L-lactic Acid Content Research Articles

Substances Secreted by Lactobacillus spp. from the Urinary Tract Microbiota Play a Protective Role against Proteus mirabilis Infections and Their Complications.

Proteus mirabilis urinary tract infections can lead to serious complications such as development of urinary stones. Lactobacillus spp., belonging to the natural microbiota of the urinary tract, exhibit a number of antagonistic mechanisms against uropathogens, including the secretion of organic acids. In this study, we determined the anti-adhesion, anti-cytotoxicity and anti-crystallization properties of the substances secreted by Lactobacillus. For this purpose, membrane inserts with a pore diameter 0.4 μm were used, which prevent mixing of cultured cells, simultaneously enabling the diffusion of metabolic products. The intensity of crystallization was assessed by measuring the levels of Ca2+, Mg2+ and NH3 and by observing crystals using microscopic methods. The cytotoxicity of the HCV-29 cell line was determined using the LDH and MTT assays, and the impact of lactobacilli on P. mirabilis adhesion to the bladder epithelium was assessed by establishing CFU/mL after cell lysis. It was shown that in the presence of L. gasseri the adhesion of P. mirabilis and the cytotoxicity of the cells decreased. The degree of crystallization was also inhibited in all experimental models. Moreover, it was demonstrated that L. gasseri is characterized by the secretion of a high concentration of L-lactic acid. These results indicate that L-lactic acid secreted by L. gasseri has a significant impact on the crystallization process and pathogenicity of P. mirabilis.

Fermentative L-Lactic Acid Production Using Bacillus coagulans from Corn Stalk Deconstructed by an Anaerobic Microbial Community

This study investigated the feasibility of producing L-lactic acid (LA) from dry corn stalk (DCS) that was pretreated by ensiling by an anaerobic microbial community consisting of Bacillus coagulans, Lactobacillus fermentum, and Enterococcus durans. After 28 days of ensiling, the LA and acetic acid content in the microsilage was 2.04 ± 0.08% and 0.38 ± 0.01%, respectively, and the pH was 4.47 ± 0.13. Enterococcus and Lactobacillus became the dominant microbiota during the ensiling process. Twenty-eight-day-old microsilage was then subjected to fermentation by B. coagulans to produce LA in a simultaneous saccharification and co-fermentation process. The enzymatic hydrolysis yield reached >96%. The maximal concentration of LA reached 18.54 ± 0.52 g/L with a substrate concentration of 5%, where the yield of LA was 0.31 ± 0.01 g/g DCS and the optical purity of the product LA was >97%. Anaerobic ensiling is viable for the pretreatment of biomass for the production of value-added chemicals.

Exploration on Solvatochromic Sensing Probe of Phenol Blue for L‐Lactic Acid Detection: Effective Strategy to Obtain Reliable Capacitive Sensor

AbstractThis study systematically investigates a capacitive sensor applied with phenol blue (PhB)‐based sensing medium for detection of L‐lactic acid (LA), as a health monitoring indicator. PhB is a substance with solvatochromic effect, inducing the change in capacitance by exposure to polar molecules. However, the capacitive LA sensor with a flat‐structured PhB/polyvinylchloride (PVC) composite‐sensing medium is observed to have a problem in that sensing capacitance variation saturate quickly with increasing the LA solution concentration. This main cause can be analyzed that the interaction of proton from LA molecule with the lone pair electrons of the PhB molecule acts as a major factor on the sensing characteristics rather than the solvatochromic behavior of PhB molecule. Therefore, a strategy is adopted to introduce a porous structure to the PhB/PVC composite‐sensing medium to maximize the interaction of PhB with protons, which is implemented through solvent and non‐solvent exchange methods. Consequently, the sensitivity and linearity of the porous‐structured LA sensor are 2.99 pF mm−1 and 0.966 over LA concentrations ranging from 0 to 100 mm, respectively, which is a significant improvement over that of the flat‐structured one. Notably, the sensing performance remained unchanged even after a month of storage under normal ambient conditions.

Distribution and Quantification of Lactic Acid Enantiomers in Baijiu.

