[The method of determination for 2, 3-Butanedione in the air of workplace by high performance liquid chromatography with derivatization].

Objective: To establish a method for the determination of 2, 3-Butanedione (BUT) in the air of workplace, which including the process of collection by absorption in phosphoric acid aqueous solution and the process of analysis and detection by high performance liquid chromatography with derivatization. Methods: In October 2022, a porous glass plate absorption tube containing 10 ml of 0.01% phosphoric acid solution was used to collect BUT in the air of the workplace at a flow rate of 0.2 L/min. The absorption solution was derived by 2, 4-dinitrophenylhydrazine for 75 min and separated on a SB-C18 column (250 mm×4.6 mm, 5 µm) . At the column temperature of 30 ℃, the mixture of acetonitrile-water (V∶V, 1∶1) was eluted at the flow rate of 1.0 ml/min. It was detected by UV detector (λ=365 nm) , qualitatived by retention time and quantitatived by external standard. Results: It showed that BUT in phosphoric acid aqueous solution could be stored for at least 7 d at 4 ℃. There was a linear relationship within the determination range of 0.05-6.00 µg/ml, the linear regression equation was y=89.610x+0.133, r=0.9999. The sampling absorption efficiencies were 98.33%-100.00%, the detection limit of the method was 0.005 µg/ml, the minimum detection concentration was 0.016 mg/m(3) (based on V(0)=3.0 L) . The recovery rates were 95.96%-102.44%, the intra batch precision were 4.36%-7.78%, and the inter batch precision were 4.96%-6.06%. Conclusion: The method has the advantages of simple operation, high sensitivity and good accuracy. It can prevent the loss and degradation of BUT. It can be used for the determination of BUT in the air of workplace.

Molecular Mechanisms of the Deregulation of Muscle Contraction Induced by the R90P Mutation in Tpm3.12 and the Weakening of This Effect by BDM and W7.

Point mutations in the genes encoding the skeletal muscle isoforms of tropomyosin can cause a range of muscle diseases. The amino acid substitution of Arg for Pro residue in the 90th position (R90P) in γ-tropomyosin (Tpm3.12) is associated with congenital fiber type disproportion and muscle weakness. The molecular mechanisms underlying muscle dysfunction in this disease remain unclear. Here, we observed that this mutation causes an abnormally high Ca2+-sensitivity of myofilaments in vitro and in muscle fibers. To determine the critical conformational changes that myosin, actin, and tropomyosin undergo during the ATPase cycle and the alterations in these changes caused by R90P replacement in Tpm3.12, we used polarized fluorimetry. It was shown that the R90P mutation inhibits the ability of tropomyosin to shift towards the outer domains of actin, which is accompanied by the almost complete depression of troponin's ability to switch actin monomers off and to reduce the amount of the myosin heads weakly bound to F-actin at a low Ca2+. These changes in the behavior of tropomyosin and the troponin-tropomyosin complex, as well as in the balance of strongly and weakly bound myosin heads in the ATPase cycle may underlie the occurrence of both abnormally high Ca2+-sensitivity and muscle weakness. BDM, an inhibitor of myosin ATPase activity, and W7, a troponin C antagonist, restore the ability of tropomyosin for Ca2+-dependent movement and the ability of the troponin-tropomyosin complex to switch actin monomers off, demonstrating a weakening of the damaging effect of the R90P mutation on muscle contractility.

Advances into Understanding the Vital Role of the Mitochondrial Citrate Carrier (CIC) in Metabolic Diseases.

The mitochondrial citrate carrier (CIC) is a nuclear-encoded protein located in the inner mitochondrial membrane. By mediating efflux of citrate from the mitochondria to the cytosol, CIC links mitochondrial central carbon metabolism and cytosolic lipogenesis together. Abnormal activity or expression of CIC was found in cancers, developmental disorders and many other diseases. Recently, the specific inhibitors of CIC were proved to modify basic cellular metabolism, which in turn led to changes in disease course such as reverted steatohepatitis and cancer cell death. CIC is believed to be a key player and may serve as a novel therapeutic target in types of human metabolic diseases. Therefore, in this paper, we integrally described the structure and function of CIC. Then, we gave an overview of CIC related diseases including cancers, congenital diseases, pro-inflammatory effects and some other diseases. At the same time, the potential molecular mechanisms of CIC in the above diseases were illuminated. Finally, we illuminated some emerging areas for future investigation.

