Exogenous Glucose Interferes with Antimicrobial-Mediated ROS Accumulation and Bacterial Death.

Glucose is widely used in the reconstitution of intravenous medications, which often include antimicrobials. How glucose affects antimicrobial activity has not been comprehensively studied. The present work reports that glucose added to bacteria growing in a rich medium suppresses the bactericidal but not the bacteriostatic activity of several antimicrobial classes, thereby revealing a phenomenon called glucose-mediated antimicrobial tolerance. Glucose, at concentrations corresponding to blood-sugar levels of humans, increased survival of Escherichia coli treated with quinolones, aminoglycosides, and cephalosporins with little effect on minimal inhibitory concentration. Glucose suppressed a ROS surge stimulated by ciprofloxacin. Genes involved in phosphorylated fructose metabolism contributed to glucose-mediated tolerance, since a pfkA deficiency, which blocks the formation of fructose-1,6-bisphosphate, eliminated protection by glucose. Disrupting the pentose phosphate pathway or the TCA cycle failed to alter glucose-mediated tolerance, consistent with an upstream involvement of phosphorylated fructose. Exogenous sodium pyruvate or sodium citrate reversed glucose-mediated antimicrobial tolerance. Both metabolites bypass the effects of fructose-1,6-bisphosphate, a compound known to scavenge hydroxyl radical and chelate iron, activities that suppress ROS accumulation. Treatment with these two compounds constitutes a novel way to mitigate the glucose-mediated antimicrobial tolerance that may exist during intravenous antimicrobial therapy, especially for diabetes patients.

Sodium citrate targeting Ca2+/CAMKK2 pathway exhibits anti-tumor activity through inducing apoptosis and ferroptosis in ovarian cancer.

INTRODUCTION: Ovarian cancer (OC) is known for its high mortality rate. Although sodium citrate has anti-tumor effects in various cancers, its effect and mechanism in OC remain unclear. OBJECTIVES: To analyze the inhibitory effect of sodium citrate on ovarian cancer cells and the underlying mechanism. METHODS: Cell apoptosis was examined by TUNEL staining, flow cytometry, and ferroptosis was examined intracellular Fe2+, MDA, LPO assays, respectively. Cell metabolism was examined by OCR and ECAR measurements. Immunoblotting and immunoprecipitation were used to elucidate the mechanism. RESULTS: This study suggested that sodium citrate not only promoted ovarian cancer cell apoptosis but also triggeredferroptosis, manifested as elevated levels of Fe2+, LPO, MDA andlipid ROS production. On one hand, sodium citrate treatment led to a decrease of Ca2+ content in the cytosol by chelatingCa2+, which further inhibited the Ca2+/CAMKK2/AKT/mTOR signaling, thereby suppressing HIF1α-dependent glycolysis pathway and inducing cell apoptosis. On the other hand, the chelation of Ca2+ by sodium citrate resulted in inactivation of CAMKK2 and AMPK, leading to increase of NCOA4-mediated ferritinophagy, causing increased intracellular Fe2+ levels. More importantly, the inhibition of Ca2+/CAMKK2/AMPK signaling pathway reduced the activity of the MCU and Ca2+ concentration within the mitochondria, resulting in an increase in mitochondrial ROS. Additionally, metabolomic analysis indicated that sodium citrate treatment significantly increased de novo lipid synthesis. Altogether, these factors contributed to ferroptosis. As expected, Ca2+ supplementation successfully reversed the cell death and decreased tumor growth induced by sodium citrate. Inspiringly, it was found that coadministration of sodium citrate increased the sensitivity of OC cells to chemo-drugs. CONCLUSION: These results revealed that the sodium citrate exerted its anti-cancer activity by inhibiting Ca2+/CAMKK2-dependent cell apoptosis and ferroptosis. Sodium citrate will hopefully serve as a prospective compound for OC treatment and for improvingthe efficacy of chemo-drugs.

Acute effects of sodium citrate supplementation on competitive performance and lactate level of elite fitness challenge athletes: A crossover, placebo-controlled, double-blind study.

