Separation and utilization of pectin lyase from commercial pectic enzyme via highly methoxylated cross-linked alcohol-insoluble solid chromatography for wine methanol reduction

The isolation and utilization of pectin lyase (PL) from commercial pectic enzyme for methanol reduction in wine production was investigated. PL can be separated from pectinesterase (PE) and polygalacturonase (PG) on HM-CL-AIS affinity chromatography at pH 4; however, it is difficult to further distinguish PE from PG. Some desirable physicochemical properties such as transmittance, lightness, redness, and lower total pectin content are found in the external enzyme adding groups (PL, PE and PG, and pectic enzyme groups) in comparison to the control group. Methanol contents in pectic enzyme and the PE and PG groups increase from 628 +/- 13 (control group) to 3103 +/- 16 and 1736 +/- 67 mg/L ethanol in the final products, respectively. Nevertheless, the adding of PL does not cause any increase in methanol content. The results present in this study suggest that the HM-CL-AIS column is a simple, inexpensive, convenient, and effective method for PL purification. Moreover, the partial purified PL is a potential replacement of commercial pectic enzyme for pectin depolymerizing, methanol content reducing, and wine quality improving in wine production.     

Fate of mucilage cell wall polysaccharides during coffee fermentation.

Effects of a 20-h fermentation on cell wall polysaccharides from the mucilage of pulped coffee beans were examined and compared to those of unfermented beans, on alcohol insoluble residues (AIRs), their hot-water-soluble crude pectic substances (PECTs), and their hot-water-insoluble residues (RESs). Yields and compositions were very similar: AIRs, which consisted of approximately 30% highly methylated pectic substances, approximately 9% cellulose, and approximately 15% neutral noncellulosic polysaccharides, exhibited no apparent degradation. However, PECTs from fermented beans were shown to have undergone a slight reduction of their intrinsic viscosity and weight-average molecular weight by capillary viscosimetry and high-performance size-exclusion chromatography. After fermentation, hot-water-insoluble pectic substances of RES exhibited partial de-esterification. Removal of coffee bean mucilage by natural fermentation seems to result from a restricted pectolysis, the mechanism of which remains to be elucidated.     

Antioxidant activity and emulsion-stabilizing effect of pectic enzyme treated pectin in soy protein isolate-stabilized oil/water emulsion

The antioxidant activity of pectic enzyme treated pectin (PET-pectin) prepared from citrus pectin by enzymatic hydrolysis and its potential use as a stabilizer and an antioxidant for soy protein isolate (SPI)-stabilized oil in water (O/W) emulsion were investigated. Trolox equivalent antioxidant capacity (TEAC) was found to be positively associated with molecular weight (M(w)) of PET-pectin and negatively associated with degree of esterification (DE) of PET-pectin. PET-pectin (1 kDa and 11.6% DE) prepared from citrus pectin after 24 h of hydrolysis by commercial pectic enzyme produced by Aspergillus niger expressed higher α,α-diphenyl-β-picrylhydrazyl (DPPH) radical scavenging activity, TEAC, and reducing power than untreated citrus pectin (353 kDa and 60% DE). The addition of PET-pectin could increase both emulsifying activity (EA) and emulsion stability (ES) of SPI-stabilized O/W emulsion. When the SPI-stabilized lipid droplet was coated with the mixture of PET-pectin and pectin, the EA and ES of the emulsion were improved more than they were when the lipid droplet was coated with either pectin or PET-pectin alone. The amount of secondary oxidation products (thiobarbituric acid reactive substances) produced in the emulsion prepared with the mixture of SPI and PET-pectin was less than the amount produced in the emulsion prepared with either SPI or SPI/pectin. These results suggest that PET-pectin has an emulsion-stabilizing effect and lipid oxidation inhibition ability on SPI-stabilized emulsion. Therefore, PET-pectin can be used as a stabilizer as well as an antioxidant in plant origin in SPI-stabilized O/W emulsion and thus prolong the shelf life of food emulsion.     

