Comprehensive profiling of amino acids and derivatives in biological samples: A robust UHPLC-MS/MS method for investigating acute lung injury.

The severe respiratory dysfunctions associated with acute lung injury (ALI) and its sequelae have a high morbidity and mortality rate, are multifactorial, and lack a viable treatment. Considering the critical function that amino acids and derivatives play in the genesis of illnesses and the regulation of metabolic processes, monitoring the levels of metabolites associated with amino acids in biological matrices is necessary and interesting to study their pathological mechanisms. Exploring the dynamics of amino acids and derivatives level and searching for biomarkers provides improved clinical ideas for the diagnosis and treatment of ALI. Therefore, we developed an ultra-high-performance liquid chromatography-electrospray tandem mass spectrometry (UHPLC-MS/MS) method that can simultaneously determine the amino acid and derivatives metabolic levels to study amino acid profiles in different biological samples to facilitate clinical research of ALI. In this study, 48 amino acids and derivatives, including neurotransmitters, polyamines, purines, and other types, were quantified simultaneously in a fast, high-throughput, sensitive, and reliable manner within a 15-minute run time without derivatization. No relevant studies have been reported to quantify these 48 amino acid metabolites in three biological samples simultaneously. Satisfactory linearity (R > 0.995), inter-day and intra-day accuracy (85.17-112.67 % and 85.29-111.60 %, respectively), inter-day and intra-day precision (RSD < 13.80 % and RSD < 12.01 %, respectively), matrix effects (81.00 %-118.00 %), recovery (85.09 %-114.65 %) and stability (RSD < 14.72 %) were all demonstrated by the optimized method's successful validation for all analytes. In addition, the suggested method was effectively implemented in plasma, urine, and lung tissue from normal mice and mice with ALI, with the aim of finding potential biomarkers associated with ALI. Potential biomarkers were screened through multivariate statistical analysis and volcanic map analysis, and the changes of markers in ALI were again identified through heat map analysis and correlation analysis with biochemical indicators, which provided ideas and references for subsequent mechanism studies. Here, the technique created in this work offers a quick and dependable way to perform an integrated analysis of amino acids in a variety of biological materials, which can provide research ideas for understanding the physiopathological state of various diseases.

Application effects of NaCl substitute on the fermentation profile of Pixian douban (broad bean paste).

The effects of a reduced-salt substitute (composed of NaCl, sodium gluconate, KCl, L-histidine, and L-lysine) applied in the fermentation of traditional Pixian douban (PXDB) were explored in this study according to sensory quality, physicochemical characteristics, color, colony count, and the contents of free amino acids (FAAs), organic acids, and volatile flavor compounds. The results showed that the PXDB with a 15% salt substitution had the most attractive reddish-brown color, a mellow fragrance, and the lowest total colony count of the three pastes. The fermentation quality of the 15% salt substitute PXDB was superior to that of the control groups, its sensory quality was more readily accepted, and the contents of its amino acid nitrogen, FAAs and organic acids had increased by 0.1050, 0.3290, and 3.9068 mg/g, respectively. Moreover, the concentrations of the main aroma compounds in the PXDB containing the salt substitute were higher than those of the control. These included phenylethanol, 3-methylthiopropanol, isoamyl alcohol, furfural, benzaldehyde, phenylacetaldehyde, nonanal, isoamyl aldehyde, 4-ethylphenol, and, particularly, 2,6-dimethylpyrazine, which had increased as much as 100 times. Correlation analysis showed that Glu, Phe, Tyr, Gly, Leu, Val, Asp, Ile, citric acids, and succinic acids were all positively correlated with the main aroma and contributed to the generation of PXDB's characteristic flavor, and main aroma substances in turn positively influence PXDB flavor sensory attributes. Overall, these results showed the application of the 15% salt substitute during PXDB fermentation improved the quality of the paste and, thus, would benefit the development of reduced-salt PXDB.

Comparison of the targeted metabolomics and nutritional quality indices of the probiotic cheese enriched with microalgae.

