Non-volatile Organic Acids Research Articles

In silico advancement of Aspergillus niger gluconic acid hyperproduction recommends plans for switching acid productivity

Background: Non-toxic, non-volatile organic acid, gluconic acid (pentahydroxycaproic acid) is found in abundance in honey and plants and in wines. Gluconic acid and its salts are used widely in food, feed, pharmaceuticals, textiles, and many other industries.Methods: In the current investigation, Aspergillus niger was utilized for Gluconic acid production in submerged fermentation. Furthermore, for Gluconic acid hyperproduction, the fungus Aspergillus niger was exposed to MNNG (100 µg/ml) for 15 minutes. Aspergillus niger strain (parent and mutant) was tested and identified for its capability of producing gluconic acid. We finished the molecular identification, in which we extracted the DNA from the A. niger and amplified its ITS region using specific primers. We further sequenced the amplified product and created a phylogenetic tree employing the MEGA6 software.Results: The tree results showed that our target sequence has 100% identity with Aspergillus niger, which proved that the isolated strain belongs to the Aspergillus genera. In addition, factors related to the production of gluconic acid were confirmed by cultivating the mutant strain A. niger (MG1) and the parent strain under several conditions. These included varying the pH (4-7), temperature (30-40°C), and the concentration of the carbon source (40-100 g/l). The outcomes displayed that the best pH was 5. The most favorable temperature was 30°C, and maximum production occurred at a carbon source concentration of 100 g/l, for both mutant and parent strains.Conclusion: This study shows how microbial strains and substrates could be used to attain cost-effective hyperproduction of gluconic acid to achieve objectives of industrial significance. Keywords: Gluconic Acid; Aspergillus niger; Hyper-production; Mutation; Molecular identification; Sequencing

A comparative metabolomics study of delayed-harvested and pumpkin grafted cucumbers

Cucumber is a widely consumed vegetable crop known for its rich nutrient composition and distinctive flavor, influenced by both volatile and non-volatile compounds. Grafting and delayed harvesting are crucial strategies for increasing cucumber yield. The present study investigates the impact of delayed harvesting at different developmental stages and grafting on the metabolic profile, flavor, and overall quality of cucumber fruits Yuxiu 2 (YX) using UPLC-MS/MS and GC-MS/MS techniques. The results indicate that delayed harvesting of YX led to significant increases in length, diameter, and weight from 12 to 24 days after pollination (DAP), with minimal growth beyond 24 DAP. However, grafting did not affect these physical parameters compared to self-rooted plants. Furthermore, metabolic profiling reveals that delayed harvesting enhances the concentration of certain non-volatile metabolites, including alkaloids, organic acids, and phenolic acids, while leading to a reduction in flavonoid contents. Overall, 140 non-volatile and 26 volatile differential metabolites were screened at three developmental stages. Notably, four new organic acids (6-amino hexanoic acid, 5-amino valeric acid, 1-hydroxy-2-naphthoic acid, and succinic semialdehyde) and three novel alkaloids (3-indole acetonitrile, epinephrine, and serotonin) were identified. Volatile compounds, such as aldehydes, esters, terpenes, alcohols, and ketones, exhibit a peak in concentration at 24 DAP, followed by a decline. The characteristic cucumber flavor compound, (E,Z)-2,6-nonadienal, remains consistent across all developmental stages. In grafted cucumber fruits, a total of 113 non-volatile and 11 volatile differential metabolites were screened, and among them, ten unique non-volatile metabolites were detected in grafted fruits, contributing to the sour and bitter taste of cucumbers. Moreover, some of the metabolites like (1S,4S,4aR)-1-isopropyl-4-methyl-7-methylene-1,2,3,4,4a,5,6,7-octahydronaphthalene with pentylenetetrazol contribute to an undesirable camphor-like odor. The study concludes that while delayed harvesting and grafting practices can increase cucumber yield, they also significantly alter the fruit’s metabolic profile, impacting taste and flavor.

