Structural Elucidation of Sialylated O-Glycan Alditols Obtained from Mucins by Mass Spectrometry.

Acidic O-glycans having sialic acid and/or sulfate residue are abundantly expressed in intestinal mucins. However, structural elucidation of acidic O-glycans is a laborious and time-consuming task due to their large structural variations. Here, we describe a methodology of structural elucidation for sialylated O-glycan alditols from intestinal mucins using tandem mass spectroscopy. Methylesterification and mild periodate oxidation of sialylated O-glycan alditols assist mass analysis. This description includes the purification process of O-glycan alditols for structural analysis.

Determination of sugars and sugar alcohols in infant formula by high performance liquid chromatography with evaporative light-scattering detector.

A method was established for the simultaneous determination of five sugars (fructose, glucose, sucrose, lactose, maltose) and five sugar alcohols (erythritol, xylitol, sorbitol, mannitol, maltitol) in infant formula by high performance liquid chromatography-evaporative light scattering detector. After the samples were extracted with acetonitrile-water solution, precipitated by acetic acid, and purified with solid phase extraction cartridge, ALLChrom Rocksil Carbohydrate ES column was adopted for separation, and isocratic elution was conducted at the flow rate of 1.0 mL/min with acetonitrile-0.04 % ammonia solution as the mobile phase. The analytes were detected by an evaporative light-scattering detector, and quantified by external standard method. The linear ranges of the 10 components were 0.04-4.0 g/L with the correlation coefficients greater than 0.999, and the limits of quantification (S/N = 10) of the method were 0.08-0.4 g/100 g. The relative standard deviation of the lactose parallel samples reached 1.29 %, and the recoveries of the other 9 components ranged from 80.4 % to 99.4 % with the relative standard deviation of 2.8 %-7.1 %. The method performs well in sensitivity and separation, which is suitable for the simultaneous quantitative determination of sugars and sugar alcohols in infant formula.

Ongoings in the apple watercore: First evidence from proteomic and metabolomic analysis.

The presence of watercored fruit with translucent mesocarp has attracted immense attention due to its unique morphology and taste, however, the metabolic reconstruction between watercored and non-watercored tissues remain elusive. Herein, the combined proteomic and metabolomic approach was carried out to characterize the protein abundance and metabolic profile in watercored apple. Results demonstrated that carbohydrate metabolism was prioritized enriched in watercored apple, including highly accumulated sucrose and sugar alcohols, and the increased level of transcription factor WHIRLY2 (WHY2), which is required in sugar reallocation. More than 50% overlap of differently abundant proteins involved in calcium ion binding, starch and sucrose metabolism implied the involvement of calcium signaling in watercore development. Moreover, significantly lower calcium content was detected in watercored apples. Sucrose synthase (SUSY) protein, which is involved both in calcium ion binding and sugar metabolism, was the potential basis of apple watercore development, which provided a likely candidate in regulation of apparent quality.

Development of an Integrated Platform to Assess the Physicochemical and Toxicological Properties of Wood Combustion Particulate Matter.

Wood burning contributes to indoor and ambient particulate matter (PM) pollution and has been associated with increased morbidity and mortality. Here, we present an integrated methodology that allows to generate, sample, and characterize wood smoke derived from different moisture contents and representative combustion conditions using pine wood as a model. Flaming, smoldering, and incomplete combustion were assessed for low-moisture pine, whereas both low-moisture pine and high-moisture pine were investigated under flaming conditions. Real-time monitoring of carbon monoxide, volatile organic compounds, and aerosol number concentration/size in wood smoke was performed. The PM was size-fractionated, sampled, and characterized for elemental/organic carbon, organic functional groups, and inorganic elements. Bioactivity of PM was assessed by measuring the sterile alpha motif (SAM) pointed domain containing ETS (E-twenty-six) transcription factor (SPDEF) gene promoter activity in human embryonic kidney 293 (HEK-293T) cells, a biomarker for mucin gene expression. Findings showed that moisture content and combustion condition significantly affected the organic and inorganic elemental composition of PM0.1 as well as its bioactivity. Also, for a given moisture and combustion scenario, PM chemistry and bioactivity differed considerably with PM size. Importantly, PM0.1 from flaming combustion of low-moisture pine contained the highest abundance of the oxygenated saturated aliphatic functional group [H-C-O] and was also biologically most potent in stimulating SPDEF promoter activity, suggesting the role of organic compounds such as carbohydrates and sugar alcohols (that contain [H-C-O]) in driving mucus-related respiratory outcomes. Our platform enables further well-controlled parametric studies using a combination of in vitro and in vivo approaches to link wood burning parameters with acute and chronic inhalation health effects of wood smoke.

Metabolomics-derived marker metabolites to characterize Phaeocystis pouchetii physiology in natural plankton communities.

Phaeocystis pouchetii (Hariot) Lagerheim, 1893 regularly dominates phytoplankton blooms in higher latitudes spanning from the English Channel to the Arctic. Through zooplankton grazing and microbial activity, it is considered to be a key resource for the entire marine food web, but the actual relevance of biomass transfer to higher trophic levels is still under discussion. Cell physiology and algal nutritional state are suggested to be major factors controlling the observed variability in zooplankton grazing. However, no data have so far yielded insights into the metabolic state of Phaeocystis populations that would allow testing this hypothesis. Therefore, endometabolic markers of different growth phases were determined in laboratory batch cultures using comparative metabolomics and quantified in different phytoplankton blooms in the field. Metabolites, produced during exponential, early and late stationary growth of P. pouchetii, were profiled using gas chromatography-mass spectrometry. Then, metabolites were characterized that correlate with the growth phases using multivariate statistical analysis. Free amino acids characterized the exponential growth, whereas the early stationary phase was correlated with sugar alcohols, mono- and disaccharides. In the late stationary phase, free fatty acids, sterols and terpenes increased. These marker metabolites were then traced in Phaeocystis blooms during a cruise in the Barents Sea and North Norwegian fjords. About 50 endometabolites of P. pouchetii were detected in natural phytoplankton communities. Mannitol, scyllo-inositol, 24-methylcholesta-5,22-dien-3ß-ol, and several free fatty acids were characteristic for Phaeocystis-dominated blooms but showed variability between them. Distinct metabolic profiles were detected in the nutrient-depleted community in the inner Porsangerfjord (< 0.5 µM NO3-, < 0.1 µM PO 4 3- ), with high relative amounts of free mono- and disaccharides indicative for a limited culture. This study thereby shows how the variable physiology of phytoplankton can alter the metabolic landscape of entire plankton communities.

Comprehensive liamocin biosurfactants analysis by reversed phase liquid chromatography coupled to mass spectrometric and charged-aerosol detection.

Liamocin biosurfactants and structurally related exophilins secreted by the Aureobasidium pullulans (A. pullulans) strain NRRL62031 were firstly analyzed by hyphenation of high-performance liquid chromatography (HPLC) with high-resolution mass spectrometry (HRMS). Ten different analytes were detected and identified by their accurate masses and divided into subclasses according to their different head groups: three liamocins with arabitol as head group, three mannitol liamocins, and four exophilins. A baseline separation of congeners within the subclasses was achieved by reversed phase HPLC on a C18 stationary phase, whereas an overlap of subclasses occurred. The structures were simultaneously confirmed by online tandem mass spectrometry (MS/MS) experiments in positive and negative ionization mode. The assigned polyol head groups and thus the feasibility of this method were confirmed by gas chromatography (GC)-MS data obtained after hydrolysis and derivatization of the liamocins. Based on the varying structural characteristics of liamocins, e.g. the polyol head group (or even none for exophilins) and the degree of acetylation, different detector response in LC-MS was expected, impairing relative quantification of congeners. Therefore, a complementary quantification method was developed using HPLC coupled to charged-aerosol detection (CAD), which allows the determination of the amount of the individual liamocin species without authentic liamocin standards. Hence, the here presented hyphenated techniques facilitate comprehensive analysis of liamocin biosurfactants.

