Identification of potential quality markers in Indonesia's Arabica specialty coffee using GC/MS-based metabolomics approach.

INTRODUCTION: The cupping test is a widely used method for quality assessment of Arabica coffee. However, the cupping test is limited by the low number of certified panelists and the low throughput. Therefore, an analytical-based quality assessment may be a promising tool to complement the cupping test. A present, there is no report investigating quality marker candidates, focusing only on "specialty" grade Arabica coffee from Indonesia. OBJECTIVE: This study identified the potential quality marker(s) in Arabica Specialty coffee at different stages (green beans, roasted beans, and brewed coffee. METHODS: The metabolite profiles of ten different Arabica specialty-grade coffees were analyzed with different cup scores using gas chromatography-mass spectrometry (GC/MS). From the ten samples, green coffee beans, roasted coffee beans, and brewed coffee were selected. In addition, an orthogonal projection to latent structure (OPLS) regression analysis was conducted to obtain a potential quality marker based on the variable importance in projection (VIP). The potential quality marker(s) were validated by GC/MS metabolome profiling and OPLS analysis of different sets of samples consisting of 35 Arabica specialty-grade coffee samples. RESULTS: In Arabica coffee samples, the OPLS model of the three stages showed galactinol to have a high VIP score. Galactinol showed a consistent positive correlation with cup scores at all stages of coffee production (green beans, roasted beans, and brewed coffee). The correlation suggests galactinol is a potential quality marker after further validation using different samples. CONCLUSION: GC/MS combined with OPLS regression analysis suggested galactinol as a quality marker and provide an early screening method for Arabica coffee quality that complements the cupping test performed by certified panelists.

Metabolome analysis to investigate the effect of controlled fermentation on taste-related metabolites in terasi.

INTRODUCTION: Terasi is a fermented shrimp paste unique to Indonesia and is used in dishes to add umami and saltiness. In a previous study, the controlled fermentation of terasi was optimized using starters containing three bacterial isolates: Staphylococcus saprophyticus, Bacillus subtilis, and Lactobacillus murinus. However, the influence of controlled fermentation using these starters on the metabolites in terasi has not yet been studied. OBJECTIVES: Therefore, this study aimed to investigate the effect of controlled fermentation on taste-related metabolites in terasi using a metabolomics approach. RESULTS: Non-targeted analysis indicated that amino acids contributed to variations during fermentation. Subsequently, targeted analysis of amino acids revealed that terasi subjected to controlled fermentation using a starter with a 2:1:2 ratio of S. saprophyticus, B. subtilis, and L. murinus, respectively, resulted in a product containing D-amino acids, such as D-Asp, D-Gln, and D-Leu that was unique when compared to other terasi products prepared using controlled fermentation. Genetic analysis of isolates from the terasi produced using controlled fermentation was also carried out, and this is the first study to suggest that Staphylococcus spp. has the potential to produce D-amino acids. CONCLUSION: In conclusion, the ratio of bacterial species in starter cultures used in controlled fermentation influenced the amino acid profile of the product and starters with a higher ratio of Staphylococcus spp. may result in the production of D-amino acids.

Gas chromatography/mass spectrometry-based metabolite profiling of coffee beans obtained from different altitudes and origins with various postharvest processing.