The objective of this study is to evaluate the influence of mixed L. acidophilus LA-5 and enrichment with microalgae (C. vulgaris and A. platensis) on metabolomic formation in a brined cheese matrix. Microbiological, compositional, and metabolomic characterization were investigated during the ripening. It was found that the nutritional quality indices of the samples were based on amino acid and fatty acid characterization. Fifty-six metabolomics including fatty acids, amino acids, organic acids, minerals, and vitamins were detected using the HPLC-DAD, GC-MS, and ICP-OES-based methods. The results indicated that the enrichment with probiotic strain and microalgae led to an increase in the nutritional quality indices such as EAAI, NI, BV, MUFA/SFA, h/H, and DFA. The chemometric analysis (e.g. HCA and PCA) presented the variance between the cheese samples based on their attributes. The identification of cheese metabolomics throughout the ripening could be used for a better understanding of the functional ingredients-cheese matrix relationships and as a directive approach for novel dairy products in other metabolomic-related studies.

Shrimp Cultured in Low-Salt Saline-Alkali Water has a Better Amino Acid Nutrition and Umami─Comparison of Flavors between Saline-Alkali Water- and Seawater-Cultured Litopenaeus vannamei.

The advantages of Litopenaeus vannamei farming in saline-alkali water have gradually attracted attention, but few studies have focused on its flavor. In this study, L. vannamei cultured in saline-alkali water (SS) and ordinary seawater (CS) (both have a breeding time of 120 days) were selected for analysis (n = 5). High-performance liquid chromatography (HPLC) was used to measure free amino acids and flavoring nucleotides in the muscles of L. vannamei, while the taste activity value (TAV) and equivalent umami concentration (EUC) were used to analyze the degree of umami. The total essential amino acids (TEAA) in the SS group were 238.41 ± 46.24 mg/mL, significantly higher than that in the CS group (107.06 ± 15.65 mg/mL). The total amount of flavor nucleotides in the SS group was 2948.51 ± 233.66 µg/mL, significantly higher than those in the CS group (2530.37 ± 114.67 µg/mL). The content and TAV of some free amino acids (Glu, Cys-s) in the SS group were significantly higher. Meanwhile, due to the significant increase in IMP, the synergistic effect of free amino acids and flavored nucleotides leads to higher EUC. The significant separation of SS and CS samples in principal component analysis (PCA) indicates a significant difference between the two groups. Our results indicate that shrimp cultured in saline-alkali water has a stronger umami. This study enriches the basic theories related to the flavor of salt-alkali water crustaceans.

Evaluation of Individual Variation of d-Amino Acids in Human Plasma by a Two-Dimensional LC-MS/MS System and Application to the Early Diagnosis of Chronic Kidney Disease.

For the discovery of sensitive biomarkers of kidney function focusing on chiral amino acids, a multiple heart-cutting two-dimensional (2D) liquid chromatography-mass spectrometry (LC-MS)/MS system has been designed/developed. As the target analytes, alanine (Ala), aspartic acid, glutamic acid (Glu), leucine (Leu), lysine, methionine, phenylalanine (Phe), proline (Pro), serine (Ser), and valine were selected considering the presence of their d-forms in mammals. The 2D LC-MS/MS system consisted of the nonenantioselective reversed-phase separation of the target amino acids, the separations of the d- and l-enantiomers, and detection using MS/MS. Using the method, the plasma chiral amino acids, precolumn derivatized with 4-fluoro-7-nitro-2,1,3-benzoxadiazole, were isolated from other intrinsic substances, then determined without losing sensitivity by the fully automated whole-peak volume transfer operation from first to second dimension. In all of the tested plasma samples obtained from five healthy individuals and 15 patients with chronic kidney disease (CKD), the target chiral amino acids were determined without interference. In healthy individuals, the levels of all the tested d-amino acids were regulated in the low ranges. In contrast, the % d values of Glu, Leu, and Phe significantly increased with the progress of kidney dysfunction, besides the previously reported values of d-Ala, Pro, and Ser. Concerning Phe, the significant increase of the % d values (p < 0.05) was reported for the first time even in the mild CKD group compared to those of the healthy group; d-Phe might be a more sensitive marker than the previously reported d-forms. These results demonstrated the potential of these d-forms as the sensitive biomarkers of kidney function for the early diagnosis of CKD.