A potential flavor culture: Lactobacillus harbinensis M1 improves the organoleptic quality of fermented soymilk by high production of 2,3-butanedione and acetoin.

Lactic acid bacteria (LAB) are commonly used in soymilk fermentation to improve health-related functionality, but their contribution to sensory qualities is less valued. We characterized Lactobacillus harbinensis M1, Lactobacillus mucosae M2, Lactobacillus fermentum M4, Lactobacillus casei M8 and Lactobacillus rhamnosus C1 from naturally-fermented tofu whey, along with Streptococcus thermophilus ST3 from kefir XPL-1 fermented soymilk, to investigate their potential as starter cultures of fermented soymilk. They were characterized for antibiotic susceptibility, probiotic potential and their performance as starter cultures. All the LABs showed sensitivity to the tested antibiotics. L. casei M8 had strongest tolerance to synthetic gastrointestinal juice (<1.0 log CFU/mL loss), as well as antagonistic effects towards five food-borne pathogens. GC/MS analysis showed that L. harbinensis M1 produced significantly higher abundance (P < 0.05) of 2,3-butanedione (2.45 ppm) and acetoin (44.30 ppm), thus improving the overall sensory acceptability of fermented soymilk. The coding genes for the synthesis of 2,3-butanedione/acetoin (alsS, alsD, butA) were predicted from the whole-genome. A co-culture of L. harbinensis M1 and L. casei M8 produced a fermented soymilk product with both markedly improved flavor and good probiotic potential. It appears that L. harbinensis M1 has much potential for improving the organoleptic properties of fermented soymilk.

[Advances in flavoring-related bronchiolitis obliterans].

Occupational exposure to the diacetyl flavoring may cause bronchiolitis obliterans (BO) in workers. The first case of flavoring-related bronchiolitis obliterans was reported in 2002. Since then, similar cases have been identified among workers in various production industries in some countries and regions. At present, there are no cases reported in China. In order to improve the awareness of the disease and promote the prevention work, this article reviews the research progress of case recognition, risk factors, clinical manifestation and possible pathogenesis.

Compositional changes of the floral scent volatile emissions from Asian skunk cabbage (Symplocarpus renifolius, Araceae) over flowering sex phases.

INTRODUCTION: Flowering of the Asian skunk cabbage (Symplocarpus renifolius, Araceae) shows a sequential expression of female, bisexual and male sex phases. The protogynous thermogenic inflorescence has unpleasant odours, but the contributing chemical composition is poorly understood. OBJECTIVE: To determine the volatile composition of odour emissions from each S. renifolius flowering phase. METHODOLOGY: The dynamic headspace method was used to collect floral volatiles from six intact S. renifolius inflorescences in their natural habitat. Collected volatiles from the three flowering phases were analysed using gas chromatography-mass spectrometry/olfactometry (GC-MS/O). RESULTS: Female-phase inflorescences were characterised by an earthy-rotten-minty odour, while male-phase inflorescences typically exhibited a rotten-oily odour. Approximately 160 compounds were detected in volatiles from the three phases. Common to all phases were 3-methylbutyl 3-methylbutanoate, 1,8-cineole, dimethyl disulphide and sabinene, together accounting for 52 to 54% of total volatiles. GC-MS/O revealed that at least 28 volatiles including eight S-containing compounds contributed to the unpleasant odour of S. renifolius. Among them, dimethyl disulphide (onion-like), methional (potato-like), and the tentatively identified methyl dithioformate (garlic-like) were intense odour-active compounds in each floral phase. Additionally, 2-isopropyl-3-methoxypyrazine (IPMP) was a major contributor to the earthy odour that was characteristic of the female phase. CONCLUSIONS: No marked changes were observed in floral volatile compositions over the three flowering phases of S. renifolius. Instead, flower phase-dependent proportional changes of minor components (e.g. IPMP and 2,3-butanedione) altered the odour characteristics between the female and male phases.

Depolymerization of waste poly(methyl methacrylate) scraps and purification of depolymerized products.