INTRODUCTION: Coffee is a popular beverage because of its pleasant aroma and distinctive flavor. The flavor of coffee results from chemical transformations influenced by various intrinsic and extrinsic factors, including altitude, geographical origin, and postharvest processing. Despite is the importance of grading coffee quality, there is no report on the dominant factor that influences the metabolomic profile of green coffee beans and the correlated metabolites for each factor. OBJECTIVE: This study investigated the total metabolite profile of coffees from different altitudes and coffees subjected to different postharvest processing. METHOD: Arabica green coffee beans obtained from different geographical origins and different altitudes (400 and 800 m) and produced by different postharvest processes (dry, honey, and washed process) were used in this study. Coffee samples obtained from altitudes of 400-1600 m above sea level from various origins that were produced by the washed method were used for further study with regard to altitudes. Samples were subjected to gas chromatography/mass spectrometry (GC/MS) analysis and visualized using principal component analysis (PCA) and orthogonal partial least squares (OPLS) regression analysis. RESULTS: The PCA results showed sample separation based on postharvest processing in PC1 and sample separation based on altitude in PC2. A clear separation between samples from different altitudes was observed if the samples were subjected to the same postharvest processing method, and the samples were of the same origin. Based on this result, OPLS analysis was conducted using coffee samples obtained from various altitudes with the same postharvest processing. An OPLS model using altitude as a response variable and 79 metabolites annotated from the GC/MS analysis as an explanatory variable was constructed with good R2 and Q2 values. CONCLUSION: Postharvest processing was found to be the dominant factor affecting coffee metabolite composition; this was followed by geographical origin and altitude. The metabolites glutamic acid and galactinol were associated with the washed and honey process, while glycine, lysine, sorbose, fructose, glyceric acid, and glycolic acid were associated with the dry process. Two metabolites with high variable influence on projection scores in the OPLS model for altitude were inositol and serotonin, which showed positive and negative correlations, respectively. This is the first study to report characteristic coffee metabolites obtained from different altitudes.

Shrimp count size: GC/MS-based metabolomics approach and quantitative descriptive analysis (QDA) reveal the importance of size in white leg shrimp (Litopenaeus vannamei).

INTRODUCTION: "Count size" is a term used to represent the number of shrimps in one pound or kilogram that applies globally in the shrimp industry. Based on shrimp body weight, count sizes range over the smallest (> 70) up to the largest size (U15) of shrimp. Large shrimps are considered highly palatable; therefore, they are priced higher than the small shrimps. However, the pricing of shrimp has not been based on scientific findings since there have been no studies reporting the correlation between shrimp quality and shrimp size. OBJECTIVE: In this study, we aimed to investigate the importance of shrimp size in terms of metabolite profile and sensory properties. METHODS: Nine groups of Litopenaeus vannamei, categorized based on their body weight similarity, were collected from various sampling sites regardless of the difference in days of culture (count size 16/20, 21/25, 26/30, 41/50, and 51/60). Gas chromatography/mass spectrometry (GC/MS)-based metabolomics analysis was employed to characterize their metabolite profiles. Furthermore, a robust PLS regression model was constructed to predict the shrimp size using metabolome data. Following this, the difference in sensory attributes among commercial shrimp count sizes 21/25-41/50 was confirmed using quantitative descriptive analysis (QDA). RESULTS: Small shrimp (> 70-51/60) had higher accumulation of proteinogenic and non-proteinogenic amino acids, sugars, and organic acids compared to large shrimps (41/50-16/20). The QDA of commercial count sizes (21/25-41/50) performed by trained panelists showed that sweetness, juiciness, crispness, and red color attributes increased with an increase in shrimp size. Based on the PLS model, proline as a sweet-tasting metabolite also showed an increased level along with the shrimp size. CONCLUSIONS: These findings demonstrate the importance of shrimp count size with regard to shrimp quality.

GC/MS-based metabolic profiling for the evaluation of solid state fermentation to improve quality of Arabica coffee beans.

