微生物多级发酵对中药方剂功能活性及有效组分的影响

选用多菌株对一中药方剂（简称JGZF）进行液体多级发酵,考察其在3种处理方式（直接熬制、自然发酵、人工发酵）下微量元素、体外抗氧化活性、抑菌活性以及其他有效组分等结果的变化。采用原子吸收光谱法检测微量元素;紫外-可见分光光度计检测体外抗氧化活性,包括过氧化物歧化酶SOD活性、DPPH自由基的清除能力、羟基自由基的清除能力和总多酚含量;牛津杯抑菌圈法测定抑菌活性;UPLC-Q-TOF-MS/MS非靶标代谢组学检测JGZF的药效成分。结果表明：跟直接熬制相比,中药方剂JGZF经自然发酵和人工发酵处理后,锌、铜、铁和钙的含量得到了显著提高;人工发酵能显著提高JGZF的体外抗氧化活性,其中SOD活性提高了1.5倍,DPPH自由基的清除能力提高了2.2倍,羟基自由基的清除能力提高了4.2倍,总多酚含量提高了2.2倍;发酵后的JGZF对沙门氏菌和大肠杆菌的抑菌活性更强,但对志贺氏菌和金黄色葡萄球菌的抑菌活性不变或略有降低;JGZF经人工发酵后,药液组分含量有增有减,其中人体必需氨基酸L-苯丙氨酸、抗炎物质黄芩素等含量显著增加;五味子苷B、7-羟基香豆素、农药残留等含量有显著降低。     

Alginate beads and apple pieces as carriers for Saccharomyces cerevisiae var. boulardii,as representative of yeast functional starter cultures

This study focused on two different carriers for Saccharomyces cerevisiae var. boulardii for its targeted release as a starter or as a probiotic: the microencapsulation into alginate beads and the immobilisation on apple pieces. Beads were studied in relation to encapsulation yield (EY), viability of cells throughout the storage, kinetic of cell release, survival under conditions that mimic the transit into the gut and in vivo application (fermentation of grape juice). Concerning the microencapsulation, EY was very high (ca. 90%), cells survived for a long period (upon to 90 days), and the beads assured cell survival and their release under conditions that mimic the gut; moreover, the capsules could be used up to ten times to start a model fermentation of grape juice. Apple pieces were a good system for the immobilisation of S. boulardii; they could be proposed successfully as a reusable carrier for a starter culture, as they assured the start of the fermentation of grape juice for at least seven times.     

Alginate and Carrageenan Immobilization Effects on Schizosaccharomyces Pombe Activity and Stability

Calcium alginate and kappa-carrageenan were evaluated as entrapment matrices for whole cells of Schizosaccharomyces pombe. The fermentation activity of immobilized cells was enhanced by preincubation in nutrient medium. Alginate bead activity was significantly better than that with carrageenen beads (p<0.05). In grape must carrageenan beads disintegrated 10 times faster than alginate beads.     

Production of ethanol from liquefied cassava starch using co-immobilized cells of Zymomonas mobilis and Saccharomyces diastaticus

Co-immobilized cells of Saccharomyces diastaticus and Zymomonas mobilis produced a high ethanol concentration compared to immobilized cells of S. diastaticus during batch fermentation of liquefied cassava starch. The co-immobilized cells produced 46.7 g/l ethanol from 150 g/l liquefied cassava starch, while immobilized cells of yeast S. diastaticus produced 37.5 g/l ethanol. The concentration of ethanol produced by immobilized cells was higher than that by free cells of S. diastaticus and Z. mobilis in mixed-culture fermentation. In repeated-batch fermentation using co-immobilized cells, the ethanol concentration increased to 53.5 g/l. The co-immobilized gel beads were stable up to seven successive batches. Continuous fermentation using co-immobilized cells in a packed bed column reactor operated at a flow rate of 15 ml/h (residence time, 4 h) exhibited a maximum ethanol productivity of 8.9 g/l/h.     