A big challenge for the civilization in energy saving/waste management can be "the regeneration of monomers from the waste plastics followed by their re-polymerization" using an ideal recycling method. Herein, we investigate the thermal depolymerization of poly(methyl methacrylate) (PMMA) using thermogravimetric analysis coupled with mass spectrometry (TGA-MS). In this process, the polymer chains were decomposed to methyl methacrylate (MMA) in high yield and the degradation species were thoroughly characterized. The obtained MMA contained traces of byproducts. Firstly, the byproducts were found to be nonpolymerizable, secondly, their presence interrupt the polymerization reaction, and thirdly, they reduce the quality of re-polymerized PMMA (r-PMMA). This study reclaims that besides the main byproduct (methyl isobutyrate), traces of methyl pyruvate and 2,3-butanedione were also formed during the thermal depolymerization of PMMA. The formed 2,3-butanedione was found to be responsible for the unpleasant smell in the recovered MMA that also found itself in the r-PMMA. Further, the generated byproducts were eliminated from the r-PMMA by a dissolution/re-precipitation method. The structural characterizations of the recycled and purified PMMA were carried out by Fourier-transform-infrared spectroscopy (FT-IR), Hydrogen-1 (1H)- and Carbon-13 (13C)-nuclear magnetic resonance (NMR), differential scanning calorimetry (DSC) and gel permeation chromatography (GPC). The chemical properties of the r-PMMA and purified PMMA proved to be similar to that of the virgin commercial PMMA. This study can provide an effective and practical prototype for the recycling of waste PMMA scraps and thus reduction in pollution caused by the landfilling of waste PMMA scraps.

The Diacetyl-Exposed Human Airway Epithelial Secretome: New Insights into Flavoring-Induced Airways Disease.

Bronchiolitis obliterans (BO) is an increasingly important lung disease characterized by fibroproliferative airway lesions and decrements in lung function. Occupational exposure to the artificial food flavoring ingredient diacetyl, commonly used to impart a buttery flavor to microwave popcorn, has been associated with BO development. In the occupational setting, diacetyl vapor is first encountered by the airway epithelium. To better understand the effects of diacetyl vapor on the airway epithelium, we used an unbiased proteomic approach to characterize both the apical and basolateral secretomes of air-liquid interface cultures of primary human airway epithelial cells from four unique donors after exposure to an occupationally relevant concentration (∼1,100 ppm) of diacetyl vapor or phosphate-buffered saline as a control on alternating days. Basolateral and apical supernatants collected 48 h after the third exposure were analyzed using one-dimensional liquid chromatography tandem mass spectrometry. Paired t tests adjusted for multiple comparisons were used to assess differential expression between diacetyl and phosphate-buffered saline exposure. Of the significantly differentially expressed proteins identified, 61 were unique to the apical secretome, 81 were unique to the basolateral secretome, and 11 were present in both. Pathway enrichment analysis using publicly available databases revealed that proteins associated with matrix remodeling, including degradation, assembly, and new matrix organization, were overrepresented in the data sets. Similarly, protein modifiers of epidermal growth factor receptor signaling were significantly altered. The ordered changes in protein expression suggest that the airway epithelial response to diacetyl may contribute to BO pathogenesis.

Alignment of actin filament streams driven by myosin motors in crowded environments.

BACKGROUND: Cellular dynamics depend on cytoskeletal filaments and motor proteins. Collective movements of filaments driven by motor proteins are observed in the presence of dense filaments in in vitro systems. As multiple macromolecules exist within cells and the physiological ionic conditions affect their interactions, crowding might contribute to ordered cytoskeletal architecture because of collective behavior. METHODS: Using an in vitro reconstituted system, we observed the emergence of stripe patterns resulting from collective actin filament streaming driven by myosin motors in the presence of the crowding agent, methylcellulose (MC). RESULTS: Although at high KCl concentrations (150mM), actin filaments tended to dissociate from a myosin-coated surface, 1% MC prevented this dissociation and enabled filament movement on myosin molecules. At concentrations of actin filaments above 0.2mg/mL, the moving filaments accumulated and progressively formed long, dense bands. The bands were spaced at about 10-µm intervals. Increasing the KCl concentration up to 300mM resulted in narrowing of the spacing between the aligned bands. On the other hand, low KCl concentrations (≤25mM) induced broad streams, where actin filaments exhibited bidirectional movement. CONCLUSIONS: These results suggest that crowded environments can promote spatial patterning of the actin cytoskeleton, depending on the intensity of the myosin driving force and filament velocity, both modulated by the ionic strength. GENERAL SIGNIFICANCE: The mutual contribution of packing and driving forces provides insight into cytoskeleton organization in living cells, in which various macromolecules mingle.

