Chongde Wu

Progress in engineering acid stress resistance of lactic acid bacteria

Lactic acid bacteria (LAB) are widely used for the production of a variety of fermented foods, and are considered as probiotic due to their health-promoting effect. However, LAB encounter various environmental stresses both in industrial fermentation and application, among which acid stress is one of the most important survival challenges. Improving the acid stress resistance may contribute to the application and function of probiotic action to the host. Recently, the advent of genomics, functional genomics and high-throughput technologies have allowed for the understanding of acid tolerance mechanisms at a systems level, and many method to improve acid tolerance have been developed. This review describes the current progress in engineering acid stress resistance of LAB. Special emphasis is placed on engineering cellular microenvironment (engineering amino acid metabolism, introduction of exogenous biosynthetic capacity, and overproduction of stress response proteins) and maintaining cell membrane functionality. Moreover, strategies to improve acid tolerance and the related physiological mechanisms are also discussed.

Novel Intestinal-Targeted Ca-Alginate-Based Carrier for pH-Responsive Protection and Release of Lactic Acid Bacteria

A novel intestinal-targeted carrier for pH-responsive protection of lactic acid bacteria in stomach and rapid release of lactic acid bacteria in small intestine is successfully developed. The proposed carrier is composed of a Ca-alginate/protamine (CAP) composite shell and a Lactobacillus-casei-encapsulated Ca-alginate (CA) core. The carriers are prepared simply by a coextrusion minifluidic and subsequent adsorption method. The CAP composite shell offers not only improved protection for Lactobacillus casei to guarantee the endurance and survival in the stomach but also satisfactory intestinal-targeted characteristics to guarantee the rapid release of Lactobacillus casei in the small intestine. In the stomach, where there is an acidic environment, the diffusion channels delineated by the CA networks in the CAP composite shell of the carriers are choked with protamine molecules; as a result, it is hard for the gastric acid to diffuse across the CAP composite shell and thus the encapsulated Lactobacillus casei inside carriers can be efficiently protected. However, when they come to the small intestine, where there is a neutral environment, the carriers dissolve rapidly because of the cooperation between protamine and trypsin; consequently, the encapsulated Lactobacillus casei can be quickly released. The proposed CAP composite carrier provides a novel mode for developing efficient protection systems, responsive controlled-release systems, and intestinal-targeted drug delivery systems.

An Improved and Validated Sample Cleanup Method for Analysis of Ethyl Carbamate in Chinese Liquor

Ethyl carbamate (EC) is a potential human carcinogen widely existing in fermented foods and alcoholic beverages. The solid-phase extraction (SPE) coupled to gas chromatography mass spectrometry is a widely-used method to determine EC levels, but the accuracy varies with sample matrix and the effects of operation parameters are rarely examined. In this study, the influence factors involved in EC determination were investigated using Chinese liquor as sample matrix, and the improved method was further applied. Three types of SPE columns, including diatomite, Florisil, and primary-secondary amine, were compared in extraction efficiency, and the diatomite column exhibited the highest extraction efficiency. The optimal volumes of elution solvents with diatomite column were 15 mL for 3-mL samples solution loaded. In addition, the alcoholic strength for EC determination should be diluted below 20% (v/v) to avoid the enhancement of matrix-induced chromatographic response. Moreover, the pH neutralization could help improve EC recovery and peak resolution, reducing interfering effects. Based on these results, the improved method showed that the limit of detection, the limit of quantification, and average recoveries were 1.10 μg/L, 3.65 μg/L, and 93.06%, respectively. To further elucidate the underlying factors related to EC accumulation, partial least square regression analysis was conducted, and the results suggested that EC levels had the closest relationship with alcoholic strength among the remaining precursors.

Changes in Volatile Compounds of Chinese Luzhou-Flavor Liquor during the Fermentation and Distillation Process.