Quantification of SARS-CoV-2 monoclonal IgG mass fraction by isotope dilution mass spectrometry.

The availability of serology assays to measure antibodies against the SARS coronavirus 2 (SARS-CoV-2) expanded rapidly during the Covid-19 pandemic. The interchangeable use of such assays to monitor disease progression and immune protection requires their standardization, for which suitably characterized monoclonal antibody materials can be useful. The methods, based on isotope dilution mass spectrometry, to value assign the mass fraction of such a material in solution within the context of an international interlaboratory comparison study (CCQM-P216) are described. The mass fraction in solution of a humanized IgG monoclonal antibody (mAb) against the SARS-CoV-2 Spike glycoprotein in the study sample has been value assigned through a combination of liquid chromatography, isotope dilution mass spectrometry (LC-ID-MS) methods and size exclusion chromatography with UV detection (SEC-UV). The former were developed for the quantification of amino acids and proteotypic peptides as surrogate analytes of the mAb while the latter was applied for the determination of the relative monomeric mass fraction. High-resolution mass spectrometry (hrMS) allowed the molecular weight evaluation and ruled out the presence of significant impurities. Method trueness was assessed using a subclass homologous IgG1 material value assigned by amino acid analysis. The assigned mass fraction of monomeric SARS-CoV-2 IgG in solution was 390 ± 16 mg/g. The associated expanded uncertainty originated mainly from acid hydrolysis variability and Trypsin/Lys-C digestion variability and efficiency.

Plasma and serum metabolic analysis of healthy adults shows characteristic profiles by subjects' sex and age.

INTRODUCTION: Pre-analytical factors like sex, age, and blood processing methods introduce variability and bias, compromising data integrity, and thus deserve close attention. OBJECTIVES: This study aimed to explore the influence of participant characteristics (age and sex) and blood processing methods on the metabolic profile. METHOD: A Thermo UPLC-TSQ-Quantiva-QQQ Mass Spectrometer was used to analyze 175 metabolites across 9 classes in 208 paired serum and lithium heparin plasma samples from 51 females and 53 males. RESULTS: Comparing paired serum and plasma samples from the same cohort, out of the 13 metabolites that showed significant changes, 4 compounds related to amino acids and derivatives had lower levels in plasma, and 5 other compounds had higher levels in plasma. Sex-based analysis revealed 12 significantly different metabolites, among which most amino acids and derivatives and nitrogen-containing compounds were higher in males, and other compounds were elevated in females. Interestingly, the volcano plot also confirms the similar patterns of amino acids and derivatives higher in males. The age-based analysis suggested that metabolites may undergo substantial alterations during the 25-35-year age range, indicating a potential metabolic turning point associated with the age group. Moreover, a more distinct difference between the 25-35 and above 35 age groups compared to the below 25 and 25-35 age groups was observed, with the most significant compound decreased in the above 35 age groups. CONCLUSION: These findings may contribute to the development of comprehensive metabolomics analyses with confounding factor-based adjustment and enhance the reliability and interpretability of future large-scale investigations.

Gastric coagulation and postprandial amino acid absorption of milk is affected by mineral composition: a randomized crossover trial.

