Optimization of fermentation conditions and quality evaluation of Chaenomeles sinensis glutinous rice wine.

The present study was conducted to optimize fermentation conditions for preparation of Chaenomeles sinensis Glutinous Rice Wine (CRW). The dynamic changes of main substances in the liquor during fermentation process, aroma components, biologically active substances and antioxidant capacity in the CRW after 6 months of aging were tested. The results showed that under optimized conditions, the yield and alcohol content of wine was 44.97 and 20.00%, respectively. After aging, 64 aroma components were detected in the wine, mainly alcohols and esters. The alcohol content of the CRW was 14.8%. Polyphenols and flavonoids reached 0.23 g/L and 0.037 g/L respectively. Furthermore, the CRW had an excellent free radical scavenging ability. Supplementary Information: The online version contains supplementary material available at 10.1007/s13197-024-05934-0.

Effects of Jiuqu inoculating Rhizopus oryzae Q303 and Saccharomyces cerevisiae on chemical components and microbiota during black glutinous rice wine fermentation.

In this study, the physicochemical properties, metabolites, sensory characteristics, and microbiota of black glutinous rice wine (BGRW) during traditional fermentation (TF) and inoculated fermentation (IF) were investigated, and their correlation relationships were revealed. Results indicated that IF promoted total sugar utilization and ethanol and esters synthesis in BGRW. Altogether 67 reliable metabolites were identified, 36 of which were labeled components, primarily esters (OAV > 1). Meanwhile, the concentrations of ethanol, esters, and total flavor on the 11th day of IF were close to those on the 22nd day of TF. Combined with the sensory description indicated that the IF could improve the quality of BGRW. Besides, TF contains vast unfavorable microorganisms, such as Chaetomium, Penicillium, Stachybotrys, and Trichocladium. Therefore, IF can accelerate the fermentation and enhance the flavor of BGRW, and inhibit the growth of pathogenic and spoilage organisms. Finally, Pearson's correlation analysis demonstrated that Saccharomyces, Rhizopus, Lactobacillus, Aspergillus, and Bacillus were the pivotal functional microorganisms in BGRW. The study provided scientific and effective basis for improving the quality, shortening the fermentation cycle, and controlling pathogenic and spoilage organisms of BGRW.

The effects of an innovative pulping technique of synchronously pulping and gelatinizing treatment on raw materials properties, oenological parameters, fermentation process, and flavor characteristics of glutinous rice wine.

Liquid-state fermentation has been increasingly applied in the industrial glutinous rice wine (GRW) production. However, products brewed by this emerging technique possess some deficiencies in flavor quality. Therefore, this study firstly developed and optimized an innovative pulping technique by the synchronously pulping and gelatinizing treatment (Process I) to improve GRW flavor quality, and then revealed the influences of Process I on raw materials properties, oenological parameters, fermentation process, and flavor characteristics of GRW. Results show that Process I significantly (p < 0.05) enriched the soluble solid and crude protein content of glutinous rice milk by improving gelatinization degree and pulping efficiency, which consequently enhanced the microbial growth, glycolysis, and protein decomposition during the GRW fermentation process. GC-MS analysis shows that Process I sequentially significantly (p < 0.05) enhanced the esterification and Ehrlich or Harrison pathway during the fermentation process. This contributed to a higher content of key ester and alcohol compounds. Supplementary Information: The online version contains supplementary material available at 10.1007/s10068-022-01119-7.

Microbial Communities and Flavor Compounds during the Fermentation of Traditional Hong Qu Glutinous Rice Wine.