Exploration of carbonyl compounds in red-fleshed kiwifruit wine and perceptual interactions among non-volatile organic acids

Carbonyl compounds are vital constituents that contribute to the flavor profile of alcoholic beverages. We examined 3-nitrophenylhydrazine as a derivatizing reagent for the measurement of 34 carbonyl compounds using UPLC−MS/MS. Adding formic acid and sodium acetate to the mobile phase significantly enhanced the detection limit of carbonyl compounds. The technique exhibited a notable extraction efficiency, yielding recovery percentages ranging from 83.6% to 117.1%, coupled with exceptional sensitivity, as evidenced by detection limits spanning from 0.07 μg/L to 4.80 μg/L. The relative standard deviation was <6.9%, indicating the precision and reliability of the analytical methodology. The method was verified by analyzing carbonyl compounds from red-fleshed kiwifruit wine. Furthermore, sensory assessment revealed that the amalgamation of tartaric acid, malic acid, and citric acid contributes to sour taste perception at sub-threshold concentrations through an additive interaction with supra-threshold non-volatile organic acids such as lactic acid and acetic acid.

The fecal microbiota of the mouse-eared bat (Myotis velifer) with new records of microbial taxa for bats.

Studies on the fecal microbiome of wild animals reveal valuable information on the feeding habits of the host and the possible roles of bacteria in digestion. In this work we characterized the fecal microbiota of seven male and seven female Myotis velifer bats using the V3-V4 regions of the 16S rRNA gene. Fecal samples were collected at the El Salitre cave in Mexico. We obtained 81 amplicon sequence variants, identifying four phyla, 12 families and 14 genera for females and seven phyla, 21 families and 26 genera for males. The phylum Synergistota is reported for the first time in bats. The most abundant phyla were Pseudomonadota and Fusobacteriota. Male feces showed a greater taxonomic richness than those from females. This study revealed that the fecal microbiota of M. velifer had a unique and more diverse composition compared to the microbiota reported for other bats. We identified 24 families and two abundant genera Cetobacterium and Haematospirillum in both males and females. Cetobacterium may produce vitamin B12 that is not produced by animals and Haematospirillum, which has been reported as an emerging human pathogen, may produce non-volatile organic acids. These genera had not been previously reported in the bat microbiota.

Mechanism of aroma enhancement methods in accelerating Congou black tea acidification subjected to room temperature storage

Acidification of aroma-enhanced black tea during storage was studied. UPLC-Q-TOF/MS (Ultra Performance Liquid Chromatography and Quadrupole-Time of Flight Mass Spectrometer) and HPLC (High-Performance Liquid Chromatography) analysis of non-volatile substances and organic acids revealed a decrease of soluble sugars and amino acids in aroma-enhanced black tea, while an increase in organic acids such as oxalic acid, malic acid and quinic acid. Further in vitro experiments indicated that the acidification of aroma-enhanced tea during storage can be attributed to decomposition of sugars and amino acids by heating, oxidation of aromatic aldehydes. Meanwhile, the amino acids, catechins, soluble sugars and flavonoids that constitute the taste of black tea are further reduced, changing the taste composition of tea infusion and further increasing its acidity. This study revealed the reasons for black tea acidification during aroma enhancement and storage and provided a theoretical basis for improving black tea quality.

Sensory attributes, chemical and microbiological properties of cigars aged with different media.

Introduction: Aging is an important process to improve the quality of cigar, but the effect of aging with media on cigar has not been reported.Therefore, this study aimed to prepare different aging media and clarify the influence of media on cigar quality. Methods: Effective media were first screened by sensory evaluation, then the effects of aging media on the chemical composition and microbial community of cigar were investigated. Results: The results showed that: 1) As aging media, coffee formula and cocoa formula could optimize the smoke and aroma characteristics of cigar, and 30days was the appropriate period for cigar aging. 2) Aging with coffee or cocoa media could increase the content of amino acids, non-volatile organic acids, malic acid and aroma components. Particularly, the content of aroma components increased from 2.48mg g-1 (W-30) to 3.21mg g-1 (C-30) and 3.70mg g-1 (K-30), respectively. 3) Aging with coffee can improve the diversity of bacteria and fungi on the cigar surface and change the succession rule of bacterial community. In contrast, aging with cocoa had no significant effect on microbial diversity of cigar. Discussion: In this study, the influence of aging media on cigar quality was analyzed multidimensionally for the first time, which provided a reference for the development of new aging media and technologies to improve cigar quality.