Background: In vitro studies suggest that casein coagulation of milk is influenced by its mineral composition, and may therefore affect the dynamics of protein digestion, gastric emptying and appearance of amino acids (AA) in the blood, but this remains to be confirmed in vivo. Objective: This study aimed to compare gastrointestinal digestion between two milks with the same total calcium content but different casein mineralization (CM). Design: Fifteen males (age 30.9 ± 13.8 years, BMI 22.5 ± 2.2 kg m-2) participated in this randomized cross-over study with two treatments. Participants underwent gastric magnetic resonance imaging (MRI) scans at the baseline and every 10 min up to 90 min after consumption of 600 ml milk with low or high CM. Blood samples were taken at the baseline and up to 5 hours postprandially. Primary outcomes were postprandial plasma AA concentrations and gastric emptying rate. Secondary outcomes were postprandial glucose and insulin levels, gastric coagulation as estimated by image texture metrics, and appetite ratings. Results: Gastric content volume over time was similar for both treatments. However, gastric content image analysis suggested that the liquid fraction emptied quicker in the high CM milk, while the coagulum emptied slower. Relative to high CM, low CM showed earlier appearance of AAs that are more dominant in casein, such as proline (MD 4.18 µmol L-1, 95% CI [2.38-5.98], p < 0.001), while there was no difference in appearance of AAs that are more dominant in whey protein, such as leucine. The image texture metrics homogeneity and busyness differed significantly between treatments (MD 0.007, 95% CI [0.001, 0.012], p = 0.022; MD 0.005, 95% CI [0.001, 0.010], p = 0.012) likely because of a reduced coagulation in the low CM milk. Conclusions: Mineral composition of milk can influence postprandial serum AA kinetics, likely due to differences in coagulation dynamics. The clinical trial registry number is NL8959 (https://clinicaltrials.gov).

Evaluation of different isotope dilution mass spectrometry strategies for the characterization of naturally abundant and isotopically labelled peptide standards.

Natural abundance and isotopically labelled tryptic peptides are routinely employed as standards in quantitative proteomics. The certification of the peptide content is usually carried out by amino acid analysis using isotope dilution mass spectrometry (IDMS) after the acid hydrolysis of the peptide. For the validation and traceability of the amino acid analysis procedure, expensive certified peptides must be employed. In this work we evaluate different IDMS alternatives which will reduce the amount of certified peptide required for validation of the amino acid analysis procedure. In this context, the characterization of both natural and isotopically labelled synthetic angiotensin I peptides was carried out. First, we applied a fast procedure for peptide hydrolysis based on microwave-assisted digestion and employed two certified peptide reference materials SRM 998 angiotensin I and CRM 6901-b C-peptide for validation of the hydrolysis procedure. The amino acids proline, leucine, isoleucine, valine, tyrosine, arginine and phenylalanine were evaluated for their suitability for peptide certification by IDMS by both liquid chromatography with tandem mass spectrometry (LC-MS/MS) and gas chromatography with mass spectrometry (GC)-MS/MS. Then, natural angiotensin I and 13C1-labelled angiotensin I were synthesized in-house and purified by preparative liquid chromatography. The concentration of the 13C1-labelled angiotensin I peptide was established by reverse IDMS in its native form using SRM 998 angiotensin I as reference. The concentration of the natural synthesized peptide was determined by IDMS both using the 13C1-labelled peptide in its native form and by amino acid analysis showing comparable results. Finally, the synthetic naturally abundant angiotensin I peptide was employed as "in-house" standard for the validation of subsequent peptide characterization procedures. Therefore, the novelty of this work relies on, first, the development of a faster hydrolysis procedure assisted by focused microwaves, providing complete hydrolysis in 150 min, and secondly, a validation strategy combining GC-MS and LC-MS/MS that allowed us to certify the purity of an in-house-synthesized peptide standard that can be employed as quality control in further experiments.

Exploring the chemical diversity of Capsicum chinense cultivars using NMR-based metabolomics and machine learning methods.

The habanero pepper (Capsicum chinense) is a prominent spicy fruit integral to the historical, social, cultural, and economic fabric of the Yucatan peninsula in Mexico. This study leverages the power of 1H NMR spectroscopy coupled with machine learning algorithms to dissect the metabolomic profile of eleven C. chinense cultivars, including those grown by INIFAP (Habanero-Jaguar, Antillano-HRA 1-1, Antillano-HRA 7-1, Habanero-HAm-18A, Habanero-HC-23C, and Jolokia-NJolokia-22) and commercial hybrids (Habanero-Rey Votán, Habanero-Kabal, Balam, USAPR10117, and Rey Pakal). A total of fifty metabolites, encompassing sugars, amino acids, short-chain organic acids, and nucleosides, were identified from the 1H NMR spectra. The optimized machine learning model proficiently predicted the similarity percentage between the INIFAP-grown cultivars and commercial hybrids, thereby facilitating a comprehensive comparison. Biomarkers unique to each cultivar were delineated, revealing that the Habanero-Rey Votán cultivar is characterized by the highest concentration of sugars. In contrast, the Balam cultivar is rich in amino acids and short-chain organic acids, sharing a similar metabolomic profile with the Jolokia-NJolokia-22 cultivar. The findings of this study underscore the efficacy and reliability of NMR-based metabolomics as a robust tool for differentiating C. chinense cultivars based on their intricate chemical profiles. This approach not only contributes to the scientific understanding of the metabolomic diversity among habanero peppers but also holds potential implications for food science, agriculture, and the culinary arts.