Enantiomers of lactic acid were investigated in Baijiu, including soy sauce aroma-type Baijiu (SSB), strong aroma-type Baijiu (STB), and light aroma-type Baijiu (LTB), via high-performance liquid chromatography with a chiral separation column. The natural concentration and enantiomeric distribution of lactic acid were studied, and their contribution to the flavor of Chinese Baijiu was evaluated based on recognition threshold. The results showed that there were significant differences in the content of lactic acid and the ratio of enantiomeric isomers among different aroma types and storage year. In SSB, the concentrations of D-lactic acid and L-lactic acid were higher, with the highest concentrations of 1985.58 ± 11.34 mg/L and 975.31 ± 14.03 mg/L, respectively. In STB, the highest concentrations of D-lactic acid and L-lactic acid were 1048.00 ± 11.46 mg/L and 939.83 ± 0.23 mg/L, respectively. In LTB, the highest concentrations of D-lactic acid and L-lactic acid were 760.90 ± 9.45 mg/L and 558.33 ± 3.06 mg/L, respectively. The average D/L enantiomeric ratios were 78:22 ± 16.16 and 80:20 ± 9.72 in the Commercial Baijiu products of SSB and STB, respectively. The average D/L enantiomeric ratio in LTB was 90:10 ± 6.08. D-lactic acid in JSHS vintage Baijiu showed a wave variation with aging, while L-lactic acid gradually increased during aging, and the average D/L enantiomeric ratio was 76:24 ± 4.26. The concentration of D-lactic acid in XJCT vintage Baijiu also showed a wave variation with aging, and the concentration of L-lactic acid tended to be stable during aging, with an average D/L enantiomeric ratio of 88:12 ± 2.80. The content of the two configurations of lactic acid in the LZLJ vintage Baijiu showed a decreasing trend during aging, with an average D/L enantiomeric ratio of 60:40 ± 11.99. The recognition threshold of D-lactic acid in 46% ethanol solution was 194.18 mg/L with sour taste; while the L-lactic acid was 98.19 mg/L with sour taste. The recognition threshold of L-lactic acid was about half that of D-lactic acid, indicating that L-lactic acid has a stronger sour taste. The taste activity values (TAVs) of D-lactic acid and L-lactic acid were greater than 1 in most of the Baijiu samples, and the TAV of D-lactic acid was greater than that of L-lactic acid. The study showed that the lactic acid enantiomers contributed to the taste perception of Baijiu in most of the samples, and D-lactic acid contributed more to the Baijiu taste than L-lactic acid.

Multi-omic Analysis of the Gut Microbiome in Rats with Lithium-Pilocarpine-Induced Temporal Lobe Epilepsy.

Evidence supports that the gut microbiota and bacteria-dependent metabolites influence the maintenance of epileptic brain activity. However, the alterations in the gut microbiota between epileptic versus healthy individuals are poorly understood. We used a multi-omic approach to evaluate the changes in the composition of gut metagenome as well in the fecal metabolomic profile in rats before and after being submitted to status epilepticus (SE)-induced temporal lobe epilepsy (TLE). The 16S ribosomal RNA (rRNA) sequencing of fecal samples coupled to bioinformatic analysis revealed taxonomic, compositional, and functional shifts in epileptic rats. The species richness (Chao1 index) was significantly lower in the post-TLE group, and the β-diversity analysis revealed clustering separated from the pre-TLE group. The taxonomic abundance analysis showed a significant increase of phylum Desulfobacterota and a decrease of Patescibacteria in the post-TLE group. The DESEq2 and LEfSe analysis resulted in 18 genera significantly enriched between post-TLE and pre-TLE groups at the genus level. We observed that epileptic rats present a peculiar metabolic phenotype, including a lower concentration of D-glucose and L-lactic acid and a higher concentration of L-glutamic acid and glycine. The microbiota-host metabolic correlation analysis showed that the genera differentially abundant in post-TLE rats are associated with the altered metabolites, especially the proinflammatory Desulfovibrio and Marvinbryantia, which were enriched in epileptic animals and positively correlated with these excitatory neurotransmitters and carbohydrate metabolites. Therefore, our data revealed a correlation between dysbacteriosis in epileptic animals and fecal metabolites that are known to be relevant for maintaining epileptic brain activity by enhancing chronic inflammation, an excitatory-inhibitory imbalance, and/or a metabolic disturbance. These data are promising and suggest that targeting the gut microbiota could provide a novel avenue for preventing and treating acquired epilepsy. However, the causal relationship between these microbial/metabolite components and the SRS occurrence still needs further exploration.