Impact of maturation and growth temperature on cell-size distribution, heat-resistance, compatible solute composition and transcription profiles of Penicillium roqueforti conidia.

Penicillium roqueforti is a major cause of fungal food spoilage. Its conidia are the main dispersal structures of this fungus and therefore the main cause of food contamination. These stress resistant asexual spores can be killed by preservation methods such as heat treatment. Here, the effects of cultivation time and temperature on thermal resistance of P. roqueforti conidia were studied. To this end, cultures were grown for 3, 5, 7 and 10 days at 25 °C or for 7 days at 15, 25 and 30 °C. Conidia of 3- and 10-day-old cultures that had been grown at 25 °C had D56-values of 1.99 ± 0.15 min and 5.31 ± 1.04 min, respectively. The effect of cultivation temperature was most pronounced between P. roqueforti conidia cultured for 7 days at 15 °C and 30 °C, where D56-values of 1.12 ± 0.05 min and 4.19 ± 0.11 min were found, respectively. Notably, D56-values were not higher when increasing both cultivation time and temperature by growing for 10 days at 30 °C. A correlation was found between heat resistance of conidia and levels of trehalose and arabitol, while this was not found for glycerol, mannitol and erythritol. RNA-sequencing showed that the expression profiles of conidia of 3- to 10-day-old cultures that had been grown at 25 °C were distinct from conidia that had been formed at 15 °C and 30 °C for 7 days. Only 33 genes were upregulated at both prolonged incubation time and increased growth temperature. Their encoded proteins as well as trehalose and arabitol may form the core of heat resistance of P. roqueforti conidia.

Maltitol: Analytical Determination Methods, Applications in the Food Industry, Metabolism and Health Impacts.

Bulk sweetener maltitol belongs to the polyols family and there have been several dietary applications in the past few years, during which the food industry has used it in many food products: bakery and dairy products, chocolate, sweets. This review paper addresses and discusses in detail the most relevant aspects concerning the analytical methods employed to determine maltitol's food safety and industry applications, its metabolism and its impacts on human health. According to our main research outcome, we can assume that maltitol at lower doses poses little risk to humans and is a good alternative to using sucrose. However, it causes diarrhoea and foetus complications at high doses. Regarding its determination, high-performance liquid chromatography proved the primary method in various food matrices. The future role of maltitol in the food industry is likely to become more relevant as processors seek alternative sweeteners in product formulation without compromising health.

[Sugar and sugar alcohols in infants Ying Yang Bao based on derivatization and gas chromatography-mass spectrometry].

OBJECTIVE: To establish a gas chromatography-mass spectrometry(GC-MS) method for determination of seven sugars and sugar alcohols in infants Ying Yang Bao nutritional supplements. METHODS: The samples were extracted with pure water and diluted with 95% ethanol. After being dried by nitrogen, methoxyamine hydrochloride oxime was dissolved in pyridine and derivatized by MSTFA. The capillary column TG-5 Ms(30 m×0. 25 mm, 0. 25 µm) was used for determination by GC-MS. RESULTS: The limits of detection(LODs)were 1. 0-3. 0 mg/g and the limits of quantification(LOQs)were 3. 3-10. 0 mg/g. The average recoveries of seven kinds of sugar and sugar alcohols were 86. 7%-96. 7%, and the relative standard deviation was less than 5. 1%(n=6). The contents of seven sugars and sugar alcohols in soybean matrix nutritional supplements were determined in the range of 0. 25-13. 70 g/100 g, which was consistent with the nutrition label of the products. CONCLUSION: The method is convenient, mild and fit for batch sample analysis.

How Do Trichoderma Genus Fungi Win a Nutritional Competition Battle against Soft Fruit Pathogens? A Report on Niche Overlap Nutritional Potentiates.