INTRODUCTION: Coffee fermentation has been reported as one key process in aroma and flavor development of coffee. However, natural fermentation often results in inconsistent quality of coffee. In this study, second fermentation using isolates from feces of civet (Luwak) and Cilembu sweet potato were used to improve the quality of Arabica green coffee beans. OBJECTIVES: The aim of this research was to improve the quality of various Arabica coffee from different origins in Indonesia by controlled-second fermentation. METHOD: The Arabica coffee beans used in this study were from three origins in Indonesia: Kintamani-Bali (I), Aceh-Gayo (II) and Nagarawangi-Sumedang (III). The second fermentation was done using three bacterial isolates coded as BF5C(2); UciSp14; and AF7E which belong to Bacillus genus. Quality assessment of fermented coffee was performed by cupping test following Specialty Coffee Association of America (SCAA) protocol by licensed Q graders, GC/MS metabolite profiling, and total polyphenol content measurement. RESULTS: The controlled-second fermentation for 4-8 h was successful to increase total polyphenol content well as to improve the complexity of coffee taste and coffee quality (cupping score > 84). Comparative GC/MS analysis showed that fermentation of coffee beans resulted in alteration of metabolite profiles of coffee beans compared with control, while still maintaining the characteristics of coffee based on each origin. CONCLUSION: The controlled-second fermentation was effective to increase the quality of coffee and alter metabolite composition of coffee that were associated with changes in taste profile of coffee. This report may serve as basis for producing coffee with better taste quality with higher polyphenols content through standardized fermentation production in industrial scale.

Metabolite profiling of whiteleg shrimp Litopenaeus vannamei from super-intensive culture in closed aquaculture systems: a recirculating aquaculture system and a hybrid zero water discharge-recirculating aquaculture system.

INTRODUCTION: The production of the whiteleg shrimp Litopenaeus vannamei now accounts for approximately 75% of the total shrimp production in Indonesia. The techniques used to produce whiteleg shrimp in Indonesia are still dominated by conventional rearing strategies using open-pond systems, which often contribute to unpredictable culture performance and weak sustainability. Alternative production strategies of closed aquaculture systems, including the recirculating aquaculture system (RAS) and hybrid zero water discharge-recirculating aquaculture system (hybrid system), have been developed and implemented for higher productivity, stability and sustainability of whiteleg shrimp grow-out production in Indonesia. Despite the positive aspects of the application of closed aquaculture systems in shrimp aquaculture, the differences in the characteristics of shrimp grown in closed RAS and hybrid systems compared to open-pond systems remain unclear. OBJECTIVE: This study aims to investigate the differences in the metabolite profiles of shrimp grown in intensive closed aquaculture systems, including an RAS and hybrid system, compared to those of shrimp grown in a semi-intensive, open, earthen pond system by means of non-targeted GC-MS metabolite profiling. METHODS: Shrimp cultured in the closed systems (RAS and hybrid system) and an open system (pond) were harvested and subjected to GC-MS non-targeted metabolomics analysis. A total of 112 metabolites were annotated from shrimp samples and subjected to principal component analysis (PCA). RESULTS: The metabolites annotated from GC-MS mainly included organic compounds, proteinogenic and non-proteinogenic amino acids, sugars, nucleosides and fatty acids. The results of principal component analysis showed several metabolites with high variable importance in projection (VIP) scores, including shikimic acid, ß-alanine, uric acid, hypoxanthine, inosine, homocysteine, methionine, phenylalanine, tryptophan and lysine, as the main metabolites differentiating the shrimp grown in the three production systems. CONCLUSION: Our findings showed that shrimp cultured in different aquaculture systems exhibited distinct metabolite profiles, and the metabolites showing high VIP scores, including shikimic acid, ß-alanine, uric acid, hypoxanthine, inosine, homocysteine, methionine, phenylalanine, tryptophan and lysine, may serve as candidate markers to indicate the differences in shrimp from different production systems.

Metabolic repair through emergence of new pathways in Escherichia coli.

Escherichia coli can derive all essential metabolites and cofactors through a highly evolved metabolic system. Damage of pathways may affect cell growth and physiology, but the strategies by which damaged metabolic pathways can be circumvented remain intriguing. Here, we use a ΔpanD (encoding for aspartate 1-decarboxylase) strain of E. coli that is unable to produce the ß-alanine required for CoA biosynthesis to demonstrate that metabolic systems can overcome pathway damage by extensively rerouting metabolic pathways and modifying existing enzymes for unnatural functions. Using directed cell evolution, rewiring and repurposing of uracil metabolism allowed formation of an alternative ß-alanine biosynthetic pathway. After this pathway was deleted, a second was evolved that used a gain-of-function mutation on ornithine decarboxylase (SpeC) to alter reaction and substrate specificity toward an oxidative decarboxylation-deamination reaction. After deletion of both pathways, yet another independent pathway emerged using polyamine biosynthesis, demonstrating the vast capacity of metabolic repair.