Purpose: The performance of sodium citrate has been investigated in high-intensity exercises, but fewer studies have addressed the role of citrate in weight-bearing exercises. Methods: Twenty fitness challenge athletes, aged 24-32 years, volunteered to participate in this crossover, placebo-controlled, double-blind study. Initially, ten athletes were given a placebo and asked to complete a fitness challenge (i.e., chin-ups, squat jumps, dips, walking lunges, sit-ups, and burpees-devil press). Another ten athletes were supplemented with sodium citrate 0.5 g/kg body mass supplements 3 h prior to performing the fitness challenges. The same procedures were completed two days later with the supplement and placebo dextrose groups switched in a cross-over design. Athletes and assessors were blinded for the experimental condition (placebo vs. verum). Lactate levels were measured 5 min after exercise. The athletes' performance on each item of the fitness challenge as well as their lactate levels, were compared. Differences between the means of the measured variables were contrasted using a dependent t-test. Results: Supplementing sodium citrate substantially improved athletes' performance in all six fitness challenge items (p < 0.05, 0.69

Anisotropically Conductive Hydrogels with Directionally Aligned PEDOT:PSS in a PVA Matrix.

Electrical anisotropy, which is characterized by the efficient transmission of electrical signals in specific directions, is prevalent in both natural and engineered systems. However, traditional anisotropically conductive materials are often rigid and dry, thus limiting their utility in applications aiming for the seamless integration of various technologies with biological tissues. In the present study, we introduce a method for precisely controlling the microstructures of conductive and insulating polymers to create highly anisotropically conductive composite hydrogels. Our methodology involves combining aligned poly(vinyl alcohol) microfibrils, infused poly(3,4-ethylenedioxythiophene) polystyrenesulfonate, and sodium citrate precipitation to form dense, aligned conductive paths. This significantly enhances the electrical conductivity anisotropy (σ∥/σ⊥ ≈ 60.8) within these composite hydrogels.

Sellado antibiótico de reservorios subcutáneos: estabilidad de soluciones con vancomicina / Antimicrobial lock solutions for implantable central venous devices: stability of vancomycin solutions

Los reservorios subcutáneos son un tipo de catéter venoso central (CVC). Cuando se usan catéteres venosos centrales (CVC), el personal sanitario necesita evitar dos grandes riesgos: formación de coágulos e infecciones bacterianas. Para prevenir y evitar la contaminación de los catéteres en los pacientes hospitalizados y ambulatorios, se han implementado diversas alternativas, como el llamado “sellado antibiótico de catéteres” (SAC). De este modo, se ha sugerido la utilización de soluciones con agentes antimicrobianos, a las que se suelen adicionar sustancias con efecto anticoagulante y/o con efecto antibiofilm. Empero, se requiere que la estabilidad de dichas soluciones sea comprobada mediante técnicas como la cromatografía líquida de alta resolución (HPLC), además de las pruebas de eficacia antimicrobiana, para así poder establecer la seguridad de los pacientes. En este entorno, se plantea el presente trabajo de revisión bibliográfica, con el objetivo de incluir las investigaciones de mayor representación clínica a este respecto, para evidenciar el comportamiento de las soluciones de sellado antibiótico de catéteres en distintas condiciones de almacenamiento y uso. En particular, esta revisión se centra en soluciones con vancomicina. De acuerdo con los estudios consultados, las soluciones de vancomicina con citrato de sodio (agente quelante) son las que presentan las mejores características en cuanto a estabilidad físico-química y eficacia como soluciones de sellado.(AU)

Subcutaneous reservoirs are a type of central venous catheter. When using central venous catheters, healthcare workers need to avoid two major risks: clot formation and bacterial infections. To prevent and avoid catheter contamination in both hospitalized patients and outpatients, several strategies have been carried out, such as the so-called ” antibiotic-based catheter lock solution”. Therefore, it has been suggested to implement the use of solutions with antimicrobial agents, to which anticoagulant and/or antibiofilm substances are often added.However, the stability of such solutions needs to be tested by techniques such as high performance liquid chromatography (HPLC), in addition to antimicrobial efficacy testing, in order to establish patient safety. In consequence, this literature review aims to include the most clinically representative research towards these aspects, to demonstrate the behaviour of antibiotic-based catheter lock solutions under different conditions of storage and use. In particular, this review focuses on solutions containing vancomycin. According to the studies consulted, vancomycin solutions with sodium citrate (chelating agent) present the best stability characteristics in terms of physicochemical properties and efficacy.(AU)

Sodium Citrate Nanoparticles Induce Dual-Path Pyroptosis for Enhanced Antitumor Immunotherapy through Synergistic Ion Overload and Metabolic Disturbance.