Enzymatic degradation of cell wall polysaccharides from mango (Mangifera indica L.) puree

Ripe mango puree (Smith cultivar) was treated with fungal polysaccharidases containing pectinolytic, hemicellulolytic, and cellulolytic activities for 2 h at 50 degrees C. A loss of 30% of the cell wall material (CWM) was measured. CWM polysaccharides were hydrolyzed to varying degrees: 88, 65, and 65% of, respectively, galacturonic acid-, arabinose-, and rhamnose-containing polymers were hydrolyzed, whereas 50% of cellulose was degraded. After 30 min of treatment, the ethanol precipitation test on the serum was negative, indicating that pectic substances were rapidly hydrolyzed. Oligogalacturonic acids (degree of polymerization, 1-12) were observed in the serum. A viscosity drop of 90% was measured after 2 h, confirming the dominant role of pectic substances in puree viscosity.     

Pectic methyl and nonmethyl esters in potato cell walls.

Because pectins are released from potatoes and other plants under conditions that cleave ester linkages, it has been suggested that there are other galaturonoyl ester cross-links between pectin chains in addition to the known non-cross-linking methyl esters. A microscale titration method and a copper binding method were developed for the measurement of total polymer carboxyl (essentially pectic) ester content in potato cell walls. Relative to the uronic acid content of the cell walls, the degree of total esterification was 57-58%. Comparison with levels of methanol released on ester hydrolysis allowed nonmethyl uronoyl esters to be estimated to be 14-15% relative to total uronic acid. The possibility of nonmethyl-esterified linkages being formed in potato cell walls by a side-reaction catalyzed by pectin methyl esterase (PME) was investigated, but no increase in nonmethyl-esterified pectin was observed under conditions where pectin was being effectively de-esterified by endogenous PME activity.     

Effect of fermentation and autoclaving on dietary fiber fractions and antinutritional factors of beans (Phaseolus vulgaris L.)

The effect of fermentation on antinutritional factors and also on total dietary fiber (TDF), insoluble (IDF) and soluble (SDF) dietary fiber fractions was studied in beans (Phaseolus vulgaris L.). The processes studied were two types of fermentation (lactic acid and natural), and a portion of the obtained flours were processed by autoclaving. The dietary fiber (DF) content and its components were determined using the enzymatic-gravimetric and enzymatic-chemical methods. The TDF content ranged from 24.5% dry matter (DM) in the raw to 25.2% DM in the processed beans. All the processing treatments significantly decreased the SDF content, and irrelevant changes were noticed in the IDF content of processed beans. Cellulose content of all samples was reduced by the processing treatments. Correspondingly, higher amounts of resistant starch was observed in the processed beans, except in the lactic acid fermented ones. However, the levels of pectic polysaccharides and Klason lignin were higher in the samples fermented by Lactobacillus plantarum. The action of microorganisms was determinant for the different degradation of the bean cell wall, disrupting the protein-carbohydrate integration, thus reducing the solubility of DF.     

Change in particle size of pectin reacted with pectinesterase isozymes from pea (Pisum sativum L.) sprout

Four pectinesterase (PE) isozymes were isolated by CM-Sepharose CL-6B chromatography from etiolated pea (Pisum sativum L.) sprouts and then reacted with citrus pectin (degree of esterification = 68%, 30-100 kDa) to observe the change in pectin particle size using a laser particle size analyzer. After incubation of a pectin-PE mixture (pH 6.5) at 30 degrees C for 4 h, PE 1 was observed to catalyze the transacylation reaction most remarkably, increasing the particle size from approximately 50-70 to approximately 250-350 nm, followed by PE 3, PE 2, and PE 4.     

Evaluation and Strategies to Improve Fermentation Characteristics of Modified Dry‐Grind Corn Processes

New corn fractionation technologies that produce higher value coproducts from dry‐grind processing have been developed. Wet fractionation technologies involve a short soaking of corn followed by milling to recover germ and pericarp fiber in an aqueous medium before fermentation of degermed defibered slurry. In dry fractionation technologies, a dry degerm defiber (3D) process (similar to conventional corn dry‐milling) is used to separate germ and pericarp fiber before fermentation of the endosperm fraction. The effect of dry and wet fractionation technologies on the fermentation rates and ethanol yields were studied and compared with the conventional dry‐grind process. The wet process had the highest fermentation rate. The endosperm fraction obtained from 3D process had lowest fermentation rate and highest residual sugars at the end of fermentation. Strategies to improve the fermentation characteristics of endosperm fraction from 3D process were evaluated using two saccharification and fermentation processes. The endosperm fraction obtained from 3D process was liquefied by enzymatic hydrolysis and fermented using either separate saccharification (SS) and fermentation or simultaneous saccharification and fermentation (SSF). Corn germ soak water and B‐vitamins were added during fermentation to study the effect of micronutrient addition. Ethanol and sugar profiles were measured using HPLC. The endosperm fraction fermented using SSF produced higher ethanol yields than SS. Addition of B‐vitamins and germ soak water during SSF improved fermentation of 3D process and resulted in 2.6 and 2.3% (v/v) higher ethanol concentrations and fermentation rates compared with 3D process treatment with no addition of micronutrients.     

Transacylation and de-esterification reactions of pectin as catalyzed by pectinesterases from tomato and citrus

The optimal conditions for the de-esterification reaction of tomato pectinesterase (PE) and citrus PE was 0.1-0.2 M NaCl and at pH 7.5-8.5, 65 degrees C, almost identical to those for the transacylation reaction as observed by turbidity (absorbance at 400 nm) change. Among the PEs tested, pea pod PE presented the most remarkable catalysis on the transacylation reaction, and 1.5% pectin solution was determined to be suitable for this reaction. Low methoxy pectin with a DE (degree of esterification) of 31% displayed a slow turbidity increase, revealing that the extent of DE was influential on the transacylation. Besides citrus pectin, apple pectin was also proved to progress transacylation reaction by PEs from tomato and citrus sources as apparently observed by turbidity method.     

Characterization of pectinesterase inhibitor in jelly fig (Ficus awkeotsang Makino) achenes

Pectinesterase inhibitor (PEI) extract prepared from intact jelly fig (Ficus awkeotsang Makino) achenes was separated by membrane (MWCO 3 and 10 kDa) and fractionated by a Sepharose G-50 gel permeation chromatography. Results from Sepharose G-50 gel permeation chromatography and concanavalin A Sepharose chromatography revealed PEI as polypeptides with molecular weights ranging from 3.5 to 4.5 kDa. Incubation of a PE (1 unit/mL)-PEI (2 mg/mL) mixture for 1 min decreased the PE activity by approximately 50%. On the basis of the results of Lineweaver-Burk double-reciprocal plots, Michaelis constant (K(m)) and V(max) values for jelly fig achenes PE (pH 6.0, 30 degrees C) were 0.78 mM -OCH3 and 1.09 microequiv of -COOH/min, respectively. In addition, PEI competitively inhibited both citrus and jelly fig achenes PEs.     

Stabilization of caseinate-covered oil droplets during acidification with high methoxyl pectin

Polysaccharides are widely used in the food industry to modify the stability of protein-based drinks. However, an in depth knowledge of the interactions occurring in the system is still lacking. In this study, the interactions between sodium caseinate and high methoxyl pectin under acidification conditions were studied nondestructively and without dilution using transmission diffusing wave spectroscopy. Oil-in-water emulsions were prepared with 10% soybean oil and 0.5% sodium caseinate. Various concentrations of pectin (ranging from 0 to 0.2%) were added, and the emulsions were acidified with glucono-delta-lactone. With acidification, a "sol-gel" transition occurred and emulsions containing pectin were more stable at lower pH than those without pectin. Furthermore, the sol-gel transition of the mixtures was more sudden for control emulsions without pectin. While in control samples the final solidlike emulsion after gelation tended to be more inhomogeneous and more dissimilar to the starting emulsion, emulsions with pectin in solution gelled later under acidification. With a sufficient amount of pectin, the emulsions showed no aggregation and the destabilization pH varied depending on the amount of pectin present in the emulsions. At intermediate pH values (pH > 5.5), the emulsions displayed a decrease in particle size, more pronounced in samples containing pectin. The results collected using light scattering in concentrated systems, 10% (v/v) in our case, suggested that pectin stabilizes the emulsion oil droplets forming a network of oil droplets loosely connected by strands of pectin.     