UNLABELLED The aim of this study was to investigate the dynamic of volatile compounds in the Zaopei during the fermentation and distillation process by headspace solid-phase microextraction-gas chromatography mass spectrometry (HS-SPME-GCMS). Physicochemical properties analysis of Zaopei (fermented grains [FG], fermented grains mixed with sorghum [FGS], streamed grains [SG], and streamed grains mixed with Daqu [SGD]) showed distinct changes. A total number of 66 volatile compounds in the Zaopei were identified, in which butanoic acid, hexanoic acid, ethyl hexanoate, ethyl lactate, ethyl octanoate, hexyl hexanoate, ethyl hydrocinnamate, ethyl oleate, ethyl hexadecanoate, and ethyl linoleate were considered to be the dominant compounds due to their high concentrations. FG had the highest volatile compounds (112.43 mg/kg), which significantly decreased by 17.05% in the FGS, 67.12% in the SG, and 73.75% in the SGD. Furthermore, about 61.49% of volatile compounds of FGS were evaporated into raw liquor, whereas head, heart, and tail liquor accounted for 29.84%, 39.49%, and 30.67%, respectively. Each volatile class generally presented a decreasing trend, except for furans. Especially, the percentage of esters was 55.51% to 67.41% in the Zaopei, and reached 92.60% to 97.67% in the raw liquor. Principal component analysis based ordination of volatile compounds data segregated FGS and SGD samples. In addition, radar diagrams of the odor activity values suggested that intense flavor of fruit was weakened most from FG to SGD. PRACTICAL APPLICATION The dynamic of volatile compounds in the Zaopei during the fermentation and distillation process was tested by SPME-GCMS. The result of this study demonstrated that both volatile compounds of Zaopei and thermal reaction during distillation simply determined the unique feature of raw liquor. This study was conducted based on the real products from liquor manufactory, so it is practicable that the method can be used in an industry setting.

Reduced nitrite and biogenic amine concentrations and improved flavor components of Chinese sauerkraut via co-culture of Lactobacillus plantarum and Zygosaccharomyces rouxii

The aim of this study was to investigate the effect of inoculated fermentation on the quality of Chinese sauerkraut. To this end we studied a co-culture system consisting of Lactobacillus plantarum Shanghai brewing 1.08 and Zygosaccharomyces rouxii CGMCC 3791 during inoculated sauerkraut fermentation. The nitrite concentrations in pickled cabbage and radish inoculated with starter cultures of L. plantarum and Z. rouxii were significantly lower than those in the spontaneous fermentation system during the whole fermentation process. In addition, co-culture of L. plantarum and Z. rouxii during the production of sauerkraut decreased the formation of biogenic amines in the pickled vegetables. Using gas chromatography–mass spectrometry we also compared the levels of volatile compounds in inoculated and naturally fermented Chinese sauerkraut. Sixty compounds were identified, with the sauerkraut inoculated with starter cultures containing overall higher contents of volatile compounds, including acids, alcohols, esters, and phenols. The structure of the microbial community during the production of sauerkraut was studied using phospholipid fatty-acid (PLFA) analysis. This analysis revealed that the brine of inoculated sauerkraut contained significantly higher contents of Gram-positive and fungal PLFAs and a lower content of Gram-negative PLFAs, suggesting that the improved quality of inoculated Chinese sauerkraut may be ascribed to the inhibition of the growth of Gram-positive during sauerkraut fermentation. These results may indicate a new strategy to enhance the quality of Chinese sauerkraut.

Difference of microbial community stressed in artificial pit muds for Luzhou‐flavour liquor brewing revealed by multiphase culture‐independent technology

Artificial pit muds (APMs) is produced by peats, aged pit muds, yellow and black clays etc. and is one of essential factors for Luzhou‐flavour liquor production. The microbial community of APMs significantly influence the quality of Luzhou‐flavour liquor. The aim of this study was to investigate the differences in bacterial, archaeal and fungal community of APMs, starters and materials.

Genomics of lactic acid bacteria: Current status and potential applications

Abstract Lactic acid bacteria (LAB) are widely used for the production of a variety of foods and feed raw materials where they contribute to flavor and texture of the fermented products. In addition, specific LAB strains are considered as probiotic due to their health-promoting effects in consumers. Recently, the genome sequencing of LAB is booming and the increased amount of published genomics data brings unprecedented opportunity for us to reveal the important traits of LAB. This review describes the recent progress on LAB genomics and special emphasis is placed on understanding the industry-related physiological features based on genomics analysis. Moreover, strategies to engineer metabolic capacity and stress tolerance of LAB with improved industrial performance are also discussed.