Metabolic reprogramming, as one of the characteristics of cancer, is associated with tumorigenesis, growth, or migration, and the modulation of metabolic pathways has emerged as a novel approach for cancer therapy. However, the conventional metabolism-mediated apoptosis process in tumor cells exhibits limited immunogenicity and inadequate activation of antitumor immunity. Herein, phospholipid-coated sodium citrate nanoparticles (PSCT NPs) are successfully prepared, which dissolve in tumor cells and then release significant amounts of citrate ions and Na+ ions. Massive quantities of ions lead to increased intracellular osmotic pressure, which activates the caspase-1/gasdermin D (GSDMD) mediated pyroptosis pathway. Simultaneously, citrate induces activation of the caspase-8/gasdermin C (GSDMC) pathway. The combined action of these two pathways synergistically causes intense pyroptosis, exhibiting remarkable antitumor immune responses and tumor growth inhibition. This discovery provides new insight into the potential of nanomaterials in modulating metabolism and altering cell death patterns to enhance antitumor immunotherapy.

Double-edged effect of sodium citrate in Nile tilapia (Oreochromis niloticus): Promoting lipid and protein deposition vs. causing hyperglycemia and insulin resistance.

Citrate is an essential substrate for energy metabolism that plays critical roles in regulating glucose and lipid metabolic homeostasis. However, the action of citrate in regulating nutrient metabolism in fish remains poorly understood. Here, we investigated the effects of dietary sodium citrate on growth performance and systematic energy metabolism in juvenile Nile tilapia (Oreochromis niloticus). A total of 270 Nile tilapia (2.81 ± 0.01 g) were randomly divided into three groups (3 replicates per group, 30 fish per replicate) and fed with control diet (35% protein and 6% lipid), 2% and 4% sodium citrate diets, respectively, for 8 weeks. The results showed that sodium citrate exhibited no effect on growth performance (P > 0.05). The whole-body crude protein, serum triglyceride and hepatic glycogen contents were significantly increased in the 4% sodium citrate group (P < 0.05), but not in the 2% sodium citrate group (P > 0.05). The 4% sodium citrate treatment significantly increased the serum glucose and insulin levels at the end of feeding trial and also in the glucose tolerance test (P < 0.05). The 4% sodium citrate significantly enhanced the hepatic phosphofructokinase activity and inhibited the expression of pyruvate dehydrogenase kinase isozyme 2 and phosphor-pyruvate dehydrogenase E1 component subunit alpha proteins (P < 0.05). Additionally, the 4% sodium citrate significantly increased hepatic triglyceride and acetyl-CoA levels, while the expressions of carnitine palmitoyl transferase 1a protein were significantly down-regulated by the 4% sodium citrate (P < 0.05). Besides, the 4% sodium citrate induced crude protein deposition in muscle by activating mTOR signaling and inhibiting AMPK signaling (P < 0.05). Furthermore, the 4% sodium citrate significantly suppressed serum aspartate aminotransferase and alanine aminotransferase activities, along with the lowered expression of pro-inflammatory genes, such as nfκb, tnfα and il8 (P < 0.05). Although the 4% sodium citrate significantly increased phosphor-nuclear factor-kB p65 protein expression (P < 0.05), no significant tissue damage or inflammation occurred. Taken together, dietary supplementation of sodium citrate could exhibit a double-edged effect in Nile tilapia, with the positive aspect in promoting nutrient deposition and the negative aspect in causing hyperglycemia and insulin resistance.

Variations in the full blood count parameters among apparently healthy humans in the Ho municipality using ethylenediamine tetraacetic acid (EDTA), sodium citrate and lithium heparin anticoagulants: A laboratory-based cross-sectional analytical study.