Visual screening of potassium solubilizing bacteria for increasing the solubilization of potassium minerals

Research on potassium solubilizing bacteria (KSB) has recently gained attention, most of it has been focused on the size of the halo zone; however, information regarding KSB growth duration and their potential to reduce the use of chemical fertilizers for sustainable agriculture and to be used as a biofertilizer are still lacking. The aim of this study was to offer a complete, long-term observation of KSB growth. We monitored the growth of two KSB strains that were isolated from the rhizosphere for 7 days on Modified Aleksandrov Medium (MAM) plates. We specifically designed the observation times. These methods were used to distinguish KSB’s capability to solubilize potassium minerals. The results indicated that KSB-12 isolate was capable of producing organic acid and thus solubilizing potassium faster than KSB-19 isolate at 1-4 days of incubation. Interestingly, the solubilizing zone of the KSB-12 isolate decrease after 5 days of incubation, while KSB-19 isolate continued to grow, maintain cell numbers, produce organic acids, and continuously release potassium. Different types of volatile and non-volatile organic acids have an effect on this phenomenon. In conclusion, we provided a well-designed time series study regarding the potassium solubilization efficiency of KSB, determining the fastest and longest strains of potential KSBs.

Effects of coffee and cocoa as fermentation additives on sensory quality and chemical compositions of cigar tobacco leaves

Exogenous additives applied for cigar tobacco leaves fermentation played an important role in sensory quality of cigar. In this study, effects of additives including coffee and cocoa on the quality of cigar tobacco leaves were explored. The changes of sensory quality, conventional chemical components, nonvolatile organic acids, and aroma compositions in tobacco leaves were studied. Results showed that the richness, mellowness and matureness of aroma, as well as the fluentness and sweetness of smoke in cigar were improved with the introduction of coffee or cocoa. According to chemical composition analysis, fermentation with coffee or cocoa increased the contents of alkaloids and sugar in tobacco leaves, which endowed the tobacco a relatively good performance on the sweetness and aroma richness. Besides, the decrease of citric acid, as well as the increase of saturated fatty acid and aroma components were found with additives, which was beneficial to improve the mellowness and aroma richness of cigar. It was suggested that coffee and cocoa exhibited positive effects on the chemical composition, especially increased the contents of sugar, saturated fatty acids and aromatic components, thus improving the sensory quality of tobacco leaves.

Preparation and Taste Profiling of the Enzymatic Protein Hydrolysate from a by-Product of Red Snow Crab Processing as a Natural Seasoning Compound.

The red snow crab (Chionoecetes japonicus) is the most industrially processed in the Republic of Korea, and the meat is very popular, owing to its savory taste and flavor. Its body meat production comprises a two-step separation to increase meat yield. However, during the secondary separation, broken shell debris is occasionally entrained in the meat products, which is a concern for manufacturers. As the residues from first separation contain 39.9% protein, it can be utilized as an enzymatic protein hydrolysate (FPH) rich in free amino acids (FAAs). A combination of flavourzyme and alcalase (1:1) superiorly hydrolyzed the protein of the residues, and the best hydrolysis condition was suggested at 60 °C for 15 h with fourfold water and 2% enzyme addition, achieving a 57.4% degree of hydrolysis. The EPH was mostly composed of FAAs containing most essential amino acids; however, bitter-tasting amino acids accounted for 46.4% of the FAAs. To reduce the bitter taste, different nonvolatile organic acids were considered as masking agents, and citric and malic acids were effective, though the umami taste is slightly decreased. In conclusion, the crab processing residues can be utilized as an FAA-based natural seasoning compound through enzymatic hydrolysis and organic acid treatment.