Formation mechanisms of ethyl acetate and organic acids in Kluyveromyces marxianus L1-1 in Chinese acid rice soup

This study aims to explore the formation mechanism of ethyl acetate and organic acids in acid rice soup (rice-acid soup) inoculated with Kluyveromyces marxianus L1-1 through the complementary analysis of transcriptome and proteome. The quantity of K. marxianus L1-1 varied significantly in the fermentation process of rice-acid soup and the first and third days were the two key turning points in the growth phase of K. marxianus L1-1. Importantly, the concentrations of ethyl acetate, ethanol, acetic acid, and L-lactic acid increased from day 1 to day 3. At least 4231 genes and 2937 proteins were identified and 610 differentially expressed proteins were annotated to 30 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways based on the analysis results of transcriptome and proteome. The key genes and proteins including up-regulated alcohol dehydrogenase family, alcohol O-acetyltransferase, acetyl-CoA C-acetyltransferase, acyl-coenzyme A thioester hydrolase, and down-regulated aldehyde dehydrogenase family were involved in glycolysis/gluconeogenesis pathways, starch and sucrose metabolism pathways, amino sugar and nucleotide sugar metabolism pathways, tricarboxylic acid (TCA) cycle, and pyruvate metabolism pathways, thus promoting the formation of ethyl acetate, organic acids, alcohols, and other esters. Our results revealed the formation mechanisms of ethyl acetate and organic acids in rice-acid soup inoculated with K. marxianus L1-1.

Enhanced lactic acid production with indigenous microbiota from date pulp waste and keratin protein hydrolysate from chicken feather waste

A substantial amount of keratin, a key protein with excellent material-forming properties, can be sustainably extracted from chicken feathers, which are commonly discarded in the poultry industry. The keratin protein found in chicken feathers can serve as a nitrogen source and a pH regulator for microbes. In this study, the use of indigenous bacteria and extracted keratin protein hydrolysate from chicken feathers as a nitrogen source improved lactic acid production from date pulp waste by fermentation. After six days of optimal dark fermentation time at pH 6, the maximum concentration of L-lactic acid (46.2 g/L) was obtained with keratin protein hydrolysate and date pulp waste substrate. Under the same conditions, the value was 2.12 times higher than that of the date pulp waste substrate (21.7 g/L) without the addition of keratin protein hydrolysate. Under optimal conditions, volumetric lactic acid productivity and yield were 7.7 g/L/day and 0.3 g/g substrate, respectively. Keratin protein hydrolysate proved to be the most effective neutralizing agent for L-lactic acid production at a concentration of 6 g/L. The combination of keratin protein hydrolysate and date pulp waste was studied for the first time as a substrate for lactic acid production. Keratin protein hydrolysate was found to buffer pH changes during fermentation. Poultry and plant by-products are usually available in agriculturally challenged areas. Therefore, this study provides an excellent opportunity to recycle waste and improve the overall circular economy of such agro-industrial complexes.

Photosensitized oxidation of cholesterol and altered oxysterol levels in sour cream: Effects of addition of cucumber pickles

The aim of this research was to examine whether sour cream (18% fat) is an exogenous source of oxysterols and whether it is possible to improve its antioxidative properties and to modulate cholesterol transformation by adding cucumber. To determine whether cucumber modifies the properties of sour cream, fresh cucumber or cucumber pickle (pH 3.3; 1.5% lactic acid) was added in an amount of 20%. The sour cream samples were then stored under light (450 lx, 590 cd, 120 lm) for 3 wk. After storage, the addition of the cucumber pickle increased total mesophilic aerobic bacteria from 7.5 to 9.3 log cfu/g and increased the l-lactic acid content from 6.1 to 9.7 g/L. The total conjugated linoleic acid content in sour cream with cucumber pickle also increased to 4.5 mg/g fat after storage, whereas the cholesterol content decreased to 3.44 g/kg fat. Importantly, with the addition of cucumber pickle, the total content of cholesterol oxidization products (COP) did not change after storage (1.7 mg/kg fat). By contrast, the total COP content in the control sour cream sample increased from 1.7 to 7.3 mg/kg fat over 3 wk of storage. The dominant COP before and after storage was 7β-hydroxycholesterol. Thus, despite exposure to light, adding cucumber pickle to sour cream modulates cholesterol transformation and effectively inhibits the formation of oxysterols.