Exploring the dynamics of bioactive metabolites changes in barley grains (Hordeum vulgare L.) during roasting: Insights from UPLC-QqQ-MS/MS analysis combined to chemometrics.

This investigation delves into the dynamic metabolic shifts within barley grains during the roasting process, employing UPLC-QqQ-MS/MS analysis. The complex spectrum of metabolites before and after roasting is revealed. The resulting data, unveils substantial transformations in chemical composition during roasting. A total of 62 chromatographic peaks spanning phenolic compounds, flavones, Millard Reaction Products, amino acids, lignans, vitamins, folates, and anthocyanins were annotated. Leveraging UPLC-QqQ-MS/MS analysis, we scrutinized the intricate metabolite profile before and after roasting where the roasting process was found to trigger dynamic changes across diverse metabolite classes particularly Millard Reaction Products, produced through the Maillard reaction, with dihydro-5-methyl-5H-cyclopentapyrazine, maltol and hydroxy maltol emerging as discerning markers of roasting progression. Amino acids and sugars showed degradation, while beta-glucan, a signature barley sugar, experienced notable decline. Folate derivatives witnessed pronounced reduction, aligning with the heat sensitivity of folates. Harnessing the power of multivariate data analysis, the consequences of roasting materialize through distinct clusters in PCA and OPLS-DA plots. Noteworthy, roasting duration governs the trajectory of metabolic divergence, culminating in the identification of roasting-specific markers. Epigallocatechin, procyanidin B, 10-HCO-H4 folate, and hordatine A emerge as pivotal discriminators. Orthogonal Projection to Latent Structure (OPLS) analysis linked anti-inflammatory activity with 30-min, 1-hour, and 1.5-hour roasted samples, with hordatine B in addition to some Millard Reaction Products being correlated with pro-inflammatory marker downregulation.. This study encapsulates the intricate metabolic metamorphosis ignited by roasting in barley grains, offering a holistic comprehension of their potential health-enhancing attributes. Key metabolites act as poignant indicators of these transformations, substantiating the complex interplay between roasting and the barley grain metabolome.

Effect of extraction pH on amino acids, nutritional, in-vitro protein digestibility, intermolecular interactions, and functional properties of guar germ proteins.

The chemical compositions, intermolecular interactions, and functional properties of guar germ proteins (GGP) were investigated at different extraction pH (7 to 11). The protein efficiency ratio, essential amino acid index (46.53), predicted biological value (39.02), nutritional index (42.67), and protein purity (91.69 %) were found to be highest at pH 9. The in-vitro protein digestibility of GGP sample was highest at pH 11. From SDS-PAGE, the band intensity (<10 kDa) became thinner with an increase in extraction pH from 7 to 9 and then thicker. Meanwhile, smallest particle size and weaker ionic and hydrogen bonds were found at pH 11. The ß-sheet content was more dominating in GGP samples. Moreover, higher denaturation temperatures of GGP samples indicated that protein molecules had a compact tertiary structure. Furthermore, the GGP extracted at pH 7 showed better functional properties. The principal component analysis suggested that pH 9 was more suitable for isolating GGP.

Using temporal dominance of sensations (TDS), multiple-sip TDS, chemical composition evaluation, and microstructural analysis to assess the impact of repeated reheating cycles on chicken soup.