Effect of different levels of malt extract on antinutrients, <Emphasis Type="Italic">in vitro</Emphasis> digestibilities and viscosity during fermentation of <Emphasis Type="Italic">Jeungpyun</Emphasis>

The effect of different levels of malt extract on the fermentation of jeungpyun (a Korean fermented steamed rice cake) was investigated. Relative to the control batter, the fermentation rate increased with increasing level of malt extract due to its effect in expediting the fermentation. The contents of total phenols and phytic acid in the jeungpyun batter were reduced during fermentation, and even more so after steaming of the batter. The reduced content of antinutritional factors in the batter was accompanied by increases in in vitro protein and starch digestibilities. The effect of different levels of malt extract on the viscosity and hardness was similar. These results indicate that the addition of malt extract during fermentation may produce a healthier and more nutritious jeungpyun. It also facilitated the production of thin fermented batter, thus affecting the textural characteristics of jeungpyun.     

The Influence of Inoculum Level on Fermentation and Flavour Compounds of White Wines Made from cv. Emir

In this study, the influence of the addition of a commercial wine yeast (Saccharomyces cerevisiae) at inocula of 1 × 10⁴ to 1 × 10⁷ cells/ml in Emir must was investigated with a focus on yeast growth, fermentation rate, ethyl alcohol and flavour compound formation. Spontaneous fermentation without inoculation was also performed. Higher peak counts were observed with higher amounts of S. cerevisiae yeast. Addition of various amounts of yeast led to the earlier disappearance of non‐Saccharomyces yeasts. The fermentation rate was improved with higher amounts of yeast, but ethanol production was not affected. Concentrations of higher alcohols increased with increasing inoculum levels, especially inoculum sizes of 1 × 10⁶ cells/ml and 1 × 10⁷ cells/ml. The amount of ethyl acetate was reduced with increased inoculum levels.     

In vitro digestibility of Bacillus fermented soya bean

Bacillus fermented legume products include among others dawadawa and soumbala made from African locust bean, and natto and kinema made from soya bean. Bacillus subtilis is the dominant species involved in the fermentation. During Bacillus fermentation for 48 h of autoclaved soya bean the quantity of soluble and dialyzable matter increased from 22% and 6% up to 65% and 40%, respectively. Protein and carbohydrate degradation during fermentation of soya bean with several Bacillus spp. was investigated and appeared to be substantial during the first 18 h of fermentation resulting in the release of high levels of peptides and oligosaccharides. In vitro digestibility was increased from 29% up to 33–43% after Bacillus fermentation for 48 h. It was shown that Bacillus spp. were able to degrade soya bean macromolecules to a large extent resulting in water-soluble low molecular weight compounds. In vitro digestion of Bacillus fermented soya bean using gastrointestinal enzymes only slightly increased the amount of dialyzable matter, which clearly demonstrated the beneficial effect of Bacillus fermentation on food nutrient availability.     

Impact of various maceration techniques on the phenolic and volatile composition of Chenin Blanc wines

Chenin Blanc, cultivated in cool climate regions, was chosen to evaluate the impact of different maceration techniques on polyphenolic profiles and volatile compounds. Control with no skin contact (CK), cold maceration (CM), cryogenic maceration (CR), separate fermentation before blending (SFB) and extended skin contact during fermentation (ESF) were the five maceration techniques being studied. SFB and ESF techniques resulted in significantly increased polyphenolic compounds compared to the control (P < 0.05), while CM and CR maceration techniques resulted in almost no effect. Meanwhile, for aromatic components, the maximum change was observed in ESF wine, which had significantly highest levels of terpenes, alcohols, esters and acids. However, compared with control wine, CR wine had higher levels of terpenes and esters and SFB wine had higher levels of terpenes and alcohols. This study provides information about prefermentation techniques and additional options for oenologists to produce Chenin Blanc wines with different characteristics.     