Airway injury in an in vitro human epithelium-fibroblast model of diacetyl vapor exposure: diacetyl-induced basal/suprabasal spongiosis.

Inhalation exposure to diacetyl (DA) is associated with obliterative bronchiolitis (OB) in workers and induces OB-like fibrotic airway lesions in rats. The pathogenesis of OB is poorly understood in part due to complex interactions between airway epithelial, mesenchymal and blood-derived inflammatory cells. DA-induced airway toxicity in the absence of recruited-inflammatory/immune cells was characterized using an air-liquid interface (ALI) model consisting of human airway epithelium with (Epi/FT) and without (Epi) a mesenchymal component. ALI cultures were exposed to 25 mM DA-derived vapors (using vapor cups) for 1 h on day 0, 2 and 4. In some experiments, the tissues were exposed to 2,3-hexanedione (Hex) which is structurally-similar, but much less fibrogenic than DA. Lactate dehydrogenase activity and day 6 histopathologic changes associated with epithelial injury, including basal/suprabasal spongiosis, were increased following exposure of Epi/FT tissues to DA but not control or Hex vapors. IL-1a, IL-6, IL-8, sIL-1Ra, TGFa, MCP-3 and TNFa proteins were increased following DA exposure of Epi/FT tissues; only IL-1a, IL-8, sIL-1Ra and TGFa were increased following exposure of Epi tissues. MMP-1, MMP-3 and TIMP-1 proteins were increased following DA exposure of Epi/FT tissues; whereas MMP-2, MMP-7 and TIMP-2 were decreased, and production was largely dependent upon the presence of sub-epithelial stromal matrix/fibroblasts. Hex-induced protein changes were minimal. This in vitro study demonstrated that exposure of human airways to DA vapors induced epithelial injury (with the histopathologic feature of basal/suprabasal spongiosis) and increased release of pro-inflammatory and pro-fibrotic cytokines/chemokines as well as MMPs/TIMPs in the absence of recruited-inflammatory cells.

α,ß-Dicarbonyl reduction is mediated by the Saccharomyces Old Yellow Enzyme.

The undesirable flavor compounds diacetyl and 2,3-pentanedione are vicinal diketones (VDKs) formed by extracellular oxidative decarboxylation of intermediate metabolites of the isoleucine, leucine and valine (ILV) biosynthetic pathway. These VDKs are taken up by Saccharomyces and enzymatically converted to acetoin and 3-hydroxy-2-pentanone, respectively. Purification of a highly enriched diacetyl reductase fraction from Saccharomyces cerevisiae in conjunction with mass spectrometry identified Old Yellow Enzyme (Oye) as an enzyme capable of catalyzing VDK reduction. Kinetic analysis of recombinant Oye1p, Oye2p and Oye3p isoforms confirmed that all three isoforms reduced diacetyl and 2,3-pentanedione in an NADPH-dependent reaction. Transcriptomic analysis of S. cerevisiae (ale) and S. pastorianus (lager) yeast during industrial fermentations showed that the transcripts for OYE1, OYE2, arabinose dehydrogenase (ARA1), α-acetolactate synthase (ILV2) and α-acetohydroxyacid reductoisomerase (ILV5) were differentially regulated in a manner that correlated with changes in extracellular levels of VDKs. These studies provide insights into the mechanism for reducing VDKs and decreasing maturation times of beer which are of commercial importance.

Chemical Reactivity and Respiratory Toxicity of the α-Diketone Flavoring Agents: 2,3-Butanedione, 2,3-Pentanedione, and 2,3-Hexanedione.

