Revealing the intrinsic relationship between microbial communities and physicochemical properties during ripening of Xuanwei ham.

To clarify the relationship between microorganisms and physicochemical indicators of Xuanwei ham. Six ham samples for the first, second and third year were selected, respectively. The changes of physicochemical properties, the free fatty acids and microbial communities of Xuanwei ham were investigated by GC-MS and high-throughput sequencing technology. Results showed that scores of colour, overall acceptability, texture, taste and aroma were the highest in the third year sample. With increasing ripening time, moisture content, water activity (Aw), lightness (L*), springiness, and resilience decreased continuously, and yellowness (b*) was the highest in the second year sample. 31 free fatty acids were detected, and unsaturated fatty acids such as palmitoleic acid, oleic acid, and linoleic acid were the major fatty acids. The content of palmitoleic acid, oleic acid and eicosenoic acid increased significantly during processing. At the phylum level, the dominant bacteria were Proteobacteria and Firmicutes, and fungi were Ascomycota. At the genus level, the dominant bacteria were Staphylococcus and Psychrobacter, and fungi were Aspergillus. Correlation analysis showed that water content and Aw were closely related to microorganisms, and most unsaturated fatty acids were significantly correlated with microorganisms. These findings showed that microorganisms played an important role in the quality of Xuanwei ham, and provided a scientific basis for the quality control of Xuanwei ham.

Effect of cooking methods on flavor profiles of Xuanwei Ham: Analytical insights into aromatic composition and sensory attributes.

To examine flavor variations in Xuanwei ham due to different cooking methods, we selected one-year cured Xuanwei ham and applied four techniques: dry frying (DF), baking (BA), steaming (ST), and boiling (BO). Organoleptic evaluation revealed ST received the highest overall sensory score. High-performance liquid chromatography (HPLC) revealed that the total nucleotide content was significantly different (P < 0.05), lactic acid predominated the measured organic acids. Solid-phase microextraction-gas chromatography-mass spectrometry (SPME-GC-MS) and chromatography-electronic nose (GC-E-Nose) data indicated that ST resulting in significantly higher total volatile compounds than the other methods (P < 0.05). SPME-GC-MS detected 55 volatile compounds, and 12 characteristic flavor substances were identified using orthogonal partial least squares discriminant analysis (OPLS-DA) (VIP > 1). This study aimed to support comprehensive research on the flavor characteristics of cooked Xuanwei ham and guide the selection of appropriate processing methods.

Separation, enrichment and cytoprotection of antioxidant peptides from Xuanwei ham using aqueous two-phase extraction.

An ethanol/(NH4)2SO4 biphasic (aqueous two-phase) system was designed to effectively separate antioxidant peptides from Xuanwei ham, and its potential to prevent ultraviolet A-induced damage to skin cells was explored. Optimization via single factor experiments and response surface methodology revealed that under 20 % ethanol aqueous solution (w/w), 25.5 % (NH4)2SO4 aqueous solution (w/w), and pH 8.80 conditions, the optimal extraction ratio was 59.0 ± 1.73 %. In vitro antioxidant activity and cellular assays showed that the peptide purified in the upper phase exhibited strong antioxidant activity, increasing the viability of HaCat cells damaged by UVA irradiation from 56.14 ± 1.05 % to 66.3 ± 1.76 %. We used an in silico peptide screening strategy and identified 10 with potential antioxidant activity, emphasizing the important role of amino acids Pro, Gly, and Ala in antioxidant activity.

Insight into the Relationship between the Causes of Off-Odour and Microorganism Communities in Xuanwei Ham.