Fermentation of Allium chinense Bulbs With Lactobacillus plantarum ZDY 2013 Shows Enhanced Biofunctionalities,and Nutritional and Chemical Properties

In this study, fermentation of Allium chinense bulbs was carried out with Lactobacillus plantarum ZDY 2013. A decrease in pH from 6.8 to 3.5 and a stable lactic acid bacteria population were observed during 7‐d fermentation. The total phenolic content increased by 2.7‐fold in the aqueous and ethanol extracts of A. chinense bulbs after fermentation. Antioxidant capacity including 2,2‐diphenyl‐1‐picrylhydrazyl radical‐scavenging effect and reducing power of both extracts was significantly (P < 0.05) improved after fermentation. Antagonistic test against 6 pathogens showed that fermentation significantly (P < 0.05) enhanced the antimicrobial activity in both extracts of fermented bulbs, especially in the ethanol extracts of fermented bulbs against L. monocytogenes. Analysis of the free amino acid (FAA) profile by ion‐exchange chromatography revealed that fermentation significantly (P < 0.05) increased total FAA content. In addition, among 27 kinds of volatile components analyzed by headspace–solid phase microextraction–gas chromatography–tandem mass spectrometry, sulfur‐containing compounds accounted for 65.23%, but decreased to 43.65% after fermentation. Our results suggested that fermentation of A. chinense bulbs with L. plantarum could improve their biofunctionalities, and nutritional and chemical properties.     

Biochemical changes during the natural fermentation of the african oil bean (Pentaclethra macrophylla) for the production of Ugba

Changes in the activities of major enzymes and in the proportion of the principal chemical components during the nataral fermentation of ugba from African oil bean (Pentaclethra macrophylla Bentham) were studied. The activities of the enzymes were related to changes in chemical composition. α-Amylase, proteolytic and lipolytic enzyme activity remained detectable throughout the 72-h fermentation period, and reached a maximum at 24-36 h. The levels of amino nitrogen, ammonia nitrogen, reducing sugar, titratable acidity and pH increased with time. A seven-fold increase in the amino nitrogen level occurred during the fermentation. The temperature of the fermentation and the moisture level reached maximum values at 24 h and 48 h of fermentation respectively. These results could provide useful indices for the improved evaluation of ugba fermentation.     

An assessment of the protein quality and vitamin b content of commonly used fermented products of legumes and millets

Fermented products of legumes, Bengal gram dhal (Cicer arietinum) and green gram dhal (Phaseolus aureus), and millets, bajra (Pennisetum typhoideum), jowar (Sorghum vulgare) and ragi (Eleucine coracana), were analysed for protein quality and vitamin B content. Fermentation reduced total crude protein by 6-8 % in the legume products and 4-6% in the millet products of bajra and ragi. No reduction in crude protein was observed in fermented jowar products. Temperature of batters increased considerably on fermentation, while pH decreased. The protein efficiency ratio (PER) and the true digestibility (TD) of legume products were not altered by fermentation, the biological value (BV) and net protein utilisation, (NPU), of Bengal gram dhokla improved significantly (P<0.05). The TD of fermented jowar increased significantly (P < 0.05) but not that of ragi, bajra and the legume products. BV and NPU of both jowar and ragi products increased significantly on fermentation (P<0.05) but not that of bajra. The thiamin and riboflavin contents of both the legume and millet products increased with increase in fermentation time. Steaming and cooking after fermentation reduced the thiamin and riboflavin content. However, fermentation after cooking of millet batters increased the levels of both vitamins markedly.     