GC/MS based metabolite profiling of Indonesian specialty coffee from different species and geographical origin.

INTRODUCTION: The consumption of high quality coffee such as specialty Arabica and fine Robusta coffee is increasing steadily in recent years. Development of single origin coffee is an important strategy to maintain coffee quality, grade and high cupping score. Indonesia is a top exporting country for Arabica coffee with high variety of specialty coffees from different origins. Despite its long standing reputation in global coffee market, very few is known about the variability among Indonesian specialty coffees. OBJECTIVES: This study aims to observe metabolite variability among Indonesian coffees from different species and geographical origins by means of non-targeted GC/MS metabolite profiling. METHODS: Sixty-four compounds were tentatively identified from 16 green and roasted coffee beans from different species and cultivation areas in Indonesia and were subjected to principal component analysis (PCA). Ten Specialty Arabica coffee and five Fine Robusta representing all important high quality coffees of Indonesia were also analyzed independently to further classify Indonesian coffee according to their origin. RESULTS: PCA results of 16 green and roasted coffee beans of different species and cultivation areas showed that samples were separated along PC1 based on different roasting condition (green and roasted) with 52.9% variance and were separated along PC2 based on different species with 19.3% variance. The result from this study showed the clustering of samples based on three major cultivation areas in Indonesia (western, central, eastern part). Metabolites showing higher concentration in Sulawesi, Papua, Flores and Sumatra samples were glycerol, glucuno-1,5-lactone, gluconic acid and sorbitol. A clear distinction in galactitol and galactinol concentration between all samples from eastern part of Indonesia and western and middle part of Indonesia was also observed. CONCLUSIONS: Our results showed that each region (western, central and eastern part of Indonesia) has signature compounds that may serve as discriminant markers for coffee authentication. This is the first report on the classification of Indonesian specialty coffee based on their metabolic profiles and can act as a basis for marker identification for routine procedures in industry.

Directed strain evolution restructures metabolism for 1-butanol production in minimal media.

Engineering a microbial strain for production sometimes entails metabolic modifications that impair essential physiological processes for growth or production. Restoring these functions may require amending a variety of non-obvious physiological networks, and thus, rational design strategies may not be practical. Here we demonstrate that growth and production may be restored by evolution that repairs impaired metabolic function. Furthermore, we use genomics, metabolomics and proteomics to identify several underlying mutations and metabolic perturbations that allow metabolism to repair. Previously, high titers of butanol production were achieved by Escherichia coli using a growth-coupled, modified Clostridial CoA-dependent pathway after all native fermentative pathways were deleted. However, production was only observed in rich media. Native metabolic function of the host was unable to support growth and production in minimal media. We use directed cell evolution to repair this phenotype and observed improved growth, titers and butanol yields. We found a mutation in pcnB which resulted in decreased plasmid copy numbers and pathway enzymes to balance resource utilization. Increased protein abundance was measured for biosynthetic pathways, glycolytic enzymes have increased activity, and adenosyl energy charge was increased. We also found mutations in the ArcAB two-component system and integration host factor (IHF) that tune redox metabolism to alter byproduct formation. These results demonstrate that directed strain evolution can enable systematic adaptations to repair metabolic function and enhance microbial production. Furthermore, these results demonstrate the versatile repair capabilities of cell metabolism and highlight important aspects of cell physiology that are required for production in minimal media.

Identifying metabolic elements that contribute to productivity of 1-propanol bioproduction using metabolomic analysis.