To expound on the correlation between the microorganism communities and the formation of off-odour in Xuanwei ham, the microorganism communities and volatile compounds were investigated in the biceps femoris (BF) and semimembranosus (SM) of Xuanwei ham with different quality grades (normal ham and spoiled ham). The single molecule real-time sequencing showed that differential bacteria and fungi were more varied in normal hams than in spoiled hams. Headspace solid-phase microextraction-gas chromatography (HS-SPME-GC-MS) results indicated that aldehydes and alcohols were significantly higher in spoiled hams than those in normal hams (p < 0.05). The off-odour of spoiled hams was dominated by ichthyic, malodourous, sweaty, putrid, sour, and unpleasant odours produced by compounds such as trimethylamine (SM: 13.05 µg/kg), hexanal (BF: 206.46 µg/kg), octanal (BF: 59.52 µg/kg), methanethiol (SM: 12.85 µg/kg), and valeric acid (BF: 15.08 µg/kg), which are positively correlated with Bacillus cereus, Bacillus subtilis, Bacillus licheniformis, Pseudomonas sp., Aspergillus ruber, and Moraxella osloensis. Furthermore, the physicochemical property and quality characteristics results showed that high moisture (BF: 56.32 g/100 g), pH (BF: 6.63), thiobarbituric acid reactive substances (TBARS) (SM: 1.98 MDA/kg), and low NaCl content (SM: 6.31%) were also responsible for the spoilage of hams with off-odour. This study provided a deep insight into the off-odour of Xuanwei ham from the perspective of microorganism communities and a theoretical basis for improving the flavour and overall quality of Xuanwei hams.

Identification, flavor characteristics and molecular docking of umami taste peptides of Xuanwei ham.

To better understand the palatable properties of Xuanwei ham, the aqueous extract was isolated, analyzed and combin with sensory evaluation. Of umami-tasting activity and umami-enhancing impact, four new peptides (MDAIKKMQ, RKYEEVAR, YVGDEAQSKRG, and VNVDEVGGEALGR) were extracted and identified by ultrafiltration, gel separation, reverse performance liquid chromatography, and nano-LC-MS / MS. Sensory evaluation results showed that all of them had umami activity and enhanced umami taste, among which VNVDEVGGEALGR had the best effect. These peptides' umami taste thresholds ranged from 0.25 to 0.8 mg/mL. The MSG solution's umami taste threshold ranged from 0.125 to 0.5 mg/mL. Molecular docking results showed that the four umami peptides could be embedded into the binding pocket of the T1R3 cavity of the umami taste receptor T1R1/T1R3, wherein the binding sites Asp219, Asp150, and Thr179 may play crucial roles, and Glu222, Asp108, Glu217 and Glu148 play auxiliary roles. Hydrogen bonding and hydrophobic interactions were the most prominent interaction forces. This study helps to clarify the flavor characteristics of Xuanwei ham and could improve new processing technology.

The modulatory effects on enterohepatic cholesterol metabolism of novel cholesterol-lowering peptides from gastrointestinal digestion of Xuanwei ham.

The aim of this study was to investigate the effects and mechanism of in vitro protein digestive products of Xuanwei ham with different ripening periods on cholesterol metabolism and hypercholesterolemia. The results showed that compared with other gastrointestinal digestion (GID) groups, the GID group of Xuanwei ham with 3-year ripening period (XWH3-GID) inhibited the expression of Niemann-Pick C1-like 1 (NPC1L1) and acetyl-CoA acetyltransferase 2 (ACAT2) through hepatocyte nuclear factor 1-alpha (HNF-1α), which in turn effectively inhibited cholesterol absorption in Caco-2 cell monolayers. Following absorption by Caco-2 cell monolayers, the XWH3-GID group suppressed the expression and secretion of proprotein convertase subtilisin/kexin type 9 (PCSK9) via HNF-1α, which enhanced the protein expression and fluorescence intensity of low density lipoprotein receptor (LDLR) on the HepG2 cell membrane, and thus promoted the uptake of low density lipoprotein (LDL). Importantly, three novel peptides (LFP, PKF and VPFP) derived from titin were identified after intestinal epithelial transport in the XWH3-GID group, which could exert cholesterol-lowering effects through inhibiting intestinal cholesterol absorption and promoting peripheral hepatic LDL uptake, and effectively ameliorate western diet-induced hypercholesterolemia in ApoE-/- mice. These results suggest that Xuanwei ham with 3-year ripening period can be used as a source of cholesterol-lowering peptides and has potential to intervene in hypercholesterolemia.

