The state-of-the-art determination of urinary nucleosides using chromatographic techniques “hyphenated” with advanced bioinformatic methods

Over the last decade metabolomics has gained increasing popularity and significance in life sciences. Together with genomics, transcriptomics and proteomics, metabolomics provides additional information on specific reactions occurring in humans, allowing us to understand some of the metabolic pathways in pathological processes. Abnormal levels of such metabolites as nucleosides in the urine of cancer patients (abnormal in relation to the levels observed in healthy volunteers) seem to be an original potential diagnostic marker of carcinogenesis. However, the expectations regarding the diagnostic value of nucleosides may only be justified once an appropriate analytical procedure has been applied for their determination. The achievement of good specificity, sensitivity and reproducibility of the analysis depends on the right choice of the phases (e.g. sample pretreatment procedure), the analytical technique and the bioinformatic approach. Improving the techniques and methods applied implies greater interest in exploration of reliable diagnostic markers. This review covers the last 11 years of determination of urinary nucleosides conducted with the use of high-performance liquid chromatography in conjunction with various types of detection, sample pretreatment methods as well as bioinformatic data processing procedures

Subcellular optogenetic inhibition of G proteins generates signaling gradients and cell migration

Cells sense gradients of extracellular cues and generate polarized responses such as cell migration and neurite initiation. There is static information on the intracellular signaling molecules involved in these responses, but how they dynamically orchestrate polarized cell behaviors is not well understood. A limitation has been the lack of methods to exert spatial and temporal control over specific signaling molecules inside a living cell. Here we introduce optogenetic tools that act downstream of native G protein–coupled receptor (GPCRs) and provide direct control over the activity of endogenous heterotrimeric G protein subunits. Light-triggered recruitment of a truncated regulator of G protein signaling (RGS) protein or a Gβγ-sequestering domain to a selected region on the plasma membrane results in localized inhibition of G protein signaling. In immune cells exposed to spatially uniform chemoattractants, these optogenetic tools allow us to create reversible gradients of signaling activity. Migratory responses generated by this approach show that a gradient of active G protein αi and βγ subunits is sufficient to generate directed cell migration. They also provide the most direct evidence so for a global inhibition pathway triggered by Gi signaling in directional sensing and adaptation. These optogenetic tools can be applied to interrogate the mechanistic basis of other GPCR-modulated cellular functions

An Ion-Exchange Atomic-Absorption Method for the Measurement of Ionized Calcium and Magnesium At Micromolar Concentrations

A quantitative ion-exchange/atomic absorption method is described for measuring the concentration of free (hydrated) calcium and magnesium in solution at micromolar levels. Sample solutions are pumped through a micro-column of strong acid-type cation-exchange resin until equilibrium has been achieved between resin and solution. After removal of interstitial solution by first air, then water, the sorbed metal ion is eluted from the resin with nitric acid directly into an atomic absorption spectrophotometer. In a 0.1 M 1:1 electrolyte, here KNO3, the amount of metal ion sorbed on the resin is directly proportional to the free metal ion concentration in solution over a concentration range of 1.25 to 5 x 10(-5) mol/L (12-50 mumol/L). Selectivities for free calcium and magnesium in the presence of complexing ligands such as citrate and phosphate compare well with calculated values

An Ion-Exchange Atomic-Absorption Method for the Determination of Ionized Calcium At Millimolar Levels

A quantitative ion exchange/atomic absorption method is described for measuring the fraction of free calcium in solution at millimolar levels. Sample solutions are equilibrated with a micro-column of strong-acid type cation-exchange resin under electrolyte concentrations (0.75 mol/L NaNO3) high enough to provide conditions where the calcium sorbed on the resin is proportional to the free calcium in solution. After a water wash, the sorbed calcium is eluted from the resin with nitric acid directly into an atomic absorption spectrophotometer. Free calcium levels were estimated using calibration curves obtained over the range 0-5 mM calcium with a precision of +/- 0.05 mM. The effect of magnesium, at concentrations twice that of calcium, on measured calcium levels is negligible. The method is pH insensitive over the range 5-7. Selectivity for free calcium in the presence of complexing ligands such as citrate and phosphate is high. The method shows promise for the determination of ionized calcium in biological systems