As a traditional Chinese rice wine, Hong Qu glutinous rice wine (HQW) is popular among consumers due to its unique flavor. However, its quality changes during fermentation, and the potential relationships between flavor and microbes have not been systematically researched. In this work, physicochemical properties (pH, total sugar, alcohol, amino acid nitrogen), flavor compounds (organic acids, free amino acids, and volatile compounds), and microbial communities were investigated. The results revealed that Pantoea, Lactiplantibacillus, Lactobacillus, Leuconostoc, and Weissella predominated the bacterial genera, and Monascus was the predominant fungal genus. Organic acids, free amino acids, and key volatile compounds (esters and alcohols) significantly increased during fermentation. The correlations analysis showed that Lactiplantibacillus was closely associated with flavor compounds formation. This study deepens our understanding of the roles of microorganisms in flavor formation on traditional HQW fermentation.

Correlations between microbiota with physicochemical properties and volatile flavor components in black glutinous rice wine fermentation.

Black glutinous rice wine (BGRW) is a popular traditional Chinese rice wine; however, the flavors profiles associated with microbiota changes during its fermentation have not yet been evaluated. In this study, we explored the correlations between microbial communities with physicochemical properties and flavor components during BGRW fermentation. High-throughput sequencing was used to identify the microbial community composition of BGRW at different fermentation stages, and physicochemical properties and volatile flavor compounds (VFCs) were identified via fermentation features testing and headspace solid phase microextraction gas chromatography mass spectrometry. First, we revealed Pantoea and Kosakonia predominated bacterial genera the early stage of BGRW fermentation, Leuconostoc, Pediococcus, Bacillus, and Lactobacillus predominated bacterial genera the later stage, while Rhizopus and Saccharomyces were the predominant fungal genera throughout fermentation. Second, total sugars, titratable acids, pH, ethanol, amino acid nitrogen, and 43 VFCs were detected during fermentation. Twenty-three VFCs were differentially produced according to the linear discriminant analysis effect size method. With the increase of the fermentation time, the kinds and contents of esters and alcohols were also increased, while acids decreased. Moreover, 12 microbial genera, Lactococcus, Pediococcus, Leuconostoc, Lactobacillus, Cronobacter, Pantoea, Weissella, Enterococcus, Rhizopus, Myceliophthora, Cystofilobasidium, and Aspergillus were found to be highly correlated (|ρ| > 0.7 and P < 0.05) with physicochemical properties and VFCs, by redundancy analysis (RDA) and two-way orthogonal partial least squares (O2PLS) analysis. Ultimately, based on the results, a metabolic map of dominant genera in BGRW was established. Our findings provided detailed information on the dynamic changes of physicochemical properties and VFCs and selection of beneficial strains to improve the quality of BGRW.

Comparison of microbial communities and amino acid metabolites in different traditional fermentation starters used during the fermentation of Hong Qu glutinous rice wine.

Hong Qu glutinous rice wine (HQGRW) is produced from glutinous rice with the addition of the traditional fermentation starter Hong Qu (mainly Gutian Hong Qu and Wuyi Hong Qu) has been added. It is unpalatable and rejected by consumers because the bitter and umami tastes are too high. The objective of this study was to compare the dynamics of the microbial communities and amino acids especially those in the different traditional fermentation starters used during HQGRW fermentation, and elucidate the key microbes responsible for amino acids. Three widely-used types of Hong Qu starters were used which can make different bitterness and umami in our previous studies, namely, black Wuyi Hong Qu (WB), red Wuyi Hong Qu (WR), and Gutian Hong Qu (GT). The living dynamics of fungal and bacterial communities during the fermentation were determined by high-throughput sequencing and rRNA gene sequencing technology for the first time. The content of amino acids in the HQGRW were determined by reverse-phase high-performance liquid chromatography analysis. The results showed that there were differences between fungal communities during the fermentation process in Wuyi Hong Qu and Gutian Hong Qu starters and between bacterial communities during the fermentation process in the three types of starters. The amino acid content of the samples showed an increasing trend in each group. The total amino acids, as well as the bitter, sweet, umami, astringent amino acids, in the GT Hong Qu group increased more slowly during fermentation, as comparerd to those in WB and WR groups. Furthermore, Meyerozyma, Saccharomyces, Bacillus, Rhizopus, Pediococcus, Monascus, and Halomonas were strongly positively correlated with the content of bitter and umami amino acids (|r| > 0.6 with FDR adjusted P < 0.05) by Spearman's correlation analysis. To conclude, these findings may contribute to a better understanding of the bitter and umami amino acid production mechanism during traditional fermentation and helpful in improving the taste of HQGRW.