We present a case study report into nutritional competition between Trichoderma spp. isolated from wild raspberries and fungal phytopathogenic isolates (Colletotrichum sp., Botrytis sp., Verticillium sp. and Phytophthora sp.), which infect soft fruit ecological plantations. The competition was evaluated on the basis of nutritional potentiates. Namely, these were consumption and growth, calculated on the basis of substrate utilization located on Biolog® Filamentous Fungi (FF) plates. The niche size, total niche overlap and Trichoderma spp. competitiveness indices along with the occurrence of a stressful metabolic situation towards substrates highlighted the unfolding step-by-step approach. Therefore, the Trichoderma spp. and pathogen niche characteristics were provided. As a result, the substrates in the presence of which Trichoderma spp. nutritionally outcompete pathogens were denoted. These were adonitol, D-arabitol, i-erythritol, glycerol, D-mannitol and D-sorbitol. These substrates may serve as additives in biopreparations of Trichoderma spp. dedicated to plantations contaminated by phytopathogens of the genera Colletotrichum sp., Botrytis sp., Verticillium sp. and Phytophthora sp.

[Survey of Label Information of Foods with Function Claims].

The consumption of health food products, such as Foods with Function Claims, has grown in Japan. Significant information, such as possible side effects or drug interactions, are expected to be described on the packaging to help consumers to make an informed choice about products. In this study, we checked the items described on the packaging of Foods with Function Claims containing eicosapentaenoic acid (EPA) and/or docosahexaenoic acid (DHA), Salacinol/Fagomine/Neokotalanol, or Varyl-Tyrosine/Lactotripeptide. We found that the label information on the package have issues that need to be addressed; for example, the description about a warning for concomitant use with antithrombotic drugs was found in only 29.7% of EPA and/or DHA containing products (44 out of 148). Providing information for safe usage of products to consumers is pivotal. Therefore, improving product labeling, and further pharmaceutical support in case of taking health foods, should be considered.

Differential Metabolic Responses Caused by the Most Important Insect Pest of Coffee Worldwide, the Coffee Berry Borer (Hypothenemus hampei).

The world's coffee supply is threatened by the coffee berry borer, Hypothenemus hampei, the most destructive pest affecting coffee production and quality. This study hypothesized that coffee berry borer infestation induces distinct metabolic responses in the green coffee seeds of Coffea arabica and Coffea canephora (robusta). A targeted metabolomics approach was conducted using liquid chromatography tandem mass spectrometry to quantify intracellular metabolites in infested and uninfested arabica and robusta green seeds. In parallel, the seed biomass content and composition were assessed for the same conditions. Coffee berry borer attack induced increases in the levels of chlorogenic acids in arabica seeds, whereas organic acids and sugar alcohols were more abundant in infested robusta seeds. Most importantly, a set of compounds was identified as biomarkers differentiating the metabolic response of these taxa to the coffee berry borer.

Two aspects of honeydew honey authenticity: Application of advance analytical methods and chemometrics.

Taking into account a growing market and small number of articles related to honeydew honey, a metabolomic approach associated with multivariate analysis and modelling was proposed to discriminate five varieties of honey. Advanced analytical techniques were used for determination of 20 elements, 14 carbohydrates and stable carbon isotope ratio. No chemical marker has been found within sugar compounds, but several elements (Ba, Ca, Mg, Sr, Mn, Al, Co, Ni, Se) were marked as characteristic of honey type and allow classification of three botanical origins (Abies alba, Quercus frainetto, Quercus ilex). Sugars turanose, trehalose, arabinose and raffinose, elements Ba, Sr, P, Cd and Se, and δ13C values of honey, have different concentrations in honeys of the same botanical origin but harvested in different season. In addition to a confirmation of authenticity in terms of production, the values of δ13C of protein could be a good indicator of botanical origin.

The use of of inulin, maltitol and lecithin as fat replacers and plasticizers in a model reduced-fat mozzarella cheese-like product.