Determination of Physicochemical and Functional Properties of Coconut Oil by Incorporating Bioactive Compounds in Selected Spices

Lipid oxidation has been identified as a major deterioration process of vegetable oils, which leads to the production of primary and secondary oxidative compounds that are harmful to human health. Oleoresins of ginger, garlic, nutmeg, pepper, cloves, and cinnamon were extracted and incorporated into coconut oil, and change occurrence on physicochemical properties, thermal stability, shelf life, and antioxidant activity was monitored against the same properties of pure coconut oil. Lipid oxidation was assessed in terms of the free fatty acid level and peroxide value. For the comparison purpose, another oil sample was prepared by incorporating vitamin E too. Results revealed that both peroxide value and FFA of pure and flavored coconut oil samples after a one-week storage period were 3.989±0.006 and 3.626±0.002 mEq/kg and 0.646±0.001 and 0.604±0.002 (%), respectively. Saponification value, iodine value, smoke point, and the flashpoint of flavored oil were decreased while increasing the viscosity during storage. The highest phenolic content and DPPH free radical scavenging activity were found in flavored coconut oil. Since spices containing antioxidants, the thermal stability of flavored oil was better than that of pure coconut oil. Both oleoresins and vitamin E-incorporated samples showed the same pattern of increment of FFA and peroxide value during storage; however, those increments were slower than those of pure coconut oil

Comprehensive Study on the Acrylamide Content of High Thermally Processed Foods

Acrylamide (AA) formation in starch-based processed foods at elevated temperatures is a serious health issue as it is a toxic and carcinogenic substance. However, the formation of more AA entangles with modern-day fast food industries, and a considerable amount of this ingredient is being consumed by fast food eaters inadvertently throughout the world. This article reviews the factors responsible for AA formation pathways, investigation techniques of AA, toxicity, and health-related issues followed by mitigation methods that have been studied in the past few decades comprehensively. Predominantly, AA and hydroxymethylfurfural (HMF) are produced via the Maillard reaction and can be highlighted as the major heat-induced toxins formulated in bread and bakery products. Epidemiological studies have shown that there is a strong relationship between AA accumulation in the body and the increased risk of cancers. The scientific community is still in a dearth of technology in producing AA-free starch-protein-fat-based thermally processed food products. Therefore, this paper may facilitate the food scientists to their endeavor in developing mitigation techniques pertaining to the formation of AA and HMF in baked foods in the future

Chronic Kidney Disease of Unknown Etiology (CKDu) in Sri Lanka: Hematological Changes and Pro-Inflammation Suggest Likely Predictors of Advance Disease, as Renal Outcomes Show Prevalent Normoalbuminuria

CKDu needs to be characterized in fundamental areas to improve etiological understanding and disease management. In a cross-sectional study, blood cell profile and plasma inflammatory cytokines were followed by automated analysis and sandwich ELISA, respectively. Disease development stages and proteinuria were ascertained by eGFR and UACR. Comparison among control and stages (ANOVA/Dunnett’s MRT) revealed time-specific changes (p < 0.05), including decreased erythrocytes (G5) and hematocrit (G5), and increased MCHC (G3b, G4), MCV (G5), and MCH (G5). CKDu decreased (p < 0.05) lymphocytes (G3b, G4, G5), monocytes (G3b), MPV (G3b, G4, G5), and plateletcrit (G3b, G4), and increased basophils (G3a, G3b, G4), N/L (G4) and PLR (G4–G5). MCHC and aforesaid leukocyte variables were in correlation (rho > ±0.03, p < 0.05, Pearson’s test) with disease development. MCP-1 and IL-6 spiked (p > 0.05) at G3b. Multivariate analyses confirmed that MCP-1, lymphocytes, and BMI were related to renal dysfunction, pointing to inflammation, compromised immunity, and muscle wasting as CKDu effects. Nonproteinuric CKDu was prevalent (23.2–35.6% of total CKDu) with (p < 0.05) elevated basophils (G3a), N/L (G4), and depleted lymphocytes (G4). In both forms, G1–G2 were unaffected, and the earliest change was G3a basophils. Results suggest that MCP-1, lymphocyte count, N/L, and PLR may verify the stage and predict impending ESRD in advance proteinuric CKDu