Major Metabolites and Microbial Community of Fermented Black Glutinous Rice Wine With Different Starters.

Black glutinous rice wine (BGRW) is a traditional Chinese rice wine that is brewed using multiple strains. However, the roles of these microorganisms, particularly their contributions to aroma formation, are poorly understood. Accordingly, the main goal of this study was to determine the microbial communities and major metabolites of different traditional fermentation starters. Anshun (AS) starter and Xingyi (XY) starter were used for BGRW to provide insight into their potential contributions to the variation in flavor and aroma. High-throughput sequencing of the microbial community using the Illumina MiSeq platform revealed significant differences during fermentation between the two starter groups. Pediococcus, Leuconostoc, and Bacillus were the dominant bacterial genera in the AS group, whereas Leuconostoc, Pediococcus, and Gluconobacter were the dominant genera in the XY group. In addition, Rhizopus, Saccharomyces, and Saccharomycopsis were the predominant fungal genera detected in both samples. The major metabolites in the two groups were identified by high-performance liquid chromatography and headspace-solid-phase microextraction gas chromatography-mass spectrometry. A total of seven organic acids along with 47 (AS) and 43 (XY) volatile metabolites were detected, among which lactic acid was the primary organic acid, and esters were the largest group in both types of wine. Principal components analysis further revealed significant differences in the dynamic succession of metabolites between the two samples. Correlation analysis showed that 22 and 17 microorganisms were strongly correlated with the production of major metabolites in AS and XY, respectively. Among them, Pediococcus, Leuconostoc, Lactobacillus, Lactococcus, and Streptococcus were shown to play crucial roles in metabolite synthesis. Overall, this study can provide a valuable resource for the further development and utilization of starters to improve the aromatic quality of BGRW.

Amino acid and microbial community dynamics during the fermentation of Hong Qu glutinous rice wine.

Hong Qu glutinous rice wine (HQGRW) is typically very bitter after fermentation due to the presence of bitter amino acids (BAA). The amino acids are considered to primarily derive from the hydrolysis of protein from the raw material by microbial populations during fermentation, and temperature also has an important effect on bitter tastes. Here, the dynamics of fungal and bacterial communities during the traditional fermentation of HQGRW were investigated using high-throughput sequencing and RNA-based rRNA gene sequencing. Both principal component analysis (PCA) and hierarchical clustering analysis (HCA) revealed significant differences between the fungal and bacterial communities during fermentation at 20 °C and those performed at 25 °C and 30 °C. The growth of Saccharomyces and some LAB apparently inhibited the growth of several pernicious bacterial taxa including acetic acid bacteria. The amino acid contents of the samples all increased continuously under the different temperature conditions. Moreover, higher temperatures were associated with higher perceptual intensity of bitterness and contents of amino acids including bitter, sweet, umami, and astringent type amino acids as well as the total amino acid content during fermentation. Furthermore, the total BAA content was strongly and positively correlated with Pediococcus, Saccharomyces, Lactobacillus, Monascus, and Halomonas relative abundances, with correlations identified by |r| > 0.6 with P adjusted P < 0.05. In conclusion, these results contribute to a better understanding of the mechanisms underlying BAA production during the traditional fermentation of HQGRW and will help improve the quality and safety of these wines.