Flow-induced crystallization of polylactides

In this work, the flow-induced crystallization of two polylactides (PLAs) with different microstructures (different l-lactic acid contents) is studied using simple shear, uniaxial extension and capillary flow experiments. In a simple shear and capillary flow, an increase in shear rate and a decrease in temperature were found to enhance the crystallization kinetics particularly for Weissenberg numbers (based on the reptation relaxation time, Wi) greater than 1 (strong flow causing chain stretching). On the other hand, in a uniaxial extensional flow, once a critical Hencky strain is achieved, crystallization starts independently of strain rate and temperature. The amount of mechanical work per unit volume imposed/dissipated onto the polymers during flow to initialize crystallization was also calculated in the simple shear, capillary, and extensional flow. The critical mechanical work for the onset of flow-induced crystallization was found to be independent of temperature and degree of molecular chain stretch (Wi) as Wi becomes greater than 1. The critical mechanical work for the onset of flow-induced crystallization in an extensional flow was found to be much smaller than that in a shear flow. The PLA sample with higher content of PLLA showed slightly higher zero-shear viscosity and a smaller thermodynamic barrier for the onset of crystallization. Finally, the degree of crystallinity increases linearly from 0% at the start of the flow-induced crystallization region and reaches a plateau at Wi equals to around 1.

Comparative measurement of D- and L-lactic acid isomers in vaginal secretions: association with high-grade cervical squamous intraepithelial lesions

Identification of low-cost protocols to identify women at elevated susceptibility to develop cervical intra-epithelial abnormalities would aid in more individualized monitoring. We evaluated whether quantitation of the D- and L-lactic acid isomers in vaginal secretions could differentiate women with normal cervical epithelia from those with a low (LSIL) or high (HSIL) grade squamous intraepithelial lesion or with cervical cancer. Vaginal samples, collected from 78 women undetgoing cervical colposcopy and biopsy, were tested for pH, bacterial composition by Gram stain (Nugent score) and concentrations of D- and L-lactic acid by a colorimetric assay. Subsequent diagnosis was 23 women with normal cervical epithelium, 10 with LSIL, 43 with HSIL and 2 with cervical cancer. Vaginal pH and Nugent score were comparable in all subject groups. The concentration of L-lactic acid, but not D-lactic acid, as well as the L/D-lactic acid ratio, were significantly elevated (p < 0.01) in women with HSIL and cervical cancer. Comparative measurement of vaginal D- and L-lactic acid isomers may provide a low-cost alternative to identification of women with an elevated susceptibility to cervical abnormalities.

Production of L (+) Lactic Acid by Lactobacillus casei Ke11: Fed Batch Fermentation Strategies

Lactic acid and its derivatives are widely used in pharmaceutical, leather, textile and food industries. However, until now there have been few systematic reports on fed-batch fermentation for efficient production and high concentration of l-lactic acid by lactic acid bacteria. This study describes the obtainment of L (+) lactic acid from sucrose using the Lactobacillus casei Ke11 strain through different feeding strategies using an accessible pH neutralizer such as CaCO3. The exponential feeding strategy can increase lactic acid production and productivity (175.84 g/L and 3.74 g/L/h, respectively) with a 95% yield, avoiding inhibition by high initial substrate concentration and, combined with the selected agent controller, avoids the cellular stress that could be caused by the high osmotic pressure of the culture media. The purification of the acid using charcoal and celite, followed by the use of a cation exchange column proved to be highly efficient, allowing a high yield of lactic acid, high removal of sugars and proteins. The described process shows great potential for the production of lactic acid, as well as the simple, efficient and low-cost purification method. This way, this work is useful to the large-scale fermentation of L. casei Ke11 for production of l-lactic acid.