Occupational exposure to 2,3-butanedione (BD) vapors has been associated with severe respiratory disease leading to the use of potentially toxic substitutes. We compared the reactivity and respiratory toxicity of BD with that of two structurally related substitutes, 2,3-pentanedione (PD) and 2,3-hexanedione (HD). Chemical reactivity of the diketones with an arginine substrate decreased with increasing chain length (BD > PD > HD). Animals were evaluated the morning after a 2-week exposure to 0, 100, 150, or 200 ppm BD, PD, or HD (postexposure) or 2 weeks later (recovery). Bronchial fibrosis was observed in 5/5 BD and 5/5 PD rats at 200 ppm and in 4/6 BD and 6/6 PD rats at 150 ppm in the postexposure groups. Following recovery, bronchial fibrosis was observed in all surviving rats exposed to 200 ppm BD (5/5) or PD (3/3) and in 2/10 BD and 7/9 PD rats exposed to 150 ppm. Bronchial fibrosis was observed only in 2/12 HD-exposed rats in the 200 ppm postexposure group. Patchy interstitial fibrosis affected lungs of recovery groups exposed to 200 ppm PD (3/3) or BD (1/5) and to 150 ppm PD (4/9) or BD (7/10) and correlated with pulmonary function deficits. BD and PD were more reactive and produced more bronchial fibrosis than HD.

Comparative key aroma compounds and sensory correlations of aromatic coconut water varieties: Insights from GC × GC-O-TOF-MS, E-nose, and sensory analysis.

Aroma is a key criterion in evaluating aromatic coconut water. A comparison regarding key aroma compounds and sensory correlations was made between Thailand Aromatic Green Dwarf (THD) and Cocos nucifera L. cv. Wenye No. 4 coconut water using E-nose and GC × GC-O-TOF-MS combined with chemometrics. Twenty-one volatile components of coconut water were identified by GC × GC-O-TOF-MS, and 5 key aroma compounds were analyzed by relative odor activity value and aroma extract dilution analysis. Moreover, the combination of the E-nose with orthogonal partial least squares was highly effective in discriminating between the two coconut water samples and screened the key sensors responsible for this differentiation. Additionally, the correlation between volatile compounds and sensory properties was established using partial least squares. The key aroma compounds of coconut water exhibited positive correlations with the corresponding sensory properties.

Effect of different isolation sources of Lactococcus lactis subsp. lactis on volatile metabolites in fermented milk.

Lactococcus lactis subsp. lactis (L. lactis subsp. lactis) is a commonly used starter cultures in fermented dairy products, contributing distinct flavor and texture characteristics with high application value. However, the strains from different isolates have different contributions to milk fermentation. Therefore, this study aimed to investigate the influence of L. lactis subsp. lactis isolated from various sources on the volatile metabolites present in fermented milk. In this study, L. lactis subsp. lactis from different isolation sources (yogurt, koumiss and goat yogurt) was utilized as a starter culture for fermentation. The volatile metabolites of fermented milk were subsequently analyzed by headspace solid phase microextraction gas chromatography-mass spectrography (HS-SPME-GC-MS). The results indicated significant differences in the structure and abundance of volatile metabolites in fermented milk produced with different isolates (R2Y = 0.96, Q2 = 0.88). Notably, the strains isolated from goat yogurt appeared to enhance the accumulation of ketones (goat yogurt vs yogurt milk: 50 %; goat yogur vs koumiss: 27.3 %)and aldehydes (goat yogurt vs yogurt milk: 21.4 %; goat yogurt vs koumiss: 54.5 %) in fermented milk than strains isolated from koumiss and yogurt milk. It significantly promoted the production of 8 flavor substances (1 substance with OAV ≥ 1 and 6 substances with OAV > 0.1) and enhanced the biosynthesis of valine, leucine, and isoleucine. This study provides valuable insights for the application of Lactococcus lactis subsp. lactis isolated from different sources in fermented dairy production and screening of potential starter cultures.

Effect of SR load and pH regulatory mechanisms on stretch-dependent Ca(2+) entry during the slow force response.

When cardiac muscle is stretched, there is an initial inotropic response that coincides with the stretch followed by a slower increase in twitch force that develops over several minutes (the "slow force response", or SFR). Unlike the initial response to stretch, the SFR is produced by an increase in Ca(2+) transient amplitude, but the cellular mechanisms that give rise to the increased transients are still debated. We have examined the relationship between the SFR, intracellular [Ca(2+)] and the inotropic state of right ventricular trabeculae from rat hearts at 37°C. The magnitude of the SFR varied with [Ca(2+)]o and stimulation frequency, so that the SFR was greatest for conditions associated with a reduced SR Ca(2+) content. The SFR was not blocked by the AT1 receptor blocker losartan, but was reduced by SN-6, an inhibitor of reverse mode Na(+)/Ca(2+)-exchange (NCX). The Na(+)/H(+)-exchange (NHE) inhibitor HOE642 had no effect in HCO3(-)-buffered solutions, but blocked 50% of the SFR in HCO3(-)-free solution. Inhibition of HCO3(-) transport by DIDS increased the SFR and made it sensitive to HOE642. The addition of cross-bridge cycle inhibitors (20mM BDM or 20µM blebbistatin) to the superfusate reduced the SFR as monitored by changes in Ca(2+). In HCO3(-)-free conditions, the SFR was associated with a slow acidification that was inhibited by BDM, and by stopping electrical stimulation. These results can be explained by stretch increasing metabolic demand and stimulating Na(+) entry via both NHE and the Na(+)/HCO3(-) transporters. This mechanism provides a novel link between inotropic state and stretch, as well as a way for the cell to compensate for increased acid load. The feedback mechanism between force and Ca(2+) transient amplitude that we describe is also limited by the degree of SR Ca(2+) load.