Effect of Xuanwei Ham Proteins with Different Ripening Periods on Lipid Metabolism, Oxidative Stress, and Gut Microbiota in Mice.

SCOPE: Consumption of dry-cured ham may have a cardiovascular disease preventive capacity. However, whether ham protein in the maintenance of health and prevention of disease is dependent on ripening periods need to be clarified. METHODS AND RESULTS: Fifty C57BL/6 mice consume a diet that contains either Casein, raw ham protein (XWH0), or Xuanwei ham protein after 1-, 2-, or 3-years of maturation (XWH1, XWH2, or XWH3) for 4 weeks. Results show that longer ripening periods in the Xuanwei ham groups are negatively correlated with body weight gain, adipose tissue weight, total cholesterol (TC), and triglycerides levels in serum and liver, and these proteins intake could maintain liver health, reduce lipid deposition and oxidative stress, regulate lipid metabolism-related genes expression, alter the gut microbiota, and promote short-chain fatty acid production. Remarkably, XWH3 intake maintains lower TC and low-density lipoprotein cholesterol levels in serum and has high antioxidative activity as well. This regulation of lipid metabolism and oxidative stress may be related to the AMPK/Nrf2 signaling. CONCLUSION: Xuanwei ham dietary proteins that have endured a specific ripening period may have preventive effects on metabolic syndrome, which provide a basis for new ideas for nutritional health interventions.

Evaluation of small molecular metabolites and sensory properties of Xuanwei ham salted with partial replacement of NaCl by KCl.

The study was to understand the effect of the partial substitution of NaCl by KCl on small molecular metabolites and sensory quality of Xuanwei ham. Thirty green hams were randomly divided into five treatments, and salted with 100% NaCl (I), 70% NaCl+30% KCl (II), 60% NaCl+40% KCl (III), 50% NaCl+50% KCl (IV) and 40% NaCl+60% KCl (V), respectively. With the increase of KCl substitution, the moisture content of Xuanwei ham increased. Non-targeted ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-Q-Exactive-MS) was used to study the effect of partial substitution of NaCl by KCl, and twenty-eight metabolites were identified as markers of small molecular metabolites in the different treatments. KCl substitution promoted the release of tryptophan, histidine, citrulline, lysine, creatine and oleic acid, which contributed to improve the flavor and taste of ham. Therefore, the treatment II and III could reduce the NaCl content of Xuanwei ham by 30% and 40%, and maintained a better sensory acceptability.

Isolation and Antibiotic Resistant Research of Tetragenococcus halophilus from Xuanwei Ham, A China High-Salt-Fermented Meat Products.

We assessed the prevalence of antibiotic resistant and antibiotic resistance genes for 49 Tetragenococcus halophilus (T. halophilus) strains isolated from Xuawei ham in China. The antibiotic resistance phenotype was detected by the Bauer-Kirby (K-B) method and the results showed that 49 isolates can be considered completely susceptible to penicillin, ampicillin, amoxicillin, cefradine, cefotaxime, tetracyclines, minocycline, doxycycline, and vancomycin, but resistant to gentamicin, streptomycin, neomycin, polymyxinB, cotrimoxazole. This resistance was sufficiently high to consider the potential for acquisition of transmissible determinants. A total of 32 isolates were resistant to ofloxacin, 4 isolates were resistant to ciprofloxacin and chloramphenicol, and 2 isolates were resistant to ceftazidime and ticarcillin. The antibiotic resistance genes were detected by routine polymerase chain reaction (PCR). Among the 26 antibiotic resistance genes, 5 varieties of antibiotic resistance genes, including acrB, blaTEM, AAda1, SulII, and GyrB were detected and the detection rates were 89.79%, 47.7%, 16.33%, 77.55%, and 75.51%, respectively. The potential acquisition of transmissible determinants for antibiotic resistance and antibiotic resistance genes identified in this study necessitate the need for a thorough antibiotic resistance safety assessment of T. halophilus before it can be considered for use in food fermentation processes.