INTRODUCTION: Previously constructed Escherichia coli strains that produce 1-propanol use the native threonine pathway, or a heterologous citramalate pathway. However, based on the energy and cofactor requirements of each pathway, a combination of the two pathways produces synergistic effects that increase the theoretical maximum yield with a simultaneous unexplained increase in productivity. OBJECTIVE: Identification of key factors that contribute to synergistic effect leading to 1-propanol yield and productivity improvement in E. coli with native threonine pathway and heterologous citramalate pathway. METHOD: A combination of snapshot metabolomic profiling and dynamic metabolic turnover analysis were used to identify system-wide perturbations that contribute to the productivity improvement. RESULT AND CONCLUSION: In the presence of both pathways, increased glucose consumption and elevated levels of glycolytic intermediates are attributed to an elevated phosphoenolpyruvate (PEP)/pyruvate ratio that is known to increase the function of the native phosphotransferase. Turnover analysis of nitrogen containing byproducts reveals that ammonia assimilation, required for the threonine pathway, is streamlined when provided with an NAD(P)H surplus in the presence of the citramalate pathway. Our study illustrates the application of metabolomics in identification of factors that alter cellular physiology for improvement of 1-propanol bioproduction.

Characteristics of kopyor coconut (Cocos nucifera L.) using sensory analysis and metabolomics-based approach.

Kopyor is a coconut with unique characteristics from Indonesia, one of the largest coconut producers in the world. Kopyor is an edible mature coconut with soft endosperm. Although this fruit is one of the most popular coconuts in the world, there are limited studies on its properties, including its sensory attributes and metabolite profiles. This study investigates the characteristics of kopyor using sensory evaluation, a widely targeted metabolomics approach, and multivariate analysis. The liquid (water) and solid (flesh) endosperms were collected as the samples. The results showed that kopyor has characteristics that distinguish it from normal mature and young coconuts. Kopyor water has a milky, creamy, nutty, bitter, and astringent taste with an oily aftertaste and mouthfeel. Kopyor flesh is soft and moist and gives a sandy mouth feel. This study analyzed the sensory attributes of the kopyor endosperm for the first time and compared it with those of normal mature and young coconuts. A gas chromatography mass spectrometry analysis showed that kopyor contained wider variety of metabolites than normal coconuts of the same age. Based on the differential analysis and orthogonal projections to latent structures-regression, kopyor water was characterized by the accumulation of flavor-related metabolites, such as amino acids and organic acids, which contributed to its sensory complexity. This study solidified the effects of maturation and endosperm type on metabolite accumulation in kopyor endosperm. This pioneering information will lead to the future use of kopyor and other unique coconuts worldwide for food, contributing to the sustainability of the coconut industry.

Effects of n-butanol production on metabolism and the photosystem in Synecococcus elongatus PCC 7942 based on metabolic flux and target proteome analyses.

Although n-butanol (BuOH) is an ideal fuel because of its superior physical properties, it has toxicity to microbes. Previously, a Synechococcus elongatus PCC 7942 derivative strain that produces BuOH from CO2 was developed by introducing six heterologous genes (BUOH-SE strain). To identify the bottleneck in BuOH production, the effects of BuOH production and its toxicity on central metabolism and the photosystem were investigated. Parental (WT) and BUOH-SE strains were cultured under autotrophic conditions. Consistent with the results of a previous study, BuOH production was observed only in the BUOH-SE strain. Isotopically non-stationary 13C-metabolic flux analysis revealed that the CO2 fixation rate was much larger than the BuOH production rate in the BUOH-SE strain (1.70 vs 0.03 mmol gDCW-1 h-1), implying that the carbon flow for BuOH biosynthesis was less affected by the entire flux distribution. No large difference was observed in the flux of metabolism between the WT and BUOH-SE strains. Contrastingly, in the photosystem, the chlorophyll content and maximum O2 evolution rate per dry cell weight of the BUOH-SE strain were decreased to 81% and 43% of the WT strain, respectively. Target proteome analysis revealed that the amounts of some proteins related to antennae (ApcA, ApcD, ApcE, and CpcC), photosystem II (PsbB, PsbU, and Psb28-2), and cytochrome b6f complex (PetB and PetC) in photosystems decreased in the BUOH-SE strain. The activation of photosynthesis would be a novel approach for further enhancing BuOH production in S. elongatus PCC 7942.