Dynamic changes of total acid and bacterial communities during the traditional fermentation of Hong Qu glutinous rice wine

Background: Hong Qu glutinous rice wine (HQGRW) is brewed under non-aseptic fermentation conditions, so it usually has a relatively high total acid content. The aim of this study was to investigate the dynamics of the bacterial communities and total acid during the fermentation of HQGRW and elucidate the correlation between total acid and bacterial communities. Results: The results showed that the period of rapid acid increase during fermentation occurred at the early stage of fermentation. There was a negative response between total acid increase and the rate of increase in alcohol during the early fermentation stage. Bacterial community analysis using high-throughput sequencing technology was found that the dominant bacterial communities changed during the traditional fermentation of HQGRW. Both principal component analysis (PCA) and hierarchical clustering analysis revealed that there was a great difference between the bacterial communities of Hong Qu starter and those identified during the fermentation process. Furthermore, the key bacteria likely to be associated with total acid were identified by Spearman's correlation analysis. Lactobacillus, unclassified Lactobacillaceae, and Pediococcus were found, which can make significant contributions to the total acid development (| r| N 0.6 with FDR adjusted P b 0.05), establishing that these bacteria can associate closely with the total acid of rice wine. Conclusions: This was the first study to investigate the correlation between bacterial communities and total acid during the fermentation of HQGRW. These findings may be helpful in the development of a set of fermentation techniques for controlling total acid.

The dynamics of volatile compounds and their correlation with the microbial succession during the traditional solid-state fermentation of Gutian Hong Qu glutinous rice wine.

Gutian Hong Qu glutinous rice wine (GHQW) is a representative alcoholic beverage in China. During its traditional brewing process, multiple strains are involved. However, the roles of these microorganisms, particularly their contributions to aroma formation, are poorly understood. Therefore, in this study, the dynamics of volatile components and microbial succession during the traditional fermentation process of GHQW were monitored, by using HS-SPME-GC/MS and high throughput 16 S rRNA and ITS sequencing techniques. Following a 32-day semi-open solid-state fermentation, massive changes in volatile profile and microbial communities were observed. Based on these, the potential correlations between volatile components and microorganisms during the fermentation were explored by using Pearson correlation analysis. It was revealed that 3 bacterial taxa, including Lactococcus lactis, Burkholderia gladioli and Cronobacter helveticus, and 5 fungal taxa, including Wickerhamomyces anomalus, Saccharomyces cerevisiae, Aspergillus vitricola, Aspergillus penicillioides and Monascus purpureus, showed strong connections with various volatile components. Thus these bacteria and fungi might play crucial roles in volatile components biosynthesis, and thus were preliminary considered as core functional microorganisms for GHQW brewing. The detailed information on the dynamic changes of volatile components and microbial communities throughout GHQW brewing can further expand our understanding of the formation mechanisms of GHQW aroma.

Changes in Cold and Hot Syndrome and Gastrointestinal Bacterial Community Structure in Mice by Intervention with Food of Different Nature.

OBJECTIVE: To reveal the effect of foods with different natures on cold or hot syndrome and gastrointestinal bacterial community structure in mice. METHODS: Forty-five 6-week-old male ICR Kunming mice of clean grade were divided into 5 groups, 9 per group, including the control (CK), hot nature herbs (HM), Hong Qu glutinous rice wine (RW), tea rice wine (TW), and cold nature herbs (CM) groups. Distilled water or corresponding herbs were administered to mice (0.01 mL/g body weight) in the 5 groups by gastric infusion respectively, once daily for 28 d. Appearance, behavior, and serum biochemical indicators, including 5-hydroxytryptamine (5-HT), thyroid stimulating hormone (TSH), noradrenaline (NE), cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), the hot nature index, as well as the gastrointestinal bacterial community structure were analyzed in all groups after treatment. RESULTS: After supplementation for 28 d, CM and TW mice showed different degrees of cold syndrome, and HM and RW mice showed different degrees of hot syndrome. Compared with the HM and RW mice, the TSH, NE, cAMP levels and hot nature indices in the CM and TW mice were significantly decreased and 5-HT and cGMP levels were significantly increased (P<0.05). There was no obvious change in appearance or behavior in CK mice. Results of clustering analysis showed that the gastrointestinal bacterial community structures were highly similar in TW and CM mice as well as in RW and HM mice, and that they were from the same branch, respectively, when the distance was 0.02. The key microbes associated with cold syndrome were Lachnospiraceae uncultured, Lactococcus, etc., and the key microbes associated with hot syndrome were S24-7 norank, Ruminococcaceae uncultured, etc. CONCLUSION: The interventions with different nature foods could change cold or hot syndrome in mice, leading to changes in gastrointestinal bacterial community structure.