Reheating chicken soup is a common culinary practice in daily life. To investigate the impact of reheating frequency on the sensory quality of chicken soup, temporal dominance of sensations (TDS) and multi-TDS were used to characterize changes in dominant sensory attributes during consumption. Additionally, E-nose and E-tongue were utilized to analyze differences in aroma and taste profiles. The alterations in sensory properties were further elucidated by analyzing variations in amino acids, volatile compounds. The findings revealed that there was no significant disparity between fresh soup and heating. However, saltiness and umami, as the most prominent dominant characteristics, intensified with increasing reheating cycles. This can be attributed to an elevation in certain amino acids that contribute to umami perception. Conversely, a reduction in some aldehydes weakened the flavor associated with fat and meat components. Moreover, enlarged oil droplets with uneven distribution within heated soup may account for the heightened greasiness sensation.

Three-Minute Enantioselective Amino Acid Analysis by Ultra-High-Performance Liquid Chromatography Drift Tube Ion Mobility-Mass Spectrometry Using a Chiral Core-Shell Tandem Column Approach.

Fast liquid chromatography (LC) amino acid enantiomer separation of 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) derivatives using a chiral core-shell particle tandem column with weak anion exchange and zwitterionic-type quinine carbamate selectors in less than 3 min was achieved. Enantiomers of all AQC-derivatized proteinogenic amino acids and some isomeric ones (24 in total plus achiral glycine) were baseline separated (Rs > 1.5 except for glutamic acid with Rs = 1.3), while peaks of distinct amino acids and structural isomers (constitutional isomers and diastereomers of leucine and threonine) of the same configuration overlapped to various degrees. For this reason, drift tube ion mobility-mass spectrometry was added (i.e., LC-IM-MS) as an additional selectivity filter without extending run time. The IM separation dimension in combination with high-resolution demultiplexing enabled confirmation of threonine isomers (threonine, allo-threonine, homoserine), while leucine, isoleucine, and allo-isoleucine have almost identical collisional cross-section (DTCCSN2) values and added no selectivity to the partial LC separation. Density functional theory (DFT) calculations show that IM separation of threonine isomers was possible due to conformational stabilization by hydrogen bond formation between the hydroxyl side chain and the urea group. Generally, the CCSN2 of protonated ions increased uniformly with addition of the AQC label, while outliers could be explained by consideration of intramolecular interactions and additional structural analysis. Preliminary validation of the enantioselective LC-IM-MS method for quantitative analysis showed compliance of accuracy and precision with common limits in bioanalytical methods, and applicability to a natural lipopeptide and a therapeutic synthetic peptide could be demonstrated.

Changes in the chemical properties and metabolite profiling of fish sauce prepared from underutilized large yellow croaker roes during fermentation at different temperatures.

Fish sauce is a popular aquatic condiment with unique flavor. In this study, the changes in the chemical properties and metabolite profiling of fish sauce from large yellow croaker roes during fermentation at different temperatures were revealed. The results found that the contents of total acid, amino acid nitrogen, total soluble nitrogen and soluble salt-free solids of fish sauce fermented at 40 °C were higher than those in other temperatures groups (25 °C and 32 °C), while the contents of total volatile basic nitrogen were lower than other temperatures. Therefore, 40 °C was the ideal fermentation temperature for fish sauce. The metabolomics analysis showed that organic acids, amino acids, nucleotide, and lipid compounds were found to participate in the biosynthesis pathway. Compared to 25 °C and 32 °C, fermented at 40 °C could increase the abundance of metabolic substances in the fish sauce, such as sugar alcohols, L-Citrulline, L-Aspartic acid, L-Cysteine, Glutathione, and L-Arginine. These results provide a theoretical basis for the production of high-quality fish sauce and the high-value utilization of fish roes.

Impact of sulfite use and acidification on chemical quality components in thermally processed watermelon juices.