Chitosan-based coating enriched with melezitose alters primary metabolites in fresh-cut pineapple during storage.

Demand for minimally processed fresh fruit is increasing due to its convenience. However, the distribution of fresh-cut fruits is limited because of their short shelf life. Pineapple, a popular tropical fruit, sold in fresh-cut form has a shelf life of approximately 5-7 days at 4 °C. Chitosan, an edible coating, is commonly used to prolong the shelf life of food products. Similarly, the sugar melezitose has been reported to change during pineapple ripening and may play a role in regulating the shelf life of pineapple. However, the direct effects of this sugar have yet to be studied. The objective of this study was to investigate the effect of chitosan coating with melezitose to prolong the shelf life of fresh-cut pineapple. Full-ripe Bogor pineapples from Okinawa, Japan, were cut into cubes and soaked in either chitosan 1.25%, melezitose 5 mg/L, or chitosan+melezitose and stored for 5 days under dark conditions (23.6 ± 0.5 °C; relative humidity, 40.0 ± 10.4%). None of the treatments significantly altered the weight loss or color changes in the fresh-cut fruit. However, treatment significantly altered the primary metabolites, namely quinic acid, sucrose, and xylitol based on orthogonal projection to latent structures data with the screening from p-value score. Moreover, cell-wall metabolism is possibly affected in pineapple cut fruit treated by chitosan-melezitose as shown from metabolite sets enrichment analysis. This study showed that chitosan added with melezitose might have potential to prolong the shelf-life of fresh-cut pineapple, providing a basis for further post-harvest studies of the whole pineapple fruit.

Improvement of the functional value of green soybean (edamame) using germination and tempe fermentation: A comparative metabolomics study.

Green soybean, also known as edamame, is a legume with high nutritional and functional value. Despite its growing popularity and potential health benefits, the functionality of green soybean has not been thoroughly studied. Previous research on the functionality of green soybean has largely focused on a limited number of specific, well-studied, bioactive metabolites, without comprehensively investigating the metabolome of this legume. Additionally, very few studies have explored the improvement of the functional value of green soybean. This study aimed to investigate the metabolome profile of green soybean, identify bioactive metabolites, and to further explore the potential improvement of the identified bioactive metabolites using germination and tempe fermentation. A total of 80 metabolites were annotated from green soybean using GC-MS and HPLC-PDA-MS. Among them, 16 important bioactive metabolites were identified: soy isoflavones daidzin, glycitin, genistin, malonyl daidzin, malonyl genistin, malonyl glycitin, acetyl daidzin, acetyl genistin, acetyl glycitin, daidzein, glycitein, and genistein, as well as other metabolites including 3,4-dihydroxybenzoic acid, 3-hydroxyanthranillic acid, 3-hydroxy-3-methylglutaric acid (meglutol), and 4-aminobutyric acid (GABA). Germination and tempe fermentation techniques were employed to potentially improve the concentrations of these bioactive metabolites. While showing improvements in amino acid contents, germination process did not improve bioactive metabolites significantly. In contrast, tempe fermentation was found to significantly increase the concentrations of daidzein, genistein, glycitein, acetyl genistin, acetyl daidzin, 3-hydroxyanthranillic acid, and meglutol (>2-fold increase with p < 0.05) while also improving amino acid levels. This study highlights the potentials of germination and fermentation to improve the functionality of legumes, particularly green soybean.

Antioxidant Activity and Metabolite Profiling of Xylocarpus granatum Extracts Using Gas Chromatography-Mass Spectrometry.