Changes in Cold and Hot Syndrome and Gastrointestinal Bacterial Community Structure in Mice by Intervention with Food of Different Nature / 中国结合医学杂志

OBJECTIVE@#To reveal the effect of foods with different natures on cold or hot syndrome and gastrointestinal bacterial community structure in mice.@*METHODS@#Forty-five 6-week-old male ICR Kunming mice of clean grade were divided into 5 groups, 9 per group, including the control (CK), hot nature herbs (HM), Hong Qu glutinous rice wine (RW), tea rice wine (TW), and cold nature herbs (CM) groups. Distilled water or corresponding herbs were administered to mice (0.01 mL/g body weight) in the 5 groups by gastric infusion respectively, once daily for 28 d. Appearance, behavior, and serum biochemical indicators, including 5-hydroxytryptamine (5-HT), thyroid stimulating hormone (TSH), noradrenaline (NE), cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), the hot nature index, as well as the gastrointestinal bacterial community structure were analyzed in all groups after treatment.@*RESULTS@#After supplementation for 28 d, CM and TW mice showed different degrees of cold syndrome, and HM and RW mice showed different degrees of hot syndrome. Compared with the HM and RW mice, the TSH, NE, cAMP levels and hot nature indices in the CM and TW mice were significantly decreased and 5-HT and cGMP levels were significantly increased (P<0.05). There was no obvious change in appearance or behavior in CK mice. Results of clustering analysis showed that the gastrointestinal bacterial community structures were highly similar in TW and CM mice as well as in RW and HM mice, and that they were from the same branch, respectively, when the distance was 0.02. The key microbes associated with cold syndrome were Lachnospiraceae uncultured, Lactococcus, etc., and the key microbes associated with hot syndrome were S24-7 norank, Ruminococcaceae uncultured, etc. CONCLUSION: The interventions with different nature foods could change cold or hot syndrome in mice, leading to changes in gastrointestinal bacterial community structure.

Microbial communities and volatile metabolites in different traditional fermentation starters used for Hong Qu glutinous rice wine.