The proteomics homology of antioxidant peptides extracted from dry-cured Xuanwei and Jinhua ham.

The objective of this study was to investigate the antioxidant activity and the homology of peptides extracted from dry-cured Xuanwei and Jinhua ham. The antioxidant activity of crude peptides was assessed by ORAC and ABTS assays and the scavenging effects on DPPH and O2- free radicals. LC-ESI-Q-TOF-MS/MS was used to analyze the peptide composition. Based on the identified peptides, homologous proteins were matched by the Peaks software. Overall, 243 and 213 peptides were identified with their parent proteins in Xuanwei and Jinhua dry-cured hams. Based on the ORAC and TEAC values, XHP showed higher antioxidant ability than JHP (P < 0.05). After further purification by G-15 chromatography, the results indicate that the oligopeptides with less than 1000 Da had greater antioxidant activity than the other two fractions. Homology-based proteomics showed that the majority of peptides originated from myosin which accounted for 26% in XHP and 32% in JHP respectively.

Antioxidant and angiotensin I-converting enzyme inhibitory activities of Xuanwei ham before and after cooking and in vitro simulated gastrointestinal digestion.

Xuanwei ham is especially rich in a large amount of peptides and free amino acids under the action of protein degradation. Some of these peptides can potentially exert bioactivities of interest for human health. Traditionally, Xuanwei ham should undergo Chinese household cooking treatments before eating. However, it has not been known how its bioactivity changes after cooking and gastrointestinal digestion. Herein, Xuanwei ham is analysed before and after cooking, as well as gastrointestinal digestion being simulated so as to evaluate and compare its effect on antioxidant and angiotensin I-converting enzyme (ACE) inhibitory activities. The antioxidant activity is analysed using five different methods, and results demonstrate that cooking has some negative effects on antioxidative capacity when determined using different antioxidant methods except for a significant increment in 1,1'-diphenyl-2-picrylhydrazyl radical-scavenging activity, while ACE inhibitory activity increases significantly after cooking compared with control samples. After gastrointestinal digestion of samples, there is a significant increment of the antioxidant and ACE inhibitory activities in comparison with control and cooked samples. Particularly, after gastrointestinal digestion, free thiols content and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) radical-cation-scavenging activity of Xuanwei ham, respectively, increase about twice and fourfold, while ACE inhibitory activity increases about twice compared to cooked samples, reaching the value of 83.73%. Therefore, through cooking the antioxidant activity and ACE inhibitory activity of Xuanwei ham are not completely lost and a part of them is still maintained, while gastrointestinal digestion produces a significant enhancement in both bioactivities, highlighting a greater potential for a beneficial physiological effect on human health after eating it.

Purification and identification of antioxidative peptides from dry-cured Xuanwei ham.

This study mainly focused on the purification and identification of antioxidative peptides generated in dry-cured Xuanwei ham. Based on scavenging effect on free radicals and ferrous ion, the antioxidant activity of crude peptides from Xuanwei ham was assessed. From the scavenging effects on 1,1-diphenyl-2-picrylhydrazyl (DPPH), hydroxyl (OH) radicals and superoxide anion (O2(-)), it was suggested that XHP generated during the ripening period had a strong antioxidant activity. By using size exclusion chromatography, anion exchange column and reversed-phase HPLC, fractions with a strong antioxidative activity were separated based on their molecular weight and polarity differences. The fraction with strong antioxidant effect was further characterized by LC-MS/MS. The results suggest that antioxidative peptides are produced during the long processing of Xuanwei ham among which the tetrapeptide Asp-Leu-Glu-Glu could be one of the main peptides that play key role in the antioxidant activity.