The potential application of Xylocarpus granatum, a mangrove species, as traditional medicine has been widely linked to its high secondary metabolite and antioxidant contents. However, few studies have been reported to identify and classify active metabolites responsible for such excellent biological activities. Therefore, the aim of this work was to determine the antioxidant activity, identify the metabolite profiles, and predict the metabolites acting as antioxidants in X. granatum extract using a gas chromatography-mass spectrometry (GC-MS)-based metabolomics approach. The seeds, stems, fruit peel, pulp, leaves, and twigs of X. granatum were macerated with ethanol. Each extract was analyzed with GC-MS, and the data were processed using mass spectrometry data-independent analysis (MS-DIAL) software to identify the metabolites. The IC50 value of plant parts of X. granatum ranged from 7.73 to 295 ppm. A total of 153 metabolites were identified and confirmed in the X. granatum extracts. Among the identified metabolites, epicatechin and epigallocatechin were the two most abundant in the stem extracts and are expected to have the greatest potential as antioxidants. Principal component analysis (PCA) succeeded in grouping all parts of the plant into three groups based on the composition of the metabolites: group 1 (stems, fruit peel, and twigs), group 2 (seeds and pulp), and group 3 (leaves).

Metabolite Changes in Indonesian Tempe Production from Raw Soybeans to Over-Fermented Tempe.

Tempe is fermented soybean from Java, Indonesia, that can serve as a functional food due to its high nutritional content and positive impact on health. Although the tempe fermentation process is known to affect its nutrient content, changes in the metabolite profile during tempe production have not been comprehensively examined. Thus, this research applied a metabolomics approach to investigate the metabolite profile in each step of tempe production, from soybean soaking to over-fermentation. Fourteen samples of raw soybeans, i.e., soaked soybeans (24 h), steamed soybeans, fungal fermented soybeans, and over-fermented soybeans (up to 72 h), were collected. Untargeted metabolomics by gas chromatography/mass spectrometry (GC-MS) was used to determine soybean transformations from various fermentation times and identify disparity-related metabolites. The results showed that soybeans samples clustered together on the basis of the different fermentation steps. The results also showed that sugar, sugar alcohol, organic acids, and amino acids, as well as fermentation time, contributed to the soybean metabolite profile transformations. During the fermentation of tempe, sugars and sugar alcohols accumulated at the beginning of the process before gradually decreasing as fermentation progressed. Specifically, at the beginning of the fermentation, gentiobiose, galactinol, and glucarate were accumulated, and several metabolites such as glutamine, 4-hydroxyphenylacetic acid, and homocysteine increased along with the progression of fermentation. In addition, notable isoflavones daidzein and genistein increased from 24 h of fermentation until 72 h. This is the first report that provides a complete description of the metabolic profile of the tempe production from soybean soaking to over-fermentation. Through this study, the dynamic changes at each step of tempe production were revealed. This information can be beneficial to the tempe industry for the improvement of product quality based on metabolite profiling.

Unique metabolite profiles of Indonesian cocoa beans from different origins and their correlation with temperature.

Chocolate flavors vary depending on the origin of the cocoa beans used. Differences in soil, microorganisms, and environmental factors contribute to the formation of flavor precursors in cocoa beans. During cocoa bean fermentation, environmental temperature has been shown to alter metabolite concentrations. However, the correlation between the metabolite profile of cocoa beans and the temperature of their region of origin has not been fully defined. In this study, the metabolite profiles of Indonesian cocoa beans of various origins were evaluated using gas chromatography/mass spectrometry-based analysis, and were found to differ depending on the origin of the bean. Subsequently, the correlation between metabolite profile and environmental temperature of the origin was assessed using orthogonal projection to latent structure regression (OPLS-R) analysis. The analysis revealed that seven metabolites were associated with temperature: γ-aminobutyric acid (GABA), ethanolamine, glycerol, isocitric acid + citric acid, succinic acid, malic acid, and saccharic acid. The findings of this study will be valuable to chocolate industries for the production of single-origin chocolates.