乳酸菌固态发酵过程中麦麸组分的动态变化

麦麸是小麦加工的主要副产物,营养丰富且产量大,采用乳酸菌处理麦麸可提高其附加值。为明确乳酸菌发酵对麦麸各组分的影响,作者采用植物乳杆菌、鼠李糖乳杆菌、戊糖片球菌和布氏乳杆菌分别对麦麸进行固态发酵,在48 h内每隔8 h取样,分析可溶性膳食纤维、粗蛋白质、淀粉、总酚、植酸等成分的质量分数及DPPH自由基清除能力的动态变化。结果表明,在麦麸基质中,4株乳酸菌在24 h内生长较为迅速;麦麸经乳酸菌发酵后可溶性膳食纤维质量分数显著提高,其中布氏乳杆菌发酵48 h后可溶性膳食纤维质量分数由4.72%增加至6.58%;随着发酵时间的增加,麦麸中淀粉质量分数逐渐降低,粗蛋白质量分数先增加后降低最后趋于稳定;植物乳杆菌在提高麦麸多酚质量分数方面有更好的效果,多酚质量分数由1.34 mg/g增加至3.86 mg/g,麦麸抗氧化活性显著增加;此外,乳酸菌发酵麦麸可显著降低其植酸质量分数。综合而言,植物乳杆菌和布氏乳杆菌在提高麦麸的营养特性方面具有较好的效果,可有效改善麦麸的综合利用价值。     

Immobilised yeast grape must deacidification in a recycle fixed bed reactor

Maloalcoholic fermentation (MAF) of grape must by Schizosaccharomyces pombe immobilised in calcium‐alginate double‐layer beads (ProMalic^®) was studied in Erlenmeyer flasks and in a total recycle fixed‐bed reactor operating in batch mode. The reaction is pseudo‐first order with respect to l ‐malic acid and under similar conditions deacidification is faster in the recycle reactor. This was attributed to mass transfer limitations which were confirmed in the recycle reactor by studying the influence of yeast load on the rate of MAF. Mass transfer limitations are also responsible for the lower activation energy of fermentation with the immobilised yeast (67 ± 9 kJ mol^?1) in comparison with the free cells (126 ± 19 kJ mol^?1). Alcoholic fermentation and MAF were performed simultaneously, both in the recycle reactor and in the industrial trials, confirming the efficacy of immobilised S. pombe to reduce grape must acidity without interfering with the main fermentation. Altogether, the present results are useful for the scale‐up of a recycle reactor to process large volumes of grape must.     

Fermentation of tea in aqueous suspension

The fermentation of macerated tea leaf in aqueous suspension has been studied both manometrically and in a suitably controlled fermentation vessel. Oxygen consumption proceeded rapidly at a steady rate for an initial period, termed the primary fermentation, then decreased sharply as the availability of simple polyphenol substrates became rate limiting. Maximum reaction rates were observed at 20-35 mg ml^?1 total solids, as a result of substrate inhibition. At higher concentrations of solids reaction rates were also affected by diffusion effects within the leaf particles. At 50 mg ml^?1 total solids and 25°C the system approximated to Michaelis-Menton kinetics with K_m⁰² = 5 × 10^?5M and V_max corresponding to a primary fermentation time (PFT) of 10-15 min. The PFT corresponded with the attainment of maximum total colour and maximum level of theaflavins (TF) (often >30% of total colour). Beyond the PFT total colour remained constant, TFs decreased and non-dialysable pigments increased with time. At fixed levels of dissolved oxygen and solids concentration > 60 mg ml^?1 the proportion of TFs was inversely related to solids concentration. At 50 mg ml^?1 total solids TFs increased to ca 60% of total colour as the dissolved oxygen level was increased to 0.175 mM. As temperature was increased from 15-35°C the reaction rate increased, the proportion of TFs decreased slightly and thearubigins of intermediate molecular weight increased markedly.     

Effect of Cultivar,Temperature, and Environmental Conditions on the Dynamic Change of Melatonin in Mulberry Fruit Development and Wine Fermentation

High levels of melatonin have been reported in various foods but not in mulberry or its wine. This study investigated the dynamic changes of melatonin levels during mulberry fruit development and ethanol fermentation of 2 different colored mulberry cultivars (“Hongguo2? Morus nigra, black and “Baiyuwang? Morus alba, white) at 2 fermentation temperatures (16 and 25 °C). Our results showed that the melatonin level increased in the beginning of mulberry development but decreased in the end. The MnTDC gene expression level correlated with melatonin production, which implied that TDC may be the rate‐limiting enzyme of the melatonin biosynthetic process in mulberries. During mulberry fermentation, the melatonin concentration increased rapidly in the beginning and then decreased gradually. Low temperature delayed the melatonin production during fermentation. A relatively high level of melatonin was found in “Hongguo2? compared with “Baiyuwang? during fruit development and fermentation. The variation of melatonin correlated with the ethanol production rate, suggesting that melatonin may participate in physiological regulation of Saccharomyces cerevisiae during the fermentation stage.     