Hong Qu glutinous rice wine (HQGRW), as one of the most typical representatives of Chinese rice wine, is generally brewed from glutinous rice by adding two traditional wine fermentation starters-Hong Qu (HQ) and Bai Qu (BQ). The objective of this study was to determine the microbial communities and volatile metabolites of different traditional fermentation starters for HQGRW, and elucidate the potential correlation between microbiota and volatile metabolites. Both heatmap and principal component analysis (PCA) revealed the significant variances in volatile profiles among different wine starters. Microbiological analysis based on high-throughput sequencing (HTS) technology demonstrated that both of bacterial and fungal communities varied significantly in different starters. HQ was dominated mainly by bacteria of Bacillus ginsengihumi (20.17%), Pantoea sp. (10.39%), Elizabethkingia sp. (5.52%), Streptococcus sp. (5.03%) Brevundimonas sp. (3.03%), Rickettsia prowazekii (2.94%), Thermus thermophilus (2.54%), Bacillus amyloliquefaciens (1.48%), Bacillus aryabhattai (1.42%); fungi of Monascus purpureus (39.7%), Aspergillus niger (27.35%), Xeromyces bisporus (8.39%), Aspergillus penicillioides (6.89%), Aspergillus flavus (2.33%) and Pichia farinose (0.79%). By contrast, BQ contained much higher proportions of bacteria of Lactococcus lactis (10.45%), Lactobacillus brevis (9.99%), Pediococcus pentosaceus (8.29%), Weissella paramesenteroides (6.69%), Lactobacillus fermentum (4.83%), Gluconobacter thailandicus (3.93%), Lactobacillus alimentarius (3.59%), fungi of Rhizopus arrhizus (31.47%), Saccharomycopsis fibuligera (27.86%), Aspergillus niger (20.81%), Issatchenkia orientalis (3.79%), Saccharomycopsis malanga (3.15%), Clavispora lusitaniae (2.29%), Candida tropicalis (1.47%), Saccharomyces cerevisiae (1.11%) and Rhizopus microsporus (0.57%). Furthermore, core functional microbiota that might contribute to volatile flavour development was explored through Spearman's correlation-based network analysis. Lactobacillus brevis, Lactobacillus alimentarius, Lactobacillus plantarum and Aspergillus niger were found to be strongly associated with acid compounds (FDR adjusted P < 0.01), while Pichia sp., Candida sp., Monascus purpureus, Lactobacillus brevis and Lactobacillus alimentarius were positively correlated with concentrations of aromatic esters associated with fruity and floral notes (FDR adjusted P < 0.01), implying that these microorganisms might make significant contributions to the flavour of rice wine. These findings demonstrated that the aromatic quality of HQGRW may be critically influenced by the microbiota in traditional fermentation starters. To conclude, this study would contribute to the development of novel defined starter cultures for improving the aromatic quality of HQGRW.

Exploring core functional microbiota responsible for the production of volatile flavour during the traditional brewing of Wuyi Hong Qu glutinous rice wine.

The objective of this study was to explore the core functional microbiota for the production of volatile flavour during the traditional brewing of Wuyi Hong Qu glutinous rice wine, one of the most typical representatives of rice wine in China. Microbiological analysis based on high-throughput sequencing (HTS) technology demonstrated that bacteria of Lactobacillus, Bacillus, Leuconostoc, Lactococcus, Raoultella, Staphylococcus, Pediococcus, and Weissella, and fungi of Saccharomyces, Saccharomycopsis, Rhizopus, Monascus, Pichia, Wickerhamomyces, Candida, and Aspergillus were the predominant genera during the traditional fermentation process. Principal component analysis (PCA) based on the relative abundance showed that both of bacterial and fungal communities varied significantly in different fermentation phases. Some predominant microbial species or genera (including bacteria of Bacillus spp., Staphylococcus spp., Weissella spp., and P. acidilactici, and fungi of M. purpureus, R. oryzae, R. arrhizus var. arrhizus, and A. niger) were detected at the initial brewing stage, and their populations decreased as the fermentation progressed, while those of Lactobacillus, Gluconacetobacter, Leuconostoc, Pichia, Wickerhamomyces, and Saccharomyces increased to become the predominant genera at the final stage. A total of 79 volatile compounds were identified in traditional fermentation starters and during the traditional brewing process, mainly including esters, alcohols, acids, aldehydes, ketones, and phenols. Heatmaps and PCA also revealed the significant variances in the composition of volatile compounds among different samples. Furthermore, the potential correlations between microbiota succession and volatile flavour dynamics were explored through bidirectional orthogonal partial least squares (O2PLS) based correlation analysis. Three bacterial genera, namely, Gluconacetobacter, Lactobacillus, Lactococcus, and three fungal genera of Pichia, Wickerhamomyces, and Saccharomyces, were determined as the core functional microbiota for production of main volatile compounds in Wuyi Hong Qu glutinous rice wine. To conclude, information provided by this study is valuable to the development of effective strategies for the selection of beneficial bacterial and fungal strains to improve the quality of Wuyi Hong Qu glutinous rice wine.

Comparison study of the volatile profiles and microbial communities of Wuyi Qu and Gutian Qu, two major types of traditional fermentation starters of Hong Qu glutinous rice wine.