Investigation of the effect of processing on the component changes of single-origin chocolate during the bean-to-bar process.

Chocolate is one of the most popular sweets in the world. In recent years, the bean-to-bar process for chocolate production has attracted global attention. Bean-to-bar is a method of managing the whole production process from cocoa beans to chocolate bars, including single-origin chocolate (SOC). Many manufacturers aim to produce high-quality chocolate to maximize the flavor of cocoa beans. However, chocolate compounds are very complex due to many processes, and there are a limited number of studies on the SOC produced from the bean-to-bar process. Therefore, understanding the effects of processing is important for the growth of the chocolate industry. The objective of this study was to investigate the processing effect on the component changes of SOC during the bean-to-bar process. In this study, the component changes during the bean-to-bar process were monitored using gas chromatography/mass spectrometry (GC/MS). Then, the characteristics of SOC from five regions in Indonesia were further investigated. Lastly, the component profiles were combined with the data obtained from sensory evaluation. Our results showed that the influence of the manufacturing process was greater than that of the difference in the cocoa production area. Moreover, 1-pentanol, raffinose, and heptanoic acid were correlated with sweetness and dairy flavor, whereas glutamic acid, tartaric acid, 3-methyl-2-butanone, mannitol, and ethyl cinnamate were correlated with bitterness, astringency, and cocoa flavor, which were shown to be affected by fermentation, roasting, and sugar addition. This information might provide a basis for improving the chocolate production process and its quality related to the component profiles.

Effects of Soaking Tempe in Vinegar on Metabolome and Sensory Profiles.

Tempe is a fermented soybean food that is globally renowned for its high protein content. Methods of preparing tempe vary worldwide, and include soaking in vinegar before fermentation. This study aimed to determine the effects of soaking in vinegar by metabolome analysis, gas chromatography/mass spectrometry, and sensory attribute evaluation. Vinegar affected metabolism during tempe fermentation, which led to altered metabolite profiles in the final product. We validated the metabolite profiles of two types of tempe using triangle tests and rate-all-that-apply (RATA) tests, which revealed that the sensory attributes of a golden-brown color, ammonia smell, pleasant smell, salty flavor, and acceptance significantly differed (p < 0.05) between the two types of tempe. A high concentration of specific amino acids in the control tempe explained a strong ammonia smell, saltiness, and darker golden-brown sensory attributes. Tempe soaked in vinegar contained high concentrations of metabolites associated with a roasted aroma and cooked meat. In conclusion, most RATA panelists who were being introduced to tempe preferred that soaked in vinegar to the control that was not.

MALDI Mass Spectrometry Imaging Reveals the Existence of an N-Acyl-homoserine Lactone Quorum Sensing System in Pseudomonas putida Biofilms.

Quorum sensing (QS) is generally used to describe the process involving the release and recognition of signaling molecules, such as N-acyl-homoserine lactones, by bacteria to coordinate their response to population density and biofilm development. However, detailed information on the heterogeneity of QS metabolites in biofilms remains largely unknown. Here, we describe the utilization of matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) to follow the production of specific metabolites, including QS metabolites, during Pseudomonas putida biofilm development. To do so, a method to grow an agar-based biofilm was first established, and MALDI-MSI was used to detect and visualize the distribution of QS metabolites in biofilms at different cultivation times. This study demonstrated that N-acyl-homoserine lactones are homogeneously produced in the early stages of P. putida biofilm formation. In contrast, the spatial distribution of quinolones and pyochelin correlated with the swarming motility of P. putida in mature biofilms. These two metabolites are involved in the production of extracellular polymeric substances and iron chelators. Our study thus contributes to establishing the specific temporal regulation and spatial distribution of N-acyl-homoserine lactone-related metabolites and quinolone and pyochelin in P. putida biofilms.