Preparation of Bifidobacterium breve encapsulated in low methoxyl pectin beads and its effects on yogurt quality

Yogurt is a popular product worldwide partly because of the health-promoting effects of the probiotics that it contains. Probiotics with high survivability constitute a promising direction for fortified yogurt products. This study aimed to prepare Bifidobacterium breve–loaded yogurt with the bacteria surviving transit to the lower part of small intestine or colon. Bifidobacterium breve beads were prepared through an ion-crosslinking method using low methoxyl pectin as the encapsulating material. Features such as encapsulation efficiency and stability during storage and passage through the simulated gastrointestinal tract were studied in vitro. A commercial starter was used for yogurt fermentation, and B. breve with or without encapsulation was added as a probiotic supplement with the starter or 3 to 4 h after fermentation. The effects of B. breve beads on yogurt characteristics were evaluated after different fermentation processes: BC, milk fermented with marketed yogurt starter; UBFF, unencapsulated B. breve added to fresh milk and then fermented; EBFF, encapsulated B. breve added to fresh milk and then fermented; UBAF, unencapsulated B. breve added after fermentation with the starter; and EBAF, encapsulated B. breve beads added 3 to 4 h after fermentation with the starter. Evaluation was based on texture, electronic nose, and electronic tongue analyses. The particle size analysis of B. breve beads showed that they were uniform, mostly spherical, 1 to 1.5 mm in diameter with encapsulating efficiency higher than 99%. Following treatment with the simulated gastrointestinal tract conditions, the number of B. breve decreased by 1.76 and 4.82 log cfu/g for B. breve beads and unencapsulated B. breve, respectively. The EBAF group showed the lowest viscosity (2,235.67 cP) at d 0, and the lower postfermentation degree was reflected by the slow increase in yogurt viscosity. All groups kept a relatively stable pH during storage. The cohesiveness values of the EBAF and UBAF groups were significantly higher than those of the other groups. The trends in texture changes within the BC, UBFF, and EBFF groups were similar, and the UBAF and EBAF groups showed similar trends. In conclusion, B. breve beads showed good stability in vitro and improved yogurt characteristics by increasing the survival rate of the encapsulated cells. Good compatibility of low methoxyl pectin beads with yogurt was also observed.     

Microbial Succession and Metabolite Changes during Long‐Term Storage of Kimchi

Kimchi is often stored for a long period of time for a diet during the winter season because it is an essential side dish for Korean meals. In this study pH, abundance of bacteria and yeasts, bacterial communities, and metabolites were monitored periodically to investigate the fermentation process of kimchi for 120 d. Bacterial abundance increased quickly with a pH decrease after an initial pH increase during the early fermentation period. After 20 d, pH values became relatively stable and free sugars were maintained at relatively constant levels, indicating that kimchi fermentation by lactic acid bacteria (LAB) was almost completed. After that time, a decrease in bacterial abundance and a growth in Saccharomyces occurred concurrently with increased free sugar consumption and production of glycerol and ethanol. Finally, after 100 d, the growth of Candida was observed. Community analysis using pyrosequencing revealed that diverse LAB including Leuconostoc citreum, Leuconostoc holzapfelii, Lactococcus lactis, and Weissella soli were present during the early fermentation period, but the LAB community was quickly replaced with Lactobacillus sakei, Leuconostoc gasicomitatum, and Weissella koreensis as the fermentation progressed. Metabolite analysis using ¹H‐NMR showed that organic acids (lactate, acetate, and succinate) as well as bioactive substances (mannitol and gamma‐aminobutyric acid (GABA)) were produced during the kimchi fermentation, and Leuconostoc strains and Lactobacillus sakei were identified as the producers of mannitol and GABA, respectively. Practical Application In this study, we have shown that the growth inhibition of yeasts including Saccharomyces and Candida is necessary to extend the shelf life of kimchi in long‐term storage. Additionally, we have shown that a mixed culture of Leuconostoc strains and Lactobacillus sakei is necessary to produce kimchi that contains both mannitol and gamma‐aminobutyric acid.     