Nitric oxide regulates cardiac intracellular Na⁺ and Ca²âº by modulating Na/K ATPase via PKCε and phospholemman-dependent mechanism.

In the heart, Na/K-ATPase regulates intracellular Na(+) and Ca(2+) (via NCX), thereby preventing Na(+) and Ca(2+) overload and arrhythmias. Here, we test the hypothesis that nitric oxide (NO) regulates cardiac intracellular Na(+) and Ca(2+) and investigate mechanisms and physiological consequences involved. Effects of both exogenous NO (via NO-donors) and endogenously synthesized NO (via field-stimulation of ventricular myocytes) were assessed in this study. Field stimulation of rat ventricular myocytes significantly increased endogenous NO (18 ± 2 µM), PKCε activation (82 ± 12%), phospholemman phosphorylation (at Ser-63 and Ser-68) and Na/K-ATPase activity (measured by DAF-FM dye, western-blotting and biochemical assay, respectively; p<0.05, n=6) and all were abolished by Ca(2+)-chelation (EGTA 10mM) or NOS inhibition l-NAME (1mM). Exogenously added NO (spermine-NONO-ate) stimulated Na/K-ATPase (EC50=3.8 µM; n=6/grp), via decrease in Km, in PLM(WT) but not PLM(KO) or PLM(3SA) myocytes (where phospholemman cannot be phosphorylated) as measured by whole-cell perforated-patch clamp. Field-stimulation with l-NAME or PKC-inhibitor (2 µM Bis) resulted in elevated intracellular Na(+) (22 ± 1.5 and 24 ± 2 respectively, vs. 14 ± 0.6mM in controls) in SBFI-AM-loaded rat myocytes. Arrhythmia incidence was significantly increased in rat hearts paced in the presence of l-NAME (and this was reversed by l-arginine), as well as in PLM(3SA) mouse hearts but not PLM(WT) and PLM(KO). We provide physiological and biochemical evidence for a novel regulatory pathway whereby NO activates Na/K-ATPase via phospholemman phosphorylation and thereby limits Na(+) and Ca(2+) overload and arrhythmias. This article is part of a Special Issue entitled "Na(+) Regulation in Cardiac Myocytes".

Preservation of cardiomyocytes from the adult heart.

Cardiomyocytes represent one of the most useful models to conduct cardiac research. A single adult heart yields millions of cardiomyocytes, but these cells do not survive for long after isolation. We aimed to determine whether inhibition of myosin II ATPase that is essential for muscle contraction may preserve fully differentiated adult cardiomyocytes. Using inhibitors of the myosin II ATPase, blebbistatin and N-benzyl-p-toluene sulphonamide (BTS), we preserved freshly isolated fully differentiated adult primary cardiomyocytes that were stored at a refrigerated temperature. Specifically, preserved cardiomyocytes stayed viable for a 2-week period with a stable expression of cardiac genes and retained the expression of key markers characteristic of cardiomyocytes. Furthermore, voltage-clamp, action potential, calcium transient and contractility studies confirmed that the preserved cardiomyocytes are comparable to freshly isolated cells. Long-term exposure of preserved cardiomyocytes to four tyrosine kinase inhibitors, sunitinib malate, dasatinib, sorafenib tosylate and imatinib mesylate, revealed their potential to induce cardiac toxicity that was manifested with a decrease in contractility and induction of cell death, but this toxicity was not observed in acute experiments conducted over the time course amenable to freshly prepared cardiomyocytes. This study introduces the concept that the inhibition of myosin II ATPase safeguards the structure and function of fully differentiated adult cardiomyocytes. The fact that these preserved cardiomyocytes can be used for numerous days after preparation makes them a robust and versatile tool in cardiac research and allows the investigation of long-term exposure to novel drugs on cardiomyocyte function.