Study on the chemical compositions and microbial communities of cigar tobacco leaves fermented with exogenous additive

Fermentation process plays an important role in the biochemical properties and quality of cigar tobacco leaves (CTLs). In industry, exogenous additive (EA) was usually adopted for improving the quality of CTLs during fermentation. However, the mechanism of enhanced quality of CTLs fermented with EA was confused. Herein, the chemical compositions and microbial communities of CTLs during fermentation with EA were analyzed. The increased contents of total nitrogen and total sugar, as well as the improved consumption rate of reducing sugar in CTLs were found with the addition of EA. Besides, fermentation with EA reduced the content of total nonvolatile organic acid, especially unsaturated fatty acid. The contents of total and several representative aroma components were improved. Additionally, the increased abundance of Staphylococcus and decreased abundance of Aspergillus were detected. Combined with the changes of chemical compositions and microbial communities, it was confirmed that the carbohydrates and alcohols originated from EA promote the enrichment of Staphylococcus and accelerate biochemical reactions, such as Maillard reaction and esterification reaction, thus improving the contents and quality of aroma components in CTLs. This study demonstrated the mechanism of enhanced quality of CTLs fermented by EA, which provides more ideas for developing novel and efficient EAs.

Evaluation of the Perceptual Interaction among Ester Odorants and Nonvolatile Organic Acids in Baijiu by GC-MS, GC-O, Odor Threshold, and Sensory Analysis.

By applying the aroma extract dilution analysis, 13 esters were found to have high FD factors in the Laobaigan flavor type of Baijiu. These were then quantified using GC-MS. In addition, 35 nonvolatile organic acids were quantified by a derivatization method combined with GC-MS. The perceptual interactions of lactic acid and ethyl lactate and that of lactic acid and ethyl acetate were studied through the olfactory threshold. The S curve was used to evaluate the changes in the olfactory thresholds. After adding lactic acid (1142, 20 000, and 53 703 mg/L), the olfactory thresholds of ethyl lactate (1000, 724, and 295 mg/L) and ethyl acetate (398, 324, and 257 mg/L) decreased obviously, which revealed that lactic acid gave additive or synergistic odor effects for the two esters. Furthermore, it was discovered that as the concentrations of lactic acid increased, the extent of the interaction among these compounds was also greater in the mixture.

Metabolic profiles alteration of Southern Thailand traditional sweet pickled mango during the production process.

Sweet pickled mango named Ma-Muang Bao Chae-Im (MBC), a delicacy from the Southern part of Thailand, has a unique aroma and taste. The employed immersion processes (brining 1, brining 2, and immersion in a hypertonic sugar solution, sequentially) in the MBC production process bring changes to the unripe mango, which indicate the occurrence of metabolic profiles alteration during the production process. This occurrence was never been explored. Thus, this study investigated metabolic profile alteration during the MBC production process. The untargeted metabolomics profiling method was used to reveal the changes in volatile and non-volatile metabolites. Headspace solid-phase micro-extraction tandem with gas chromatography quadrupole time of flight (GC/QTOF) was employed for the volatile analysis, while metabolites derivatization for non-volatile analysis. In conclusion, a total of 82 volatile and 41 non-volatile metabolites were identified during the production process. Terpenes, terpenoids, several non-volatile organic acids, and sugars were the major mango metabolites that presented throughout the process. Gamma-aminobutyric acid (GABA) was only observed during the brining processes, which suggested the microorganism’s stress response mechanism to an acidic environment and high chloride ions in brine. Esters and alcohols were abundant during the last immersion process, which had an important role in MBC flavor characteristics. The knowledge of metabolites development during the MBC production process would be beneficial for product development and optimization.

Simultaneous quantification of volatile fatty acids and nonvolatile organic acids in Hevea brasiliensis latex.

The current method used in latex industries to determine the volatile fatty acids contents of Hevea brasiliensis latex is steam distillation. However, the accuracy of the method has been debated for some time. We assessed the accuracy of the method and developed a new, more reliable high-performance liquid chromatographic method of determining acids in latex. The volatile fatty acids (formic, acetic, propionic, butyric, and valeric acids) and nonvolatile organic acids (oxalic, malic, lactic, citric, and succinic acids) in latex are directly determined simultaneously for the first time with high sensitivity and without losses during sample preparation. To avoid errors from derivatization, an acid-resistant Prevail HPLC column and a gradient mobile phase of 25mM potassium dihydrogen phosphate (pH 2.5) and acetonitrile were employed. Under optimum conditions, the calibrations of both types of acids demonstrated satisfactory correlation coefficients of ≥0.990, with limits of detection ranging from 0.02 to 395mM. The developed method demonstrated the profiles of acids in field and concentrated latex of the same batch. Moreover, the evolution of the profiles of all studied acids in both types of latex during a 3-month period was also revealed.