Mead production: fermentative performance of yeasts entrapped in different concentrations of alginate

Mead is an alcoholic drink known since ancient times, produced by yeast fermenting diluted honey. However, the production of mead has suffered in recent years, partially owing to the lack of scientific progress in this field. In this study, two strains of Saccharomyces cerevisiae, QA23 and ICVD47, were immobilized in 2 or 4% (w/v) alginate beads to assess the most effective alginate concentration for yeast immobilization to produce mead. Neither of the alginate concentrations was able to prevent cell leakage from the beads. The fermentation length was 120 h for both yeast strains. In all cases, at the end of the fermentation, the number of cells entrapped in the beads was higher than the number of free cells, and the total 4% alginate bead wet weight was significantly higher than the 2% alginate bead wet weight. In addition, the evaluation of mead quality showed that the yeast strain had significantly more influence on the physicochemical characteristics than the alginate concentration. Although the yeasts immobilized in the two alginate concentrations were able to perform the fermentation, further research is needed in order to understand the evolution of the yeast population inside the beads throughout the fermentative process. Copyright © 2014 The Institute of Brewing & Distilling     

Effects of mediums on fermentation behaviour and aroma composition in pure and mixed culture of Saccharomyces cerevisiae with Torulaspora delbrueckii

The nutrient status and composition in mediums have a significant effect on yeast metabolism and phenotypic characteristics in wine fermentation. In this study, the effects of three frequently used mediums, including synthetic grape juice (SGJ), grape juice without grape pericarp and seeds (GJ) and grape must with grape pericarp and seeds (GMPS), on yeast fermentation behaviour and aroma compounds produced by pure and mixed culture of Saccharomyces cerevisiae T73 with Torulaspora delbrueckii TD20 were investigated after alcoholic fermentation. The results showed that high fermentation activities and cell population were always found in GJ medium irrespective of inoculated approach. More esters and higher alcohols were produced in GMPS medium fermented by pure S. cerevisiae, while SGJ medium had increased levels of fatty acids. Consistent with previous literatures, the mixed fermentation of T. delbrueckii and S. cerevisiae produced more acetate esters and fatty acids than the pure culture of S. cerevisiae, while this enological trait was only found in SGJ and GJ, not in GMPS. Our results highlighted that more attention should be paid to the fermentation medium when evaluating the enological and aromatic properties of selected yeasts used in industrial winemaking. In this regard, the combined use of GJ and GMPS medium might be a suitable choice.     

Preparation of layer-assembled W/O/W-type microencapsulated beads and application in solid-state fermentation

In this study, microcapsule beads-0–3-layers (M-0–3 indicates microencapsulated beads with 0, 1, 2, and 3 layers) were prepared, their properties were measured and characterized, and the effects of M-0–3 on solid-state fermentation were investigated. The results showed that in a liquid environment, the releasing glucoamylase activities of M-0–3 were 55.77%, 47.67%, 45.85%, and 42.87% in 360 h, respectively. In the solid environment, the reducing sugar production efficiency of M-0–3 was 29.84%, 27.72%, 19.16%, and 15.93% in 15 days, respectively. Adding M-0–3 improved the alcohol and reduced sugar content while decreasing the residual starch content of the Jiupei, indicating that adding M-0–3 was beneficial to the solid-state fermentation of Baijiu. Solid-state fermentation simulation experiments illustrated that microcapsule beads play a positive role in the production of Baijiu, enhancing raw material utilization and yield of Baijiu production.