Background: Several studies have shown that various anticoagulants used for collection of blood samples produce varying effects on haematological analyses. Tripotassium ethylenediamine tetra-acetic acid (K3EDTA), sodium citrate and lithium heparin remain the most used anticoagulants employed in hematological analysis. There is paucity of data on the effect of these anticoagulants on haematological parameters in humans in Ghana. We assessed the suitability of K3EDTA, sodium citrate and lithium heparin for routine Full Blood Count (FBC) investigation. Method: A laboratory-based analytical cross-sectional study was conducted using blood samples from 55 conveniently sampled apparently healthy tertiary students from January 2021 to October 2021. Blood samples were taken from each participant into 3 anticoagulant tubes: K3EDTA, sodium citrate and lithium heparin and FBC parameters estimated using the Mindray automated haematology analyzer. One-way ANOVA Kruskal-Wallis test, Mann-Whitney U, Intra-class correlation coefficient (ICC) analysis, Bland-Altman's plot and Lin's concordance correlation coefficient were used where appropriate to ascertain the level of variation, consistency, and agreements among and between results. Normality testing using Shapiro-Wilk test statistic revealed non-Gaussian distribution of data, hence, were presented as median, minimum, and maximum. Data generated were analyzed using STATA v15 and MedCalc v20 where appropriate for statistical analysis. P-values <0.05 were considered statistically significant. Results: The study comprised 34 males and 21 females. The median age for males (23 years: min = 20, max = 34) was statistically comparable (p = 0.2652) to that of females (22 years: min = 18, max = 34). We observed excellent consistency in the estimation of MCV (ICC = 0.94), MCH (ICC = 0.98), MCHC (ICC = 0.91), GRAN# (ICC = 0.92) and LYMPH% (ICC = 0.91) across the three anticoagulants. Heparin and K3EDTA largely agreed on most of the FBC parameters, 50.0% (7/14) including HGB, MCV, MCH, PLT, LYMPH#, GRAN# and GRAN%. Meanwhile using K3EDTA as a standard, heparin produced almost perfect agreement only in the assessment of RBC (CCC = 0.992) while a substantial agreement was observed in the assessment of HGB (0.971), HCT (0.958) and MCH (0.987). Citrate agreed substantially with K3EDTA in the assessment of LYMPH% (CCC = 0.964) and moderately in the assessment of MCV (CCC = 0.948) and MCH (0.913). Overall, compared to K3EDTA, heparin was highly precise and accurate in the estimation of HGB, RBC, HCT and MCH while citrate determined MCV and MCH more accurately and precisely. Conclusion: Citrated blood consistently produced lower FBC values compared to heparin and K3EDTA and hence suggests not reliable in the assessment of FBC among humans. Heparin agreed largely with K3EDTA in the estimation of FBC parameters and may be used as a better alternative anticoagulant in the absence of K3EDTA however with great caution.

Improving Methanol Utilization by Reducing Alcohol Oxidase Activity and Adding Co-Substrate of Sodium Citrate in Pichia pastoris.

Methanol, which produced in large quantities from low-quality coal and the hydrogenation of CO2, is a potentially renewable one-carbon (C1) feedstock for biomanufacturing. The methylotrophic yeast Pichia pastoris is an ideal host for methanol biotransformation given its natural capacity as a methanol assimilation system. However, the utilization efficiency of methanol for biochemical production is limited by the toxicity of formaldehyde. Therefore, reducing the toxicity of formaldehyde to cells remains a challenge to the engineering design of a methanol metabolism. Based on genome-scale metabolic models (GSMM) calculations, we speculated that reducing alcohol oxidase (AOX) activity would re-construct the carbon metabolic flow and promote balance between the assimilation and dissimilation of formaldehyde metabolism processes, thereby increasing the biomass formation of P. pastoris. According to experimental verification, we proved that the accumulation of intracellular formaldehyde can be decreased by reducing AOX activity. The reduced formaldehyde formation upregulated methanol dissimilation and assimilation and the central carbon metabolism, which provided more energy for the cells to grow, ultimately leading to an increased conversion of methanol to biomass, as evidenced by phenotypic and transcriptome analysis. Significantly, the methanol conversion rate of AOX-attenuated strain PC110-AOX1-464 reached 0.364 g DCW/g, representing a 14% increase compared to the control strain PC110. In addition, we also proved that adding a co-substrate of sodium citrate could further improve the conversion of methanol to biomass in the AOX-attenuated strain. It was found that the methanol conversion rate of the PC110-AOX1-464 strain with the addition of 6 g/L sodium citrate reached 0.442 g DCW/g, representing 20% and 39% increases compared to AOX-attenuated strain PC110-AOX1-464 and control strain PC110 without sodium citrate addition, respectively. The study described here provides insight into the molecular mechanism of efficient methanol utilization by regulating AOX. Reducing AOX activity and adding sodium citrate as a co-substrate are potential engineering strategies to regulate the production of chemicals from methanol in P. pastoris.

Diminishing the Pathogenesis of the Food-Borne Pathogen Serratia marcescens by Low Doses of Sodium Citrate.

Protecting food from bacterial contamination is crucial for ensuring its safety and avoiding foodborne illness. Serratia marcescens is one of the food bacterial contaminants that can form biofilms and pigments that spoil the food product and could cause infections and illness to the consumer. Food preservation is essential to diminish such bacterial contaminants or at least reduce their pathogenesis; however, it should not affect food odor, taste, and consistency and must be safe. Sodium citrate is a well-known safe food additive and the current study aims to evaluate its anti-virulence and anti-biofilm activity at low concentrations against S. marcescens. The anti-virulence and antibiofilm activities of sodium citrate were evaluated phenotypically and genotypically. The results showed the significant effect of sodium citrate on decreasing the biofilm formation and other virulence factors, such as motility and the production of prodigiosin, protease, and hemolysins. This could be owed to its downregulating effect on the virulence-encoding genes. An in vivo investigation was conducted on mice and the histopathological examination of isolated tissues from the liver and kidney of mice confirmed the anti-virulence activity of sodium citrate. In addition, an in silico docking study was conducted to evaluate the sodium citrate binding ability to S. marcescens quorum sensing (QS) receptors that regulates its virulence. Sodium citrate showed a marked virtual ability to compete on QS proteins, which could explain sodium citrate's anti-virulence effect. In conclusion, sodium citrate is a safe food additive and can be used at low concentrations to prevent contamination and biofilm formation by S. marcescens and other bacteria.

Sodium citrate effectively used in shed mediastinal blood autotransfusion after cardiac surgery.

BACKGROUND: We used sodium citrate as an alternative anticoagulation agent to heparin in the procedure of autologous blood transfusion with patients with postoperative haemorrhage after CPB. The aim of study was to evaluate the efficacy and safety of sodium citrate used in shed mediastinal blood autotransfusion after cardiac surgery. METHODS: Ninety-three patients were divided into two groups in this study. In the control group, 52 patients' shed mediastinal blood was discarded. The reinfusion group consisted of 41 patients receiving a reinfusion of washed autologous red cells from shed mediastinal blood. Each 400 mL shed blood sample was anticoagulated by 140 mL of 1.6% diluted sodium citrate in the washing procedure using a blood recovery machine. Hemoglobin (Hb), hematocrit (Hct), and electrolyte concentrations in both the patients and shed mediastinal blood were measured before and after this procedure. RESULTS: The mean volume of autotransfused shed blood was 239.5 ± 54.6 mL.The Hct of the washed red cells was 56.8 ± 6.1%. Significantly, fewer units of allogeneic blood were required per patient in the reinfusion group at 24 h postoperatively (2.91 ± 1.34 vs 4.03 ± 0.19 U, p = 0.002). At 24 h postoperatively, Hb and Hct levels were higher in the reinfusion group than in the control group. The calcium ion concentration was very low in the shed mediastinal blood, 0.25 ± 0.08 mmol/L, and was lower after washing, 0.15 ± 0.04 mmol/L. CONCLUSIONS: Sodium citrate, as an alternative anticoagulant agent, can be used in autologous shed mediastinal blood transfusion after CPB cardiac surgery. This procedure can effectively reduce the amount of allogeneic blood for patients with haemorrhage.

Sodium citrate increases the aggregation capacity of calcium ions during microbial mineralization to accelerate the formation of calcium carbonate.

The microbially induced carbonate precipitation (MICP) technique is widely used in soil heavy metal pollution control. Microbial mineralization involves extended mineralization times and slow crystallization rates. Thus, it is important to discover a method to accelerate mineralization. In this study, we selected six nucleating agents to screen and investigated the mineralization mechanism using polarized light microscopy, scanning electron microscopy, X-ray diffraction and Fourier-transform infrared spectroscopy. The results showed that sodium citrate removed 90.1% Pb better than traditional MICP and generated the highest amount of precipitation. Interestingly, due to the addition of sodium citrate (NaCit), the rate of crystallization increased and vaterite was stabilized. Moreover, we constructed a possible model to explain that NaCit increases the aggregation capacity of calcium ions during microbial mineralization to accelerate the formation of calcium carbonate (CaCO3). Thus, sodium citrate can increase the rate of MICP bioremediation, which is important for improving MICP efficiency.

