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Objectives: It is essential to obtain homogeneous drug mixtures, especially when only a fraction of the prepared dose is to be administered. This study aimed to validate a manual mixing method for guaranteeing homogeneity., Methods: One operator tested six standardised manual mixing techniques (one, five and 10 inversions, and one, five and 10 bottoms-up agitations) six times each for preparations in bags and syringes. The mixing step was reproduced experimentally by adding a small volume of analyte (0.8 mL for syringes and 6 mL for bags) to a large volume of matrix (50 mL for syringes and 300 mL for bags). Three analyte/matrix pairings were tested: water/water, water/glucose 20% and glucose 20%/water. The tracer (sodium chloride) was assayed using capillary electrophoresis. Volume measurement errors were corrected by weighing bags and syringes. In order to evaluate inter-individual variability, the 10 inversions technique was tested by 10 drug preparation technicians. Mixtures were considered acceptable if they were between 95% and 105% accurate and if the coefficient of variation was ≤5% of the average of the six repetitions., Results: Both the 10 inversions and 10 bottoms-up agitations mixing techniques ensured acceptable mixtures by the principal technician in all tested conditions. When mixing using the 10 inversions method was tested by the 10 technicians, the mixture's mean acceptability could no longer be ensured., Conclusion: Use of a standardised mixing technique did not appear to be sufficient to obtain a homogeneous mixture across technicians. Standardised guidelines for needle position, needle rinsing and speed of addition should be implemented., Competing Interests: Competing interests: None declared.

Selenium (Se) is an essential micronutrient as well as a toxic trace element in animal and human nutrition. The effects of Se in the immune system and some diseases are well documented. The objective of the present study was to examine the role of Se in reducing the hypoxia induced apoptosis in neuroblastoma cell line. Hypoxia showed an enhanced cytotoxicity, increased free radical production and apoptosis (p<0.001) which was measured in terms of DNA break down by comet assay. Hypoxia has decreased reduced Glutathione (GSH) content, Glutathione Reductase (GR), Glutathione peroxidase (GPx) and Superoxide Dismutase (SOD) activities as compared to control cells. During hypoxic condition the expression of cytochrome C, pro and active caspase-3 levels were enhanced significantly followed by nonsignificant upregulation of Bcl-2. But, the Se supplementation inhibited the cytotoxicity, free radical generation and stabilized the HIF-1alpha accumulation in cells under hypoxia. The GSH content, GR, GPx and SOD activities increased significantly in Se-treated hypoxic cells, as compared to control. Further there was an appreciable inhibition of apoptosis by upregulation of Bcl-2 proteins, in the presence of Se under hypoxia. Selenium supplementation to cells significantly inhibited the hypoxia induced DNA fragmentation and restored the antioxidant status back to control levels. This study suggests that Se supplementation prevented the cells from hypoxia induced apoptosis by triggering upregulation of Bcl-2 protein and reducing the oxidative stress.

The present study reports the cytoprotective and antioxidant properties of alcoholic leaf extract of seabuckthorn (SBT) against hypoxia induced oxidative stress in C-6 glioma cells. Exposure of cells to hypoxia for 12 h resulted in a significant increase in cytotoxicity and decrease in mitochondrial transmembrane potential compared to the controls. Further an appreciable increase in nitric oxide and reactive oxygen species (ROS) production was noted which in turn was responsible for fall in intracellular antioxidant levels and GSH/GSSG ratio. There was a significant increase in DNA damage during hypoxia as revealed by comet assay. Pretreatment of cells with alcoholic leaf extract of SBT at 200 mug/ml significantly inhibited cytotoxicity, ROS production and maintained antioxidant levels similar to that of control cells. Further, the leaf extract restored the mitochondrial integrity and prevented the DNA damage induced by hypoxia. These results indicate that the leaf extract of SBT has strong antioxidant and cytoprotective activity against hypoxia induced oxidative injury.

Objective: To evaluate the effect of oral administration of Kombucha tea (K-tea) on lead induced oxidative stress., Methods: Sprague Dawley rats were administered 1 mL of 3.8% lead acetate solution daily alone or in combination with K-tea orally for 45 d, and the antioxidant status and lipid peroxidation were evaluated., Results: Oral administration of lead acetate to rats enhanced lipid peroxidation and release of creatine phosphