest in the culture with pH controlled at 4.5-6.5, the culture with pH controlled at 6.5 resulted in the highest production of beta-galactosidase. Further, agitation at 100 rpm or more was found to enhance both the growth and production of beta-galactosidase. Fermentation conducted in a jar fermenter having the pH of the culture medium, the cultivation temperature, and the agitation speed controlled at 6.5, 37 degrees C, and 100 rpm, respectively, a maximum beta-galactosidase activity of 36.7 U/ml and a maximum transgalactosylation activity of 0.49 U/ml was achieved in 10 h of fermentation. There are ca 2.0 and 12.3 fold greater than the reported maximum beta-galactosidase and transgalactosylation activity, respectively, produced by B. longum CCRC 15708 in a flask culture system.     

Use of proteolytic enzymes to facilitate the recovery of chitin from shrimp wastes

Abstract

Two proteolytic enzymes, chymotrypsin and papain, were used to hydrolyze proteins associated with demineralized shrimp waste to recover chitin. The conditions used were optimized using response surface methodology (RSM). The factors studied were temperature (3 levels), pH (3 levels) and enzyme to waste ratio (E/W ratio)(5 levels). Of these, pH was the most important factor in the models obtained. Optimum conditions for deproteinization by chymotrypsin were determined to be around 40°C, pH 8.0 and E/W ratio of 7.1000 (w:w). With papain, a temperature of around 38°C, pH of 8.7 and E/W ratio of 10:1000 (w:w) gave the optimum response. At these conditions, the yield of protein was maximum, and ranged from 22–48% depending on the starting material. The residual protein levels in the waste after deproteinization with the enzymes were very low, i.e., 1.3 and 2.8% for chymotrypsin and papain‐treated samples, respectively.     

Investigation of the utility of pineapple juice and pineapple waste material as low-cost substrate for ethanol fermentation by Zymomonas mobilis

The utility of the juice of rotten or discarded pineapples and the waste material of the production of pineapple juice as low-cost substrates for ethanol production by Zymomonas mobilis was investigated. Z. mobilis ATCC 10988 produced 59.0 g.l(-1) ethanol in undiluted pineapple juice without nutritional supplementation and without the regulation of the pH while 42.5 g.l(-1) ethanol was obtained using a 125 g.l(-1) sucrose medium supplemented with 10 g.l(-1) yeast extract and mineral salts. Ethanol fermentation using unhydrolyzed and enzymatically hydrolyzed pineapple waste material was also investigated under various culture conditions. When a 15% (v/v) dilution of unhydrolyzed waste material without nutritional supplementation was used, more than 3.5 g.l(-1) ethanol was produced. When the media containing 15, 30, and 40% (v/v) of the hydrolyzate consisting of a 60% (v/v) suspension of pineapple waste material were used, final concentrations of ethanol were 5.0 g.l(-1), 7.6 g.l(-1), and 9.3 g.l(-1), respectively. These results suggest that pineapple juice and the waste material can be useful low-cost substrates for ethanol production by Z. mobilis without supplementation with expensive organic nitrogen complexes such as yeast extract and without the regulation of the pH during cultivation, leading to the reduction in the production costs.