The present study compared pasteurized and reconstituted (from vacuum-concentrated) watermelon juices with sulfite use (∼40 mg/L) and acidification (pH = 4.2) to fresh watermelon juices. The products were evaluated for phenolics, free amino acids, carotenoids, sugars, organic acids, and alcohols by high-performance liquid chromatography-HPLC and the volatile profile by headspace-gas chromatography/mass spectrometry(HS-GC/MS). Pasteurization had no significant impact on most of the chemical components. Furthermore, it potentiated typical watermelon aromas (E,E)-2,6-nonadienal, (Z)-3-nonen-1-ol, 4-hexen-1-ol, (E,Z)-3,6-nonadien-1-ol, 6-amino-2-methyl-2-heptanol, (E)-6-nonenal, (E)-2-nonenal, pentanal, nonanal and 1-nonanol), without off-flavor compounds formation. On the other hand, the reconstituted juice showed reduced amino acids (serine, glutamine, and tryptophan), phenolics (epicatechin gallate, myricetin, and cis-resveratrol), carotenoids (lycopene, ß-carotene, and violaxanthin) and most volatile compounds. Our results showed that sulfite and acidification could maintain watermelon juice's nutritional and quality parameters after pasteurization. The vacuum concentration and reconstitution processes negatively impacted the evaluated compounds. Our findings contribute to improving thermal processes in watermelon juices for better preservation of nutrients, flavor, and bioactive compounds.

Toward multidimensional information: A derivatization-free UHPLC-QqQ MS/MS method for amino acid components of fingerprint.

The analysis of fingerprint chemical composition is a meaningful way to excavate the multidimensional information of fingerprint, including the donor profiling information and the age of a fingerprint, which broadens the evidential values of fingerprint, especially for the partial and distorted fingerprint. But the research remains still in the pilot phases or is ongoing. Amino acids are the dominant organic substances in latent sweat fingerprint and influenced by many donor factors. Hence, their content reflects personal information of donors. Forensic science will be revolutionized if suspects can be individualized by their amino acid content. The diverse nature, distinct physicochemical properties, and ultra-micro levels of amino acids present in fingerprints make it hard to detect. A high sensitivity method for detecting and quantifying multiple amino acid components is required. UHPLC-QqQ MS/MS offers high sensitivity, high separation, simultaneous multicomponents detection, and no derivatization, making it an ideal method for detecting and analyzing amino acids in fingerprints. Therefore, in this study, we propose and validate an efficient UHPLC-QqQ MS/MS method for the extraction and analysis of 13 amino acids from fingerprint. We compared the results of amino acids of 10 different substrates and found that the inherent amino acids in most porous substrates would have been extracted along with the fingerprint amino acids, making them unsuitable for quantitative amino acid analysis. Instead, plastic sheets are ideal substrates for laboratory studies. Then, extensive experiments were conducted among 30 donors for multidimensional information analysis. The type of samples analyzed were eccrine-rich fingerprints. A Binary Logistic Regression (BLR) model was developed, and the female and male donors were successfully differentiated by amino acids in fingerprints. Two other mathematical models were also developed to verify the accuracy, and all three different mathematical models were able to identify donors of different genders with over 90% accuracy. This demonstrates that amino acids have the potential to provide more information for donors as metabolic markers. In the future, we will conduct a series of experiments to analyze more multidimensional information for individual identification by amino acid content in the fingerprint.

Self-Assembling Polypeptides in Complex Coacervation.