A Graphene-Based Enzymatic Biosensor Using a Common-Gate Field-Effect Transistor for L-Lactic Acid Detection in Blood Plasma Samples.

Lactate is an important organic molecule that is produced in excess during anaerobic metabolism when oxygen is absent in the human organism. The concentration of this substance in the body can be related to several medical conditions, such as hemorrhage, respiratory failure, and ischemia. Herein, we describe a graphene-based lactate biosensor to detect the concentrations of L-lactic acid in different fluids (buffer solution and plasma). The active surface (graphene) of the device was functionalized with lactate dehydrogenase enzyme using different substances (Nafion, chitosan, and glutaraldehyde) to guarantee stability and increase selectivity. The devices presented linear responses for the concentration ranges tested in the different fluids. An interference study was performed using ascorbic acid, uric acid, and glucose, and there was a minimum variation in the Dirac point voltage during detection of lactate in any of the samples. The stability of the devices was verified at up to 50 days while kept in a dry box at room temperature, and device operation was stable until 12 days. This study demonstrated graphene performance to monitor L-lactic acid production in human samples, indicating that this material can be implemented in more simple and low-cost devices, such as flexible sensors, for point-of-care applications.

An exploratory study of associations with spontaneous preterm birth in primigravid pregnant women with a normal cervical length

Background Predictors of spontaneous preterm birth in primigravid women remain undetermined. Aim We evaluated whether biomarkers in vaginal secretions and/or differences in the dominant bacterium in the vaginal microbiome predicted the risk for spontaneous preterm birth in primigravid women with a cervical length >25mm. Study design In a prospective study, 146 second trimester pregnant women with their first conception and a cervix >25mm were enrolled. The vaginal microbiome composition was characterized by analysis of 16S ribosomal RNA gene sequences. The concentrations of d- and l-lactic acid, matrix metalloproteinase (MMP) 2, 8 and 9 and tissue inhibitor of metalloproteinase (TIMP) 1 and 2 in vaginal secretions were measured by ELISA. Cervical length was determined by vaginal ultrasonography. Pregnancy outcome data were subsequently collected. There was a spontaneous preterm birth (SPTB) in 13 women (8.9%) while in an additional 8 women (5.5%) preterm delivery was medically indicated. Lactobacillus iners was the dominant vaginal bacterium in 61.5% of women with a SPTB but only in 31.2% of those who delivered at term (p = .0354). The vaginal concentration of TIMP-1 (p = .0419) and L-lactic acid (p = .0495) was higher in women with a SPTB as compared to those who delivered at term. Lactobacillus iners dominance was associated with elevated levels of TIMP-1 (p = .0434) and TIMP-2 (p = .0161) and lower levels of D-lactic acid (p < .0001) compared to when L. crispatus was dominant. Conclusion In this exploratory study of primigravid women, elevations in vaginal TIMP-1 and L- lactic acid and L. iners dominance in the vaginal microbiome are associated with an increased occurrence of SPTB.

Investigating the microbiota of fermented fish products (Pla-ra) from different communities of northeastern Thailand

DNA-sequencing was performed on the V3-V4 regions of 16S rRNA genes to investigate the microbial diversity of five samples of fermented freshwater fish (pla-ra) from three provinces in northeastern Thailand. The samples had salt concentrations ranging from 7 to 10%, pH values from 4.83 to 7.15, and D-/L-lactic acid concentrations of 90 to 450 mg/l. A total of 598 operational taxonomic units were annotated at various taxonomic ranks based on the SILVA Database. The lactic-acid and halophilic genera Tetragenococcus, Halanaerobium and Lactobacillus were among the dominant taxa of bacteria. The top 20 non-redundant taxa were considered in more detail. In two pla-ra samples, Tetragenococcus muriaticus was commonly identified. Halanaerobium fermentans was the most abundant species in a third sample and co-dominant in another sample. Lactobacillus rennini was dominant in the pla-ra sample from Roi Et Province. Additionally, other beneficial bacteria were detected including Staphylococcus nepalensis, Lactobacillus sakei, Lactobacillus pentosus, Weissella confusa, and Bifidobacterium bifidum. Differences between samples may be due to use of different raw materials, salt concentrations, recipes, processes and fermentation periods. The microbial communities in pla-ra provide a better understanding of the production outcomes of traditional products. Further optimization of the fermentation process, for example by using dominant bacterial taxa in starter cultures, may improve processes of food fermentation, food quality and flavor control, providing useful guidelines for industrial applications.