Promotion or Inhibition Effects of Exogenous Glutathione-Degraded Amino Acids on the Formation of 2,3-Butanedione and Pyrazines via Varied Pathways of Interaction with α-Dicarbonyl Compounds Derived from N-(1-Deoxy-d-xylulos-1-yl)-alanine.

The contribution of glutathione (GSH) and free amino acids degraded from GSH to the generation of pyrazines and 2,3-butanedione was illustrated during their interaction in the thermal treatment of the Amadori compound of alanine and xylose (ARP). GSH-degraded amino acids, glutamic acid (Glu), cysteine (Cys), and glycine (Gly), but not pyroglutamic acid (pGlu), could effectively capture α-dicarbonyls to facilitate the formation of pyrazines when ARP was heated with GSH. Deoxypentosones, the precursors of 2,3-butanedione, were largely consumed in the ARP-GSH model by the interaction with GSH and its degradative Cys compared with the ARP model. The addition of GSH and deoxypentosones inhibited the further degradation of deoxypentosones, resulting in less formation of 2,3-butanedione and other α-dicarbonyl compounds. Meanwhile, the reaction between GSH-degraded Cys and deoxypentosones to form sulfur-containing compounds such as thiols accelerated the consumption of deoxypentosones; thereby, the formation of 2,3-butanedione was severely interfered. However, this inhibition was compensated for by the GSH-degraded Gly through the addition between Gly and MGO and the subsequent deamination. The involvement of exogenous GSH could simultaneously boost the yields of 2,3-butanedione and pyrazines compared with those of ARP heated alone. As the degree of GSH degradation strengthened in the ARP-thermal-degraded GSH models, the yields of both pyrazines and 2,3-butanedione steadily increased.

A novel method to extend viability and functionality of living heart slices.

Living heart slices have recently emerged as a powerful experimental model for fundamental cardiac research. By retaining the structure and function of the native myocardium while maintaining the simplicity of cell culture models, heart slices can be easily employed in electrophysiological, pharmacological, biochemical, and structural investigations. One single heart yields many slices (>20 slices for rodents, >100 slices for porcine or human hearts), however due to the low throughput of most assays and rapid slice degeneration within 24 h of preparation, many slices remain unused and are discarded at the end of the preparation day. Here we present a novel method to extend viability and functionality of living heart slices, enabling their use in experiments over several consecutive days following preparation. By combining hypothermic conditions with inhibition of myosin II ATPase using 2,3-butanedione monoxime (BDM), slices prepared from the left ventricle of porcine hearts remain viable and exhibit preserved contractile function and morphology for up to 6 days. Electrophysiological function was also confirmed over the 6 days by extracellular field potentials recordings. This simple method not only maximizes the use of slices prepared from one single heart, thus reducing the number of animals required, but also increases data reproducibility by allowing multiple electrophysiological, pharmacological, biochemical, and structural studies to be performed from the same heart.

Characteristics of changes in volatile organic compounds and microbial communities during the storage of pickles.

The variations of volatile organic compounds (VOCs) and microbial communities of three pickles during storage at 4°C for one week were analyzed by headspace-gas chromatography-ion mobility spectrometry (HS-GC-IMS), high-throughput sequencing, and Spearman correlation analysis. A total of 50 VOCs were identified from three pickles. During storage, most alcohols, aldehydes, ketones, and esters decreased, while acids increased, and sulfides, alkenes, and phenols were relatively equal. Firmicutes, Cyanobacteria, and Proteobacteria were the predominant bacterial phyla, and Weissella, Streptophyta, Leuconostoc, Bacillariophyta, and Lactobacillus were the predominant bacterial genera in three pickles. The bacterial diversity level significantly decreased during storage (P < 0.05). Spearman correlation coefficient indicated that Leuconostoc, Lactobacillus, and Weissella were highly correlated with the flavor of pickles, while Bacillariophyta and Streptophyta were highly correlated with the flavor formation of pickles during storage. These results could contribute to a better understanding of the impact of bacteria in flavor formation during pickle storage.