产脂肪酶乳酸菌对羊肉发酵香肠脂肪酸的影响

脂肪酶在肉制品加工中会影响风味物质的积累与产生,发酵肉制品生产中常通过添加产脂肪酶菌株来改善制品的风味。通过乳酸菌在代谢过程中分解三丁酸甘油酯产生透明圈的大小,判断菌株产脂肪酶的活力,同时结合脂肪酶基因Lip0069、Lip0893表达量的变化情况,筛选产脂肪酶能力较高的乳酸菌,研究该菌株对发酵香肠中脂肪酸的影响。结果表明,瑞士乳杆菌TR1-1-3代谢过程中产酶活力最强,编码脂肪酶的基因Lip0069、Lip0893表达量最高。发酵香肠试验组共检测到34种脂肪酸,其中饱和脂肪酸（Saturated Fatty Acids, SFA）16种、单不饱和脂肪酸酸（Monounsaturated Fatty Acids, MUFA）9种和多不饱和脂肪酸（Polyunsaturated FattyAcids, PUFA）9种。含量较高的MUFA主要是油酸（C18:1C9）、棕榈油酸（C16:1）、十七碳烯酸（C17:1）、肉豆蔻烯酸（C14:1）。含量较高的PUFA主要是亚油酸（C18:2C9）、亚麻酸（C18:3N6、C18:3N3）、花生四烯酸（C20:4）、二十二碳六烯酸（C22:6,Docosahexaenoic Acid,DHA）。TR1-1-3和ZF22组发酵2 d后,MUFA的含量明显上升（P<0.05）。12 d时5个试验组MUFA的含量均达到最高,其中TR1-1-3增长比例最高,且与ZR、M组差异显著（P<0.05）。TR1-1-3组发酵2 d后PUFA的含量发生了明显的变化（P<0.05）,12 d时TR1-1-3组 PUFA的含量增长了54.17%,且与其他试验组相比差异显著（P<0.05）。乳酸菌TR1-1-3可降低产品中SFA在脂肪酸中所占比例,明显加快MUFA、PUFA的增加速度及其含量,可为研究改善香肠脂肪特性提供良好的前景。     

Combined enzymatic and high-pressure processing affect cell wall polysaccharides in berries

The effect of high-pressure processing (HPP) on cell wall polysaccharides in berries was investigated. HPP decreased the degree of methyl esterification (DM), probably by activation of pectin methyl esterase (PME), and improved the extractability of pectins. When commercial enzyme mixtures were added to mashed berries, a synergistic effect was observed between treatment with commercial enzymes and HPP. Compared to treatment at atmospheric pressure, pectic polysaccharides were degraded to a larger extent when HPP was used. In contrast, hemicelluloses were hardly affected by the added enzymes when HPP was included, although they were degraded during similar treatment at atmospheric pressure. Additionally, the activity of rhamnose-releasing enzymes present in minor quantities might be enhanced after HPP, resulting in a decrease of rhamnose in the polymeric cell wall material. These results exploring the effect of HPP at representative conditions clearly point out the potential of HPP for polysaccharide modification.     