Effects and mechanism of Lactococcus lactis thermo-sensitive hydrogel on the wound healing of full-thickness skin defects in diabetic mice

Effects and mechanism of Lactococcus lactis thermo-sensitive hydrogel on the wound healing of full-thickness skin defects in diabetic mice

Chemical Composition of Sour Beer Resulting from Supplementation the Fermentation Medium with Magnesium and Zinc Ions.

The bioavailability of minerals, such as zinc and magnesium, has a significant impact on the fermentation process. These metal ions are known to influence the growth and metabolic activity of yeast, but there are few reports on their effects on lactic acid bacteria (LAB) metabolism during sour brewing. This study aimed to evaluate the influence of magnesium and zinc ions on the metabolism of Lactobacillus brevis WLP672 during the fermentation of brewers’ wort. We carried out lactic acid fermentations using wort with different mineral compositions: without supplementation; supplemented with magnesium at 60 mg/L and 120 mg/L; and supplemented with zinc at 0.4 mg/L and 2 mg/L. The concentration of organic acids, pH of the wort and carbohydrate use was determined during fermentation, while aroma compounds, real extract and ethanol were measured after the mixed fermentation. The addition of magnesium ions resulted in the pH of the fermenting wort decreasing more quickly, an increase in the level of L-lactic acid (after 48 h of fermentation) and increased concentrations of some volatile compounds. While zinc supplementation had a negative impact on the L. brevis strain, resulting in a decrease in the L-lactic acid content and a higher pH in the beer. We conclude that zinc supplementation is not recommended in sour beer production using L. brevis WLP672.

Revealing metabolic pathways relevant to prediabetes based on metabolomics profiling analysis

AimsDahl salt-sensitive (SS) rats develop similar prediabetes lesion characteristics, such as impaired glucose tolerance (IGT), when compared with the salt resistant rat. In this study, we evaluate the risk of high glucose intake during prediabetes and reveal the metabolic pathways relevant to the pathophysiology of prediabetes to diabetes using the SS rat model and compared this with the salt-resistant consomic SS.13BN rat model. MethodsSS rats were fed with normal chow ±10% glucose solution ad libitum for five weeks. The same experimental treatment was performed on the SS.13BN rats. Metabolites derived from the serum and liver tissue were measured through biochemical and metabolomics analyses. Multivariate, pathway enrichment, and metabolic correlation network analyses were performed based on the metabolomics data. ResultsBiochemical analysis revealed that serum triglyceride (TG) significantly increased with a significant decrease in serum total cholesterol (TC) after high glucose intake in the SS rat. Metabolic pathway analysis revealed that high glucose intake interfered with galactose, glyoxylate, and dicarboxylate metabolism, most evidently in the SS rat. Hepatic l-lactic acid content increased in the SS rat after high glucose intake, whereas the opposite was observed in SS.13BN rats. Metabolic correlation network analysis based on serum metabolites revealed that urea and l-valine had higher metabolic centrality in the SS rat. ConclusionOur findings revealed that high glucose intake can significantly stimulate hypertriglyceridemia and reduce serum TC level. The profoundly altered metabolic pathway included galactose, glyoxylate, and dicarboxylate metabolism. l-lactic acid was screened as a biomarker in liver, whereas l-valine and urea were screened as hub metabolites in serum.

Comparison of Lactic Acid Production by L. casei in Batch, Fed-batch and Continuous Cultivation, Testing the use of Feather Hydrolysate as a Complex Nitrogen Source