Highly Heterogeneous Morphology of Cobalt Oxide Nanostructures for the Development of Sensitive and Selective Ascorbic Acid Non-Enzymatic Sensor.

The surface tailored metal oxide nanostructures for the development of non-enzymatic sensors are highly demanded, but it is a big task due to the wide range of complexities during the growth process. The presented study focused on the surface modification of the heterogeneous morphology of cobalt oxide (Co3O4) prepared by the hydrothermal method. Further surface modification was conducted with the use of sodium citrate as a reducing and surface modifying agent for the Co3O4 nanostructures through the high density of oxygenated terminal groups from the citrate ions. The citrate ions enabled a significant surface modification of the Co3O4 nanostructures, which further improved the electrochemical properties of the Co3O4 material toward the design of the non-enzymatic ascorbic acid sensor in a phosphate buffer solution of pH 7.4. The morphology and crystal arrays of the Co3O4 nanostructures were studied by scanning electron microscopy (SEM) and powder X-ray diffraction (XRD) techniques. These physical characterizations showed the highly tailored surface features of Co3O4 nanostructures and a significant impact on the crystal properties. The electrochemical activity of Co3O4 was studied by chronoamperometry, linear sweep voltammetry, and cyclic voltammetry (CV) for the detection of ascorbic acid. The linear range of the proposed sensor was measured from 0.5 mM to 6.5 mM and a low limit of detection of 0.001 mM was also estimated. The presented Co3O4 nanostructures exhibited significant surface roughness and surface area, consequently playing a vital role toward the selective, sensitive, and stable detection of ascorbic acid. The use of a low cost surface modifying agent such as sodium citrate could be of great interest for the surface roughness and high surface area of nanostructured materials for the improved electrochemical properties for the biomedical, energy storage, and conversion systems.

Vancomycin and daptomycin stability in heparin or sodium citrate lock solutions.

BACKGROUND: We investigated physical and chemical stability of daptomycin and vancomycin in heparin or sodium citrate lock solutions. The aim of this study was to find the optimal combination of antimicrobials and additives for lock solutions, which maximized patient safety. METHODS: Vancomycin and daptomycin were diluted with heparin or sodium citrate to achieve final concentrations of vancomycin-heparin 2.5 mg/mL-833.33 U/mL, vancomycin-citrate 2.5-33.3 mg/mL, daptomycin-heparin 5 mg/mL-800 U/mL, and daptomycin-citrate 5-32 mg/mL and they were stored at room temperature (+25°C), 4°C, -20°C, and 37°C. Physical and chemical stability were analyzed for each antibiotic-anticoagulant combination in all conditions immediately after preparation, at 24, 48, 72 h and at different time points until unstable concentrations were obtained. Daptomycin-sodium citrate microbiological activity was also studied by evaluating two Staphylococcus aureus cultures in a calcium enriched medium with a daptomycin E test, with and without sodium citrate. RESULTS: After incubation at 37°C vancomycin and daptomycin combined with heparin retained at least 90% of the initial concentration over 48 h. Vancomycin-sodium citrate solution stored at 37°C reduced more than 10% of the initial concentration at 24 h. On the other hand, daptomycin-sodium citrate preparation was stable at 37°C for 72 h but the microbiological activity of daptomycin was lower in the presence of sodium citrate. CONCLUSIONS: The purpose is to prepare vancomycin and daptomycin lock solutions combined with heparin. They should be changed at 48 h and its stability is over 3 days at 25°C and 7 days at 4°C, which allow Hospital Pharmacy Services to manage their stocks. Daptomycin-sodium citrate combination is more stable for extended periods but its bioactivity has not been demonstrated.

Enhanced acidogenic fermentation from Al-rich waste activated sludge by combining lysozyme and sodium citrate pretreatment: Perspectives of Al stabilization and enzyme activity.