酶解牛乳蛋白多肽复合果汁乳酒工艺研究

为克服传统乳酒缺陷,该文采用蛋白酶水解法及酒精酵母乳酸菌混合发酵法研究了牛乳蛋白多肽复合果汁乳酒的生产工艺。通过对比和正交试验确定了牛乳蛋白的最佳水解酶、水解条件及牛乳蛋白多肽复合果汁乳酒的发酵条件和配方。结果表明：牛乳蛋白的最佳水解酶是中性蛋白酶,最佳水解条件是：酶用量140 mg/L,温度50℃,pH 6.5,酶解5 h,牛乳蛋白水解最彻底;牛乳蛋白多肽复合果汁乳酒的最佳发酵条件为经中性蛋白酶水解的牛乳,用蔗糖调整其糖度为15%,再配以4%的猕猴桃浓缩汁,添加0.35％ kefir发酵剂在25℃发酵12 h,发酵液中的酒精及总酸含量较高;牛乳蛋白多肽复合果汁乳酒的最佳配方组成是：发酵液60%、糖6%、酸0.28%、甜味剂0.1%、稳定剂0.5%、复合香精0.06%、风味增强剂8 mg/L,乳酒品质最好。     

茶叶籽贮藏时间对毛油产率与质量的影响

为了揭示茶叶籽贮藏时间对发酵法茶叶籽毛油产率与质量的影响,每2周从贮藏的茶叶籽中取样,利用茶叶籽油发酵法生产工艺进行茶叶籽毛油生产,并对工艺中各项剩余物的含油量及毛油的重要质量指标进行了测定,结果如下。室温条件下,茶叶籽贮藏47周后,毛油产率下降了23.5%、酸值及过氧化值分别升高了44.88%及69.4%,毛油色泽基本没有变化。滤渣、发酵沉淀的质量分别升高了20.27%及23.35%;淀粉、油渣质量分别降低了6.13%及3.64%。滤渣、发酵沉淀、淀粉及油渣含油率分别升高了15.63%、22.77%、206%及12.88%。发酵沉淀质量对毛油产率的影响是正向的,淀粉、油渣及滤渣质量对毛油产率的影响是负向的;发酵沉淀及滤渣含油率对毛油产率的影响是正向的,淀粉及油渣含油量对毛油产率的影响是负向的,影响大小的排序为:油渣淀粉滤渣发酵沉淀。综合分析表明,滤渣是通过滤渣质量的增加导致毛油产率随贮藏时间下降的,其下降作用的贡献占全部下降因素的79.28%。贮藏47周后的茶叶籽仁,利用发酵法生产工艺仍然具有毛油生产价值。该研究可为茶叶籽油合理生产提供理论支撑。     

Purification of fusaproliferin from cultures of Fusarium subglutinans by preparative high-performance liquid chromatography

Thirty-six Fusarium strains were grown on cracked yellow corn and evaluated for optimum fusaproliferin production, with Fusarium subglutinans E-1583 producing the highest levels (1600 microg/g). Three solvent systems were tested for extracting fusaproliferin from the cultures of F. subglutinans E-1583. Methanol gave the highest fusaproliferin recovery, followed by methanol/1% aqueous NaCl (55:45, v/v) and acetonitrile/methanol/H(2)O (16:3:1, v/v/v). Hexane partitioning was effective in removing many impurities from the crude fusaproliferin extracts prior to the liquid chromatography step. Fusaproliferin samples were further purified by high-performance liquid chromatography (HPLC) with a C18 preparatory column using a mobile phase of acetonitrile/H(2)O (80:20, v/v). The purity of the fusaproliferin was verified by analytical HPLC, GC/MS, (1)H NMR spectroscopy, and electrospray ionization (ESI) MS. The isolated fusaproliferin was shown to be free of impurities and can be used as a standard for routine analysis. Fusaproliferin was shown to be temperature-sensitive when samples were stored at room temperature (20-24 degrees C) for more than several days. After 30 days at 4 degrees C, approximately 8% of the fusaproliferin had been transformed to deacetyl-fusaproliferin; however, samples stored at -20 degrees C for 1 year contained only trace amounts of the deacetylated form.     