ConspectusIntracellular compartmentalization plays a pivotal role in cellular function, with membrane-bound organelles and membrane-less biomolecular "condensates" playing key roles. These condensates, formed through liquid-liquid phase separation (LLPS), enable selective compartmentalization without the barrier of a lipid bilayer, thereby facilitating rapid formation and dissolution in response to stimuli. Intrinsically disordered proteins (IDPs) or proteins with intrinsically disordered regions (IDRs), which are often rich in charged and polar amino acid sequences, scaffold many condensates, often in conjunction with RNA.Comprehending the impact of IDP/IDR sequences on phase separation poses a challenge due to the extensive chemical diversity resulting from the myriad amino acids and post-translational modifications. To tackle this hurdle, one approach has been to investigate LLPS in simplified polypeptide systems, which offer a narrower scope within the chemical space for exploration. This strategy is supported by studies that have demonstrated how IDP function can largely be understood based on general chemical features, such as clusters or patterns of charged amino acids, rather than residue-level effects, and the ways in which these kinds of motifs give rise to an ensemble of conformations.Our laboratory has utilized complex coacervates assembled from oppositely charged polypeptides as a simplified material analogue to the complexity of liquid-liquid phase separated biological condensates. Complex coacervation is an associative LLPS that occurs due to the electrostatic complexation of oppositely charged macro-ions. This process is believed to be driven by the entropic gains resulting from the release of bound counterions and the reorganization of water upon complex formation. Apart from their direct applicability to IDPs, polypeptides also serve as excellent model polymers for investigating molecular interactions due to the wide range of available side-chain functionalities and the capacity to finely regulate their sequence, thus enabling precise control over interactions with guest molecules.Here, we discuss fundamental studies examining how charge patterning, hydrophobicity, chirality, and architecture affect the phase separation of polypeptide-based complex coacervates. These efforts have leveraged a combination of experimental and computational approaches that provide insight into molecular level interactions. We also examine how these parameters affect the ability of complex coacervates to incorporate globular proteins and viruses. These efforts couple directly with our fundamental studies into coacervate formation, as such "guest" molecules should not be considered as experiencing simple encapsulation and are instead active participants in the electrostatic assembly of coacervate materials. Interestingly, we observed trends in the incorporation of proteins and viruses into coacervates formed using different chain length polypeptides that are not well explained by simple electrostatic arguments and may be the result of more complex interactions between globular and polymeric species. Additionally, we describe experimental evidence supporting the potential for complex coacervates to improve the thermal stability of embedded biomolecules, such as viral vaccines.Ultimately, peptide-based coacervates have the potential to help unravel the physics behind biological condensates, while paving the way for innovative methods in compartmentalization, purification, and biomolecule stabilization. These advancements could have implications spanning medicine to biocatalysis.

Adding pulse flours to cereal-based snacks and bakery products: An overview of free asparagine quantification methods and mitigation strategies of acrylamide formation in foods.

Thermal processing techniques can lead to the formation of heat-induced toxic substances. Acrylamide is one contaminant that has received much scientific attention in recent years, and it is formed essentially during the Maillard reaction when foods rich in carbohydrates, particularly reducing sugars (glucose, fructose), and certain free amino acids, especially asparagine (ASN), are processed at high temperatures (>120°C). The highly variable free ASN concentration in raw materials makes it challenging for food businesses to keep acrylamide content below the European Commission benchmark levels, while avoiding flavor, color, and texture impacts on their products. Free ASN concentrations in crops are affected by environment, genotype, and soil fertilization, which can also influence protein content and amino acid composition. This review aims to provide an overview of free ASN and acrylamide quantification methods and mitigation strategies for acrylamide formation in foods, focusing on adding pulse flours to cereal-based snacks and bakery products. Overall, this review emphasizes the importance of these mitigation strategies in minimizing acrylamide formation in plant-based products and ensuring safer and healthier food options.

Impact of prolonged storage on quality assessment properties and constituents of honey: A systematic review.

This systematic review paper aims to discuss the trend in quality assessment properties and constituents of honey at different storage conditions and confer the possible whys and wherefores associated with the significant changes. Initially, a literature search was conducted through Google Scholar, ScienceDirect, PubMed, and Scopus databases. In total, 43 manuscripts published between 2001 and 2023 that met the inclusion and exclusion criteria were chosen for the review. As an outcome of this review, prolonged honey storage could deteriorate sensory, nutritional, and antioxidant properties and promote fermentation, granulation, microbial growth, carcinogenicity, organotoxicity, and nephrotoxicity. This systematic review also recognized that diastase activity, invertase activity, 5-hydroxymethylfurfural content, proline content, sugar content, amino acids, and vitamins could be used as indicators to distinguish fresh and stored honey based on the significant test (p-value) in the reported studies. However, all the reported studies used the simplest approach (one-way ANOVA) to identify the significant differences in the analyzed parameter during the storage period and none of them reported an approach to identify the most influential parameter at different storage conditions. In conclusion, orthogonal partial least squares discriminant analysis (supervised multivariate statistical tool) has to be employed in future studies to find the most influential parameter and could be used to potent chemical markers to distinguish fresh and stored honey because this analysis is incorporated with S-plot, variable importance of projection, and one-way ANOVA, which can produce the most accurate and precise results rather solely depending on one-way ANOVA.