Formation of ethyl carbamate in Goji wines: Effect of Goji fruit composition

Ethyl carbamate (EC) is a multisite carcinogen widely occurring in alcoholic beverages. In this investigation, solid-phase extraction combined with gas chromatography mass spectrometry was employed to determine EC contents during the fermentation and storage processes, and the effects of Goji varieties on its formation were also examined. The results indicated that natural and potential EC contents were significantly affected by the varied composition of Goji fruits. The analysis of chemical properties showed differences in hundred-grain weight, water contents, amino acids, and nitrogen-to-carbon ratio for Goji berries. Citrulline was completely degraded although it is routinely considered as a non-preferred nitrogen for yeasts. Due to compositional differences, Goji wines accumulated distinct urea levels that positively correlated with the potential EC contents. Furthermore, the temperature in both the production processes highly influenced EC formation. These results contribute to a more comprehensive understanding of EC formation, and in turn, controlling EC in the Goji wine matrix.

Spatial Distribution of Bacterial Communities and Related Biochemical Properties in Luzhou-Flavor Liquor-Fermented Grains

Grain fermenting with separate layers in a fermentation pit is the typical and experiential brewing technology for Chinese Luzhou-flavor liquor. However, it is still unclear to what extent the bacterial communities in the different layers of fermented grains (FG) effects the liquors quality. In this study, the spatial distributions of bacterial communities in Luzhou-flavor liquor FG (top, middle, and bottom layers) from 2 distinctive factories (Jiannanchun and Fenggu) were investigated using culture-independent approaches (phospholipid fatty acid [PLFA] and polymerase chain reaction-denaturing gel electrophoresis [DGGE]). The relationship between bacterial community and biochemical properties was also assessed by Canonical correspondence analysis (CCA). No significant variation in moisture was observed in spatial samples, and the highest content of acidity and total ester was detected in the bottom layer (P < 0.05). A high level of ethanol was observed in the top and middle layers of Fenggu and Jiannanchun, respectively. Significant spatial distribution of the total PLFA was only shown in the 50-y-old pits (P < 0.05), and Gram negative bacteria was the prominent community. Bacterial 16S rDNA DGGE analysis revealed that the most abundant bacterial community was in the top layers of the FG both from Fenggu and Jiannanchun, with Lactobacillaceae accounting for 30% of the total DGGE bands and Lactobacillus acetotolerans was the dominant species. FG samples from the same pit had a highly similar bacterial community structure according to the hierarchal cluster tree. CCA suggested that the moisture, acidity, ethanol, and reducing sugar were the main factors affecting the distribution of L. acetotolerans. Our results will facilitate the knowledge about the spatial distribution of bacterial communities and the relationship with their living environment.

Characterizing Soy Sauce Moromi Manufactured by High-Salt Dilute-State and Low-Salt Solid-State Fermentation Using Multiphase Analyzing Methods

Present study was to characterize the physiochemical properties, free amino acids (FAAs), volatiles and microbial communities of various moromi, respectively sampled from different stages of high-salt dilute-state (HSDS) and low-salt solid-state (LSSS) fermentation, using multiphase analyzing methods. Phospholipid fatty acids (PLFA) analysis indicated that Gram-positive bacteria were dominant bacteria and fungi were principal microbes. For DGGE analysis, dominant microbes in moromi were mainly fell into Weissella, Tetragenococcus, Candida, Pichia, and Zygosaccharomyces. During fermentation, the dominant microbes shifted from nonhalophilic and less acid-tolerant species to halophilic and acid-tolerant species. Total of 15 FAAs and 44 volatiles were identified in moromi, mainly Glu, Asp, Tyr, and acids, alcohols, esters, aldehydes, respectively. Odor activity values analysis suggested that the final moromi of LSSS fermentation had more complicated odors than that of HSDS fermentation. Conclusively, technological parameters, microbial communities, raw materials and fermentation process may result in the discrepancy of HSDS and LSSS moromi.