啤酒糟生产菌体蛋白饲料的试验研究

利用啤酒糟中的残糖和碳水化合物，适量补充氮源和菜籽粕，采用微生物固态发酵方法生产菌体蛋白饲料。试验结果表明：产品比发酵前粗蛋白提高10％以上，粗纤维降解率达10％以上，16种氨基酸总和占粗蛋白总量的比例达到了82%以上。试验证明啤酒糟为主要原料生产菌体蛋白饲料对有效利用啤酒糟资源和开发饲料蛋白具有重要的意义。     

Boron-Calcium Synergically Alleviates Aluminum Toxicity in Wheat Plants (Triticum aestivum L.)

The effects of B and Ca treatments on root growth, nutrient localization and cell wall properties in wheat ( Triticum aestivum L.) plants with and without Al stress were investigated. Seedlings were grown hydroponically in a complete nutrient solution for 7 d and then treated with B (0, 40 μM), Ca (0, 2,500 μM), and Al (0, 100 μM) in a 500 μM CaCl₂ solution for 8 d. The cell wall materials (CWM) were extracted with a phenol: acetic acid: water (2:1:1 w/v/v) solution and used for subsequent pectin extraction with trans -1,2-diami-nocyclohexane- N,N,N,N -tetraacetic acid (CDTA) and Na₂CO₃ solutions. Boron, Ca, and B + Ca treatments enhanced root growth by 19.5, 15.2, and 27.2%, respectively, compared to the control (pH 4.5). Calcium and B+Ca treatments enhanced root growth with Al stress by 43 and 54%, respectively, while B did not exert any effect. The amounts of CWM and pectin per unit of root fresh weight increased by Al treatment, whereas the Ca and B+Ca treatments slightly reduced the contents of these components. Seventy-four percent of total B, 69% of total Ca, and 85% of total Al were located in the cell wall in the B, Ca, and Al treatments, respectively and 32% of total B, 33% of total Ca, and 33% of total Al were located in the CDTA-soluble and Na₂CO₃-soluble pectin fractions. A more conspicuous localization of B was observed in the presence of Al. Aluminum treatment markedly decreased the Ca content in the cell wall as well as pectin fractions, mainly in the case of the CDTA-soluble pectin fraction. Boron + Ca treatment decreased the Al content in the cell wall and pectin fractions compared to the Ca treatment alone in the presence of Al. It is concluded that the B+Ca treatment enhanced root growth and, B and Ca uptake, and helped to maintain a normal B and Ca metabolism in the cell walls even in the presence of Al.     

Producing docosahexaenoic acid (DHA)-rich algae from biodiesel-derived crude glycerol: effects of impurities on DHA production and algal biomass composition

Crude glycerol is the primary byproduct of the biodiesel industry. Producing docosahexaenoic acid (DHA, 22:6 n-3) through fermentation of the alga Schizochytrium limacinum on crude glycerol provides a unique opportunity to utilize a large quantity of this byproduct. The objective of this work is to investigate the effects of impurities contained in the crude glycerol on DHA production and algal biomass composition. Crude glycerol streams were obtained from different biodiesel refineries. All of the glycerol samples contained methanol, soaps, and various elements including calcium, phosphorus, potassium, silicon, sodium, and zinc. Both methanol and soap were found to negatively influence algal DHA production; these two impurities can be removed from culture medium by evaporation through autoclaving (for methanol) and by precipitation through pH adjustment (for soap). The glycerol-derived algal biomass contained 45-50% lipid, 14-20% protein, and 25% carbohydrate, with 8-13% ash content. Palmitic acid (C16:0) and DHA were the two major fatty acids in the algal lipid. The algal biomass was rich in lysine and cysteine, relative to many common feedstuffs. Elemental analysis by inductively coupled plasma showed that boron, calcium, copper, iron, magnesium, phosphorus, potassium, silicon, sodium, and sulfur were present in the biomass, whereas no heavy metals (such as mercury) were detected in the algal biomass. Overall, the results show that crude glycerol was a suitable carbon source for algal fermentation. The crude glycerol-derived algal biomass had a high level of DHA and a nutritional profile similar to that of commercial algal biomass, suggesting a great potential for using crude glycerol-derived algae in omega-3-fortified food or feed.