Optimization and Validation of a Method for Analysis of Non-Volatile Organic Acids in Baijiu by Derivatization and its Application in Three Flavor-Types of Baijiu

Optimization and Validation of a Method for Analysis of Non-Volatile Organic Acids in Baijiu by Derivatization and its Application in Three Flavor-Types of Baijiu

Reconstitution of the Flavor Signature of Laobaigan-Type Baijiu Based on the Natural Concentrations of Its Odor-Active Compounds and Nonvolatile Organic Acids.

Nonvolatile organic acids (NVOAs) associated with the Laobaigan flavor type of Baijiu were analyzed by a derivatization method, and 37 NVOAs were quantified. In addition, 33 odorants were judged to have high flavor dilution factors by application of aroma extract dilution analysis and quantification by gas chromatography coupled with mass spectrometry (GC-MS) and GC combined with quadrupole time-of-flight MS. The quantitative data obtained for the odorants and NVOAs were used to recombine the overall flavor of Baijiu. The odor of the reconstitution of the odor-active volatiles and the NVOAs was more similar to that of the original Baijiu sample than the sample that only contained odor-active volatiles, and the alcoholic and sweety flavor odor characters were reduced, while others, that is, fruity, acidic, floral, jujube and grain, were amplified. It was shown for the first time that NVOAs have effects on the volatiles in Baijiu and might obviously influence the intensity of certain aroma qualities.

A flavoromics strategy for the differentiation of different types of Baijiu according to the non-volatile organic acids

Non-volatile organic acids (NVOAs) in 12 main flavor types of Baijiu were analyzed by a derivatization method combined with GC-MS and 38 NVOAs were quantified. Meanwhile, a flavoromics strategy based on the contents of NVOAs in the 12 flavor types of Baijiu was successfully used to the differentiation of Baijiu. PLS-DA models (explained variation, predictive capability) were used to consider different categories: fermentation process (0.931, 0.870), starter (0.921, 0.834), fermentation container (0.899, 0.810) and raw material (0.951, 0.909). Based on the selected categories, suitable separations were achieved, and the classification ability of these models were nearly 100%. As a result, the model demonstrated its ability to perfectly distinguish different types of Baijiu. Seventeen potential markers were identified by variable importance in projection method and were further processed using heatmap and hierarchical cluster analysis, indicating that the NVOAs had great discrimination power to differentiate Baijiu.

GC × GC-TOF/MS and UPLC-Q-TOF/MS based untargeted metabolomics coupled with physicochemical properties to reveal the characteristics of different type daqus for making soy sauce aroma and flavor type baijiu

Daqu is the starter for making baijiu (Chinese liquor). Daqu impart distinctive flavors to baijiu, but understanding of the metabolite composition and factors affecting fermentation performance of yellow, white and black daqu are limited. In this study, the relationship between the physicochemical properties of daqu and their metabolic profiles was determined. A total of 647 compounds were identified in daqu, including 401 volatile compounds, mainly aromatic compounds and pyrazines, and 246 non-volatile compounds which were mainly organic and amino acids. Correlation analysis showed that liquefying power, saccharifying power and pH were the main physicochemical properties accounted for the differences in metabolic profile between the three types of daqus. A group of 48 biomarkers (mainly alcohols, aromatics and non-volatile organic acids) selected by multivariate statistical analysis, could clearly distinguish between different types of daqu. Different metabolite phenotypes in three types of daqu were mainly caused by 21 metabolic pathways.

Pharmacometabolomics with a combination of PLS-DA and random forest algorithm analyses reveal meloxicam alters feline plasma metabolite profiles.