Abstract A comprehensive comparison of the main fermentation parameters, productivity, yield and final L-lactic acid concentration, obtained through batch, fed-batch and continuous cultivations using Lactobacillus casei CCDM 198 and a model cultivation medium was carried out. Using this data, a pulse-feed fed-batch process was established for testing chicken feather hydrolysate as a replacement for all complex nitrogen sources (yeast and beef extracts and peptone) in the medium. As comparably high values of productivity (about 4.0 g/L/h) and yield (about 98 %) were reached under all cultivation conditions, the maximum final L-lactic acid concentration (116.5 g/L), as achieved through pulse-feed fed-batch fermentation, was chosen as the most important criterion for process selection. Fed-batch cultivation with chicken feather hydrolysate as both a complex nitrogen source and a neutralizing agent for maintaining constant culture pH yielded half the concentration of L-lactic acid compared to the model medium. We demonstrate here that chicken feather hydrolysate has potential for use in the production of L-lactic acid but its utilization requires further optimization

Analysis of hypoxanthine and lactic acid levels in vitreous humor for the estimation of post-mortem interval (PMI) using LC–MS/MS

Hypoxanthine (Hx) is produced during terminal stages of purine catabolism in humans. The concentrations of Hx and l-lactic acid in vitreous humor highly correlate with post-mortem interval (PMI). In this study, we measured the concentrations of Hx and l-lactic acid in uncontrolled authentic vitreous humor from cadavers, and investigated the correlation between these molecules and PMI. A liquid chromatography–tandem mass spectrometry (LC–MS/MS) was used for the quantitative analysis of Hx and l-lactic acid in vitreous humor. These concentrations were also corrected by four seasons such as spring, summer, autumn, and winter and temperatures on the day of death such as −10 to 5 °C, 6–15 °C, 16–25 °C, and 26–32 °C that may affect the biomarker concentrations. Vitreous humors were collected from cadavers with known time of death at the National Forensic Service (NFS), Republic of Korea. The correlation between the concentrations of Hx and l-lactic acid in vitreous humors and PMI was evaluated using 79 corpses (53 males and 26 females), with sampling time ranging from 13 to 103 h after death. The average daily ambient temperature at the time of death of each sample was investigated to calibrate the correlation with PMI. Following correction of the concentrations of Hx and l-lactic acid with temperature, the correlation of Hx and l-lactic acid with PMI increased from 0.53 to 0.59 for Hx and 0.38 to 0.42 for l-lactic acid. The highest correlation of Hx and l-lactic acid concentrations with PMI was observed in the winter season, with an R2 value of 0.80 and 0.71 for Hx and l-lactic acid, respectively. The correlation of Hx and l-lactic acid with PMI was corrected by ambient temperature for each season, resulting in an increase in the R2 value to 0.88 for Hx and 0.72 for l-lactic acid. The best correlation was observed when the temperature was corrected after dividing by season.

Structure–property relation of porous poly (l-lactic acid) scaffolds fabricated using organic solvent mixtures and controlled cooling rates and its bio-compatibility with human adipose stem cells

Thermally induced phase separation method was used to make porous three-dimensional poly (l-lactic acid) scaffolds. The effect of imposed thermal profile during freezing of the poly (l-lactic acid) in dioxane solution on the scaffold was characterized by their micro-structure, porosity (%), pore sizes’ distribution, and mechanical strength. The porosity (%) decreased considerably with increasing concentrations of poly (l-lactic acid) in the solution, while a decreasing trend was observed with increasing cooling rates. The mechanical strength increases with increase in poly (l-lactic acid) concentration and also with increase in the cooling rate for both types of solvents. Therefore, mechanical strength was increased by higher cooling rates while the porosity (%) remained relatively consistent. Scaffolds made using higher concentrations of poly (l-lactic acid; 7% and 10% w/v) in solvent showed better mechanical strength which improved relatively with increasing cooling rates (1°C–40°C/min). This phenomenon of enhanced structural integrity with increasing cooling rates was more prominent in scaffolds made from higher initial poly (l-lactic acid) concentrations. Human adipose–derived stem cells were cultured on these scaffold (7% and 10% w/v) prepared by thermally induced phase separation at all cooling rates to measure the cell proliferation efficiency as a function of their micro-structural properties. Mean pore sizes played a crucial role in cell proliferation than percent porosity since all scaffolds were &gt;88% porous. The viability percent of human adipose tissue–derived adult stem cells increased consistently with longer periods of culture. Thus, poly (l-lactic acid) scaffolds prepared by thermally controlled thermally induced phase separation method could be a prime candidate for making ex vivo tissue-engineered grafts for surgical implantation.