BACKGROUND: Mono-, di- and oligosaccharides, polyhydric alcohols and lipids are three main types of plasticizers used to process food materials. In the present study, inulin, maltitol and lecithin were selected as representative oligosaccharide, polyhydric alcohol and lipid fat replacers, respectively. Their effects on the physicochemical properties of reduced-fat mozzarella cheese were evaluated. RESULTS: Lecithin reduced the hardness and increased the degree of free oil released. Inulin and lecithin decreased the hydrophobic interaction of reduced-fat cheese. Maltitol improved the elasticity of the reduced-fat cheese and increased the hydrophobic interaction within the casein matrix. Maltitol-added cheese had a lower glass transition temperature (Tg ) than the other cheeses. Maltitol significantly improved the stretchability of the reduced-fat cheese. CONCLUSION: The results obtained in the present study suggest that maltitol is an effective fat replacer in reduced-fat mozzarella cheese and might enhance the cheese's functional properties. The Tg of cheese was related to the water and fat content, fat replacer addition and cross-linking degree of casein. The relationship between Tg and the physicochemical properties of cheese will be studied in further research. © 2019 Society of Chemical Industry.

Temporal sweetness and side tastes profiles of 16 sweeteners using temporal check-all-that-apply (TCATA).

An increase in consumer awareness around the negative health impacts of consuming excess added sugars has led to a rise in the replacement of sucrose in foods and beverages. This replacement is often through the use of low or no calorie sweeteners to reduce total calories while maintaining sweetness and palatability. There are a wide variety of sweeteners with diverse physical and caloric compositions which can be used at concentrations estimated to be equi-sweet to sucrose in food products. However, many of the available sweeteners are known to differ in their temporal profiles and may have other side and aftertastes alongside sweetness that need to be considered when comparing their suitability as potential sucrose substitutes. The objective of the current study was to profile and compare the temporal sweetness and qualitative differences of 15 sweeteners to sucrose across nutritive saccharide (sucrose, dextrose, fructose, allulose (D-psicose), palatinose (isomaltulose), sucrose-allulose mixture), nutritive polyol (maltitol, erythritol, mannitol, xylitol, sorbitol) and non-nutritive (acesulfame-k, aspartame, sucralose, stevia (rebaudioside-A or rebA), luo han guo (monk fruit extract or mogroside)) groups, at equi-sweet intensity to 10% w/v sucrose based on a previous psychophysical dose-response study. Using the Temporal Check-all-that-Apply (TCATA) method, 20 participants evaluated a set of 17 sweetener samples (including a sucrose duplicate) in triplicates across three 1-h sessions for the occurrence of six attributes (sweetness, bitterness, metallic taste, chemical taste, honey taste and mouth-drying) over a 60 s period. Sucrose was characterised by a rapid sweetness onset (within first 10 s) to peak citation, and subsequent decay of sweetness with minimal side tastes noted. Acesulfame-K, stevia (rebA) and luo han guo had prominent and long-lasting citations of the undesirable bitter, metallic and chemical side tastes and significantly lower sweetness citations. Allulose, erythritol, sorbitol, aspartame and sucralose were perceived to have bitter, metallic and chemical side tastes, but retained a largely sweet taste profile, with longer residual sweetness for aspartame and sucralose. Nutritive sweeteners dextrose, fructose, maltitol, mannitol, sucrose-allulose mixture, palatinose and xylitol had the most similar temporal and qualitative taste profiles when compared to sucrose, in terms of their sweetness onset, peak sweetness citations, sweetness decay, and side taste profiles.

Metabolomics biomarkers and the risk of overall mortality and ESRD in CKD: Results from the Progredir Cohort.