Hong Qu, which mainly contains Monascus sp. and other microorganisms, as well as numerous microbial metabolites, is used as the fermentation starter of Hong Qu glutinous rice wine, a traditional alcoholic beverage. Two widely-used types of Hong Qu, namely Wuyi Qu (WYQ) and Gutian Qu (GTQ), were thoroughly compared for their fermentation properties, volatile profiles, and microbiota structures in this study. Significantly higher color value, glucoamylase and α-amylase activities were discovered in WYQ. And substantial variation in volatile components and microbial communities were also observed between them. It was identified that bacterial genus Burkholderia dominated GTQ (71.62%) and Bacillus dominated WYQ (44.73%), while Monascus purpureus was the most abundant fungal species in both types of starters (76.99%). In addition, 213 bacterial genera and 150 fungal species with low-abundance were also detected. Since the Linear Discriminant Analysis Effect Size algorithm, 14 genus-level bacterial taxa and 10 species-level fungal taxa could be utilized to distinguish these two types of starters. Moreover, the potential correlation of the volatile components and microbiota within WYQ and GTQ were further analyzed, by utilizing Partial Least Squares Discriminant Analysis. Ultimately, this study provides detailed insight into the volatile profiles and microbial communities presented in Hong Qu.

Evaluation of different PCR primers for denaturing gradient gel electrophoresis (DGGE) analysis of fungal community structure in traditional fermentation starters used for Hong Qu glutinous rice wine.

Denaturing gradient gel electrophoresis (DGGE) has become a widely used tool to examine microbial community structure. However, when DGGE is applied to evaluate the fungal community of traditional fermentation starters, the choice of hypervariable ribosomal RNA gene regions is still controversial. In the current study, several previously published fungal PCR primer sets were compared and evaluated using PCR-DGGE, with the purpose of screening a suitable primer set to study the fungal community of traditional fermentation starters for Hong Qu glutinous rice wine. Firstly, different primer sets were used to amplify different hypervariable regions from pure fungal cultures. Except NS1/FR1+ and ITS1fGC/ITS4, other primer sets (NL1+/LS2R, NL3A/NL4GC, FF390/FR1+, NS1/GCFung, NS3+/YM951r and ITS1fGC/ITS2r) amplified the target DNA sequences successfully. Secondly, the selected primer sets were further evaluated based on their resolution to distinguish different fungal cultures through DGGE fingerprints. Three primer sets (NL1+/LS2R, NS1/GCFung and ITS1fGC/ITS2r) were finally selected for investigating the fungal community structure of different traditional fermentation starters for Hong Qu glutinous rice wine. The internal transcribed spacer (ITS) region amplified by ITS1fGC/ITS2r, which is more hypervariable than the 18S rRNA gene and 26S rRNA gene, provides an excellent tool to separate amplification products of different fungal species. Results indicated that PCR-DGGE profile using ITS1fGC/ITS2r showed more abundant fungal species than that using NL1+/LS2R and NS1/GCFung. Therefore, ITS1fGC/ITS2r is the most suitable primer set for PCR-DGGE analysis of fungal community structure in traditional fermentation starters for Hong Qu glutinous rice wine. DGGE profiles based on ITS1fGC/ITS2r revealed the presence of twenty-four fungal species in traditional fermentation starter. A significant difference of fungal community can be observed directly from DGGE fingerprints and principal component analysis. The statistical analysis results based on the band intensities of fungal DGGE profile showed that Saccharomyces cerevisiae, Saccharomycopsis fibuligera, Rhizopus oryzae, Monascus purpureus and Aspergillus niger were the dominant fungal species. In conclusion, the comparison of several primer sets for fungal PCR-DGGE would be useful to enrich our knowledge of the fungal community structures associated with traditional fermentation starters, which may facilitate the development of better starter cultures for manufacturing Chinese Hong Qu glutinous rice wine.