The accumulation of poly aluminum chloride (PAC) in dewatered waste activated sludge (WAS) can cause severe Al pollution and significantly reduce the production of volatile fatty acids (VFAs) from anaerobic fermentation. Herein, the combination of lysozyme and sodium citrate pretreatment was applied to stabilize the aluminum and enhance the VFAs production via anaerobic fermentation. The complexation and stabilization of aluminum by the citrate was efficient, which is conducive to relieving the inhibition of aluminum on lysozymes and other extracellular hydrolases. Compared with the control group, the lysozyme, protease and α-glucosidase activities were obtained at 1.86, 1.72, and 1.15 times, respectively, following the pretreatment. 129.71 mg/g volatile suspended solids (VSS) of soluble proteins and 26.3 mg/g VSS of polysaccharides were obtained within 4 h, together with the degradation of 124 % more proteins and 75 % more polysaccharides within three days. This provided a sufficient number of substrates for VFA production. 588.4 mg COD/g VSS of total VFAs were obtained after the six-day anaerobic fermentation from Al-rich WAS following the combination of lysozyme and sodium citrate pretreatment, which was 7.3 times higher than that of the control group. This study presents a novel approach for enhancing VFA production in anaerobic fermentation as well as reducing risk of Al hazards from Al-rich WAS.

Development of chondrocyte-laden alginate hydrogels with modulated microstructure and properties for cartilage regeneration.

Alginate hydrogel is an attractive biomaterial for cell microencapsulation. The microarchitecture of hydrogels can regulate cellular functions. This study aims to investigate the applicability of sodium citrate buffer (SCB) as a culture medium supplement for modulating the microstructure of alginate microbeads to provide a favorable microenvironment for chondrogenic induction. The chondrocyte-laden microbeads, with and without TGF-ß3 incorporation, were produced through an encapsulator. The obtained small-sized microbeads (~300 µm) were exposed to a treatment medium containing SCB, composed of varied concentrations of sodium citrate (1.10-1.57 mM), sodium chloride (3.00-4.29 mM), and ethylenediaminetetraacetic acid (0.60-0.86 mM) to partially degrade their crosslinked structure for 3 days, followed by culture in a normal medium until day 21. Scanning electron microscope micrographs demonstrated a loose hydrogel network with an enhanced pore size in the SCB-treated microbeads. Increasing the concentration of SCB in the treatment medium reduced the calcium content of the microbeads via a Na+ /Ca2+ exchange process and improved the water absorption of the microbeads, resulting in a higher swelling ratio. All the tested SCB concentrations were non-cytotoxic. Increases in aggrecan and type II collagen gene expression and their corresponding extracellular matrix accumulation, glycosaminoglycans, and type II collagen were vividly detected in the TGF-ß3-containing microbeads with increasing SCB concentrations in the treatment medium. Our findings highlighted that the combination of SCB treatment and TGF-ß3 incorporation in the chondrocyte-laden microbeads is a promising strategy for enhancing cartilage regeneration, which may contribute to a versatile application in cell delivery and tissue engineering.

Optimization of agarose-alginate hydrogel bead components for encapsulation and transportation of stem cells.

Hydrogels have potential uses in various biological applications because of their unique characteristics. Fine-tuning of agarose-alginate (Ag-Al) hydrogel components improves the mechanical characteristics of the final construct for cell encapsulation and transportation. Formulation of suitable dissolving agents may enable the release of encapsulated cells for further applications in laboratory or clinical settings. Objectives: This study aimed at optimizing the composition of Ag-Al hydrogel beads and their dissolving agents for potential use in the transportation of stem cells. Methods: Various agarose, alginate, and CaCl2 concentrations were tested to construct hydrogel beads. The degradation rate and swelling ratio of each hydrogel sample were recorded. The optimized Ag-Al hydrogels were used for encapsulation of stem cells from human exfoliated deciduous teeth (SHED). Optimization of dissolving agents was performed and tested with the hydrogel-encapsulated cells. Data were statistically analyzed in SPSS. Results: The selected concentration of Ag-Al hydrogels components was successfully demonstrated to encapsulate SHED, which remained viable until day 10. An average of 2 min was required for degradation of the hydrogel with encapsulated SHED by a dissolving agent consisting of 100 mM sodium citrate and 100 mM EDTA. The cell viability of SHED released after day 10 of encapsulation was 29.1%. Conclusion: Alteration of Ag-Al components has considerable influence on the mechanical properties of the constructed hydrogel. The feasibility of performing the optimized cell encapsulation protocol, as well as the dissolving step, may provide a useful guide for the transportation of viable cells between countries, for medical research.