Repeated administration of meloxicam to cats is often limited by the potential damage to multiple organ systems. Identifying molecules that predict the adverse effects of meloxicam would help to monitor and individualize its administration, maximizing meloxicam's beneficial effects. The objectives of this study were to (a) determine if the repeated administration of meloxicam to cats alters the plasma metabolome and (b) identify plasma metabolites that may serve to monitor during the administration of meloxicam in cats. Purpose bred young adult cats (n=12) were treated with meloxicam at 0.3mg/kg or saline subcutaneously once daily for up to 17days. An untargeted metabolomics approach was applied to plasma samples collected prior to and at designated time points after meloxicam or saline administration. To refine the discovery of biomarkers, the machine-learning algorithms, partial least squares discriminant analysis (PLS-DA) and random forest (RF), were trained and validated using a separate unrelated group of meloxicam- and saline-treated cats (n=8). A total of 74 metabolites were included in the statistical analysis. Metabolomic analysis shows that the repeated administration of meloxicam alters multiple substances in plasma, including nonvolatile organic acids, aromatic amino acids, monosaccharides, and inorganic compounds as early as four days following administration of meloxicam. Seventeen plasma molecules were able to distinguish meloxicam-treated from saline-treated cats. The metabolomic changes discovered in this study may help to unveil unknown mechanisms of NSAID-induced side effects. In addition, some metabolites could be valuable for individualizing the administration of meloxicam to cats to mitigate adverse effects.

Effect of Rhizopus nigricans (Rhizopus stolonifera)-based novel starter culture on quality and safety attributes of doenjang, a traditional Korean soybean fermented food product

We aimed to develop a consortium of starter culture of effective microorganisms to prepare doenjang, a traditional Korean fermented food. Different ratios of Bacillus subtilis TKSP 24 (B), Aspergillus oryzae complex (A), Rhizopus nigricans (also named as Rhizopus stolonifera) (R), and Mucor racemosus 15 (M15) were selected as meju starter cultures to produce doenjang with improved quality. Microbial strain combinations (B: A: R and B: M15: R) were mixed separately at three different ratios [1:1:1 (w/w), 1:0.5:1.5 (w/w), and 1:1.5:0.5 (w/w)] to prepare BAR-1, BAR-2, BAR-3, BM15R-1, BM15R-2, and BM15R-3 doenjang samples. Quantitative analyses included free amino acids, free sugar, volatile and non-volatile organic acids, cellular antioxidant activity along with the presence of biogenic amines and aflatoxins, and microbial counts. Total free amino acids responsible for the sweet taste of doenjang were highest in BAR-2 (322.50 mg/100 g) and BM15R-3 (320.07 mg/100 g). Total volatile organic acid was highest in BAR-1 compared to other preparations. All doenjang samples had biogenic amines, especially histamine, below the toxicity level (500 mg/kg). Also, the aflatoxin and hazardous microbial count in the tested doenjang samples were below the level of toxicity. The findings suggest that use of multiple microbial strains in combination with R. nigricans as a starter culture could be a novel and effective approach to improve the nutrition and safety of fermented soybean food products of doenjang.

Effect of Weissella cibaria co-inoculation on the quality of Sichuan Pickle fermented by Lactobacillus plantarum

The effect of Weissella cibaria co-inoculation on the quality of Sichuan Pickle (SCP) fermented by Lactobacillus plantarum was evaluated. The fresh cowpea and seasoning ingredients were sterilized firstly by 70% (v/v) edible alcohol solution, and then respectively fermented with W. cibaria or L. plantarum or their mixed inoculums in brine containing 6.0% salt for 7 days at 25 °C. The cell growth, pH, non-volatile organic acids, volatile flavor compounds, residual nitrite and biogenic amines were monitored during fermentation. Compared to mono-inoculation of L. plantarum, co-inoculation accelerated cell proliferation, pH decrease and production of non-volatile organic acids in initial fermentation, and also significantly promoted the formation of volatile flavor compounds in types and contents during fermentation, especially for the aroma-active and key volatile compounds, thus improved the overall sensory quality of SCP. Furthermore, the co-inoculation decreased the concentration of nitrite and biogenic amines by 5–12% and 6–12%, respectively. Therefore, co-inoculating W. cibaria has improved the food quality of SCP fermented by mono-inoculation of L. plantarum.