INTRODUCTION: Studies on metabolomics and CKD have primarily addressed CKD incidence defined as a decline on eGFR or appearance of albuminuria in the general population, with very few evaluating hard outcomes. In the present study, we investigated the association between metabolites and mortality and ESRD in a CKD cohort. SETTING AND METHODS: Data on 454 participants of the Progredir Cohort Study, Sao Paulo, Brazil were used. Metabolomics was performed by GC-MS (Agilent MassHunter) and metabolites were identified using Agilent Fiehn GC/MS and NIST libraries. After excluding metabolites present in <50% of participants, 293 metabolites were analyzed. An FDR q value <0.05 criteria was applied in Cox models on the composite outcome (mortality or incident renal replacement therapy) adjusted for batch effect, resulting in 34 metabolites associated with the outcome. Multivariable-adjusted Cox models were then built for the composite outcome, death, and ESRD incident events. Competing risk analysis was also performed for ESRD. RESULTS: Mean age was 68±12y, mean eGFR-CKDEPI was 38.4±14.6 ml/min/1.73m2 and 57% were diabetic. After adjustments (GC-MS batch, sex, age, DM and eGFR), 18 metabolites remained significantly associated with the composite outcome. Nine metabolites were independently associated with death: D-malic acid (HR 1.84, 95%CI 1.32-2.56, p = 0.0003), acetohydroxamic acid (HR 1.90, 95%CI 1.30-2.78, p = 0.0008), butanoic acid (HR 1.59, 95%CI 1.17-2.15, p = 0.003), and docosahexaenoic acid (HR 0.58, 95%CI 0.39-0.88, p = 0.009), among the top associations. Lactose (SHR 1.49, 95%CI 1.04-2.12, p = 0.03), 2-O-glycerol-α-D-galactopyranoside (SHR 1.76, 95%CI 1.06-2.92, p = 0.03), and tyrosine (SHR 0.52, 95%CI 0.31-0.88, p = 0.02) were associated to ESRD risk, while D-threitol, mannitol and myo-inositol presented strong borderline associations. CONCLUSION: Our results identify specific metabolites related to hard outcomes in a CKD population. These findings point to the need of further exploration of these metabolites as biomarkers in CKD and the understanding of the underlying biological mechanisms related to the observed associations.

Development of Alditol Acetate Derivatives for the Determination of 15N-Enriched Amino Sugars by Gas Chromatography-Combustion-Isotope Ratio Mass Spectrometry.

Amino sugars can be used as indices to evaluate the role of soil microorganisms in active nitrogen (N) cycling in soil. This paper details the assessment of the suitability of gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS) for the analysis of 15N-enriched amino sugars as alditol acetate derivatives prior to application of a novel 15N stable isotope probing (SIP) approach to amino sugars. The efficient derivatization and cleanup of alditol acetate derivatives for GC was achieved using commercially available amino sugars, including glucosamine, mannosamine, galactosamine, and muramic acid, as laboratory standards. A VF-23ms stationary phase was found to produce optimal separations of all four compounds. The structure of the alditol acetate derivatives was confirmed using gas chromatography/mass spectrometry (GC/MS). For GC-C-IRMS determinations, implementation of a two-point normalization confirmed the optimal carrier gas flow rate to be 1.7 mL min-1. Linearity of δ15N value determinations up to δ15Nt of 469 ± 3.1‰ (where δ15Nt is the independently measured δ15N value) was confirmed when 30 nmol N was injected on-column, with the direction of deviation from δ15Nt at low sample amount dependent on the 15N abundance of the analyte. Observed between- and within-run memory effects were significant ( P < 0.007) when a highly enriched standard (469 ± 3.1‰) was run; therefore, analytical run order and variation in 15N enrichment of analytes within the same sample must be considered. The investigated parameters have confirmed the isotopic robustness of alditol acetate derivatives of amino sugars for the GC-C-IRMS analysis of 15N-enriched amino sugars in terms of linearity over an enrichment range (natural abundance to 469 ± 3.1‰) with on-column analyte amount over 30 nmol N.

Development and validation of HILIC-UHPLC-ELSD methods for determination of sugar alcohols stereoisomers and its application for bioconversion processes of crude glycerin.