Improvement of Lutein Production in Auxenochlorella protothecoides Using Its Genome-Scale Metabolic Model and a System-Oriented Approach.

Lutein is a valuable metabolite widely used in the food, pharmaceutical, cosmetic, and aquaculture industries. Marigold flowers are the most common source of commercial lutein, but cultivation area, weather conditions, and high manpower costs are among the disadvantages of lutein production from marigold flowers. Microalgae are an excellent alternative to plant sources of lutein as they do not have the limitations of plant extraction. Auxenochlorella protothecoides is a promising candidate for commercial production of lutein. In the present research, a genome-scale metabolic model was applied to introduce some strategies to improve lutein production in A. protothecoides. The effective reactions to improve lutein production were determined based on analysis of multiple optimal solutions. The enzymatic regulators of candidate reactions were identified using the BRENDA database. The effect of 13 activators was investigated experimentally. Our results showed that sodium citrate has the greatest effect on lutein production, so it was introduced as the most effective compound for increasing lutein production by A. protothecoides.

Evaluation of sodium citrate anticoagulant for the Resolution of edta-dependent pseudo Thrombocytopenia.

BACKGROUND: EDTA-dependent pseudo thrombocytopenia (EDTA-PTCP) refers to a falsely low platelet count occurring in the presence of ethylene diamine tetra-acetic acid (EDTA) anticoagulant during blood sample collection, which results in the formation of platelet clumps in vitro. This phenomenon has significant clinical implications, including unnecessary administration of platelets. Our study aims to evaluate the efficacy of sodium citrate anticoagulant for the resolution of EDTAPTCP. METHODS: This retrospective study was conducted in the haematology laboratory of Shifa International Hospital (SIH), Pakistan. Patients with pseudo thrombocytopenia (i.e. platelet count less than 150,000/ul with platelet clumps seen on peripheral smear) were included in this study if they had blood samples drawn in both EDTA and sodium citrate tubes less than 48 hours apart. Data was analyzed using IBM® SPSS Software Version 22. RESULTS: A total of 151 study participants were included in this study. The mean age was 48.95±20.69 years and the majority were female (52.3%). Wilcoxon signed-rank test showed that there was a statistically significant difference in platelet count measured in both tubes (Z = -3.223, p=0.001). Overall, blood samples processed in sodium citrate tubes showed lower platelet count than EDTA samples. Sodium citrate anticoagulant was able to correct EDTA-PTCP in 47 (31.1%) of the cases. CONCLUSIONS: Sodium citrate anticoagulant was only able to resolve one-third of our EDTA-PTCP cases. Our findings do not support the use of sodium citrate as a suitable alternative for correction of EDTA-PTCP.

Influence of blood sample storage and different types of anticoagulants on results of NK cytotoxicity tests.

Introduction: Natural killer (NK) cells are important players in the human immune response. Impaired NK function may lead to serious, life-threatening conditions. Defects may be consequences of genetic mutations or results of secondary factors such as infections, malignancies and autoimmune diseases. The cytotoxicity test is very useful, but its accessibility is limited to special immunological laboratories. Blood samples are often transported to remote centers, which takes time and requires special conditions.The aim of this study was to compare cytotoxicity assay results between samples preserved with three different anticoagulants to standardize the diagnostic procedure. Material and methods: Peripheral blood from healthy donors was taken with three anticoagulants: heparin, K2EDTA and citrate. Peripheral blood mononuclear cells (PBMC) were isolated and tested directly after blood drawing and after 24-hour storage. Cytotoxic abilities of NK cells were tested in 4 h co-culture with K562. NK cytotoxicity was measured by flow cytometry. Results: In most cases of analyzed healthy donors, cytotoxicity results were similar regardless of type of anticoagulant. However, the highest mean values were obtained in samples with citrate. There was a significant decrease in cytotoxicity after 24 hours of storage of the whole blood at ambient temperature. The mean drop in cytotoxicity results was substantial for all anticoagulants: 76% for heparin, 67% for citrate and 70% for EDTA. Conclusions: Results of spontaneous NK cytotoxicity seem to be affected by the anticoagulants used for blood protection. Commercial instant cytotoxicity testing and delayed analysis after blood storage gave the highest results in blood with sodium citrate.