The recent increase in the production of crude glycerin through the manufacture of biodiesel has imputed a commercial issue, the excess of this raw material in the market and its constant devaluation, which resulted in the need for new technologies for its use. Crude glycerin can be used in biotechnological processes for the production of high value-added compounds. This study presents novel, simple and fast methods based on ultra-high performance liquid chromatography (UHPLC) using evaporative light scattering detection (ELSD) for simultaneous analysis of ten sugar alcohols with a hydrophilic interaction chromatography (HILIC) column. The selected compounds and their possible stereoisomers have major commercial importance and they can be obtained by biotechnological routes. Under optimized conditions, threitol, erythritol, adonitol, xylitol, arabitol, iditol, sorbitol, mannitol, dulcitol and volemitol can be analyzed simultaneously within 15.0 min. The use of different column temperatures was a key parameter to reach the selectivity during the separation of some stereoisomers. Regression equations revealed a good linear relationship (R > 0.995) over the range from 50.0 to 800.0 ng. Limits of detection (LOD) and quantification (LOQ) ranged from 30.0 to 45.0 ng and 50.0-75.0 ng, respectively. The HILIC-UHPLC-ELSD methods showed good precision with low coefficient of variation (CV%) for the intra- and inter-assays experiments (≤ 5.1%) and high repeatability in terms of retention times for each analyte (≤ 0.5%). The accuracy was confirmed with an average recovery ranging from 92.3 to 107.3%. The developed methods employ an analytical technique more accessible and suitable for routine analyzes and have shown to be suitable for simultaneous analysis of sugar alcohols present in crude bioconverted glycerin samples using different classes of microorganisms.

Chemical and bioactivity screening of subcritical water extracts of chokeberry (Aronia melanocarpa) stems.

Subcritical water extracts of chokeberry (Aronia melanocarpa) stems were chemically and biologically characterised. Chemical profile was defined by GC-MS analysis whereas anti-oxidant, anti-diabetic and tyrosinase-inhibitory activities of the extracts were investigated by in vitro assays. Antioxidant activity assays revealed strong activity against DPPH radical (IC50 = 0.1 mg/mL) and reducing power (IC50 = 0.25 mg/mL). The extracts demonstrated remarkable amylase (0.59 mmol ACAE/g) and glucosidase (7.50 mmol ACAE/g) inhibitory effects. Anti-tyrosinase activity of aronia stem extracts obtained by subcritical water was calculated to be 15.87 mg KAE/g extract. GC-MS analysis of chokeberry stem subcritical water extracts revealed the presence of different chemical classes. The compounds present in the highest concentrations were polyols arabitol (13.7%), xylitol (3.5%), and glycerol (1.96%), as well as sugars such as fructose (3.04%), ribose (1.99%) and xylulose (1.18%).

Varying Inocula Permutations (Aspergillus oryzae and Bacillus amyloliquefaciens) affect Enzyme Activities and Metabolite Levels in Koji.

In this study, we investigated the altered enzymatic activities and metabolite profiles of koji fermented using varying permutations of Aspergillus oryzae and/or Bacillus amyloliquefaciens. Notably, the protease and ß-glucosidase activities were manifold increased in co-inoculated (CO) koji samples (co-inoculation of A. oryzae and B. amyloliquefaciens). Furthermore, gas chromatography-mass spectrometry (GC-MS)-based metabolite profiling indicates that levels of amino acids, organic acids, sugars, sugar alcohols, fatty acids, nucleosides, and vitamins were distinctly higher in CO, SA (sequential inoculation of A. oryzae, followed by B. amyloliquefaciens), and SB (sequential inoculation of B. amyloliquefaciens, followed by A. oryzae). The multivariate principal component analysis (PCA) plot based on GC-MS datasets indicated a clustered pattern for MA and MB (koji samples inoculated either with A. oryzae or B. amyloliquefaciens) across PC2 (20.0%). In contrast, the CO, SA, and SB metabolite profiles displayed segregated patterns across PLS1 (22.2%) and PLS2 (21.1%) in the partial least square discriminant analysis (PLS-DA) model. Intriguingly, the observed disparity in the levels of primary metabolites was engendered largely by higher relative levels of sugars and sugar alcohols in MA, SA, and CO koji samples, which was commensurate with the relative amylase activities in respective samples. Collectively, the present study emphasizes the utility of integrated biochemical and metabolomic approaches for achieving the optimal permutation of fermentative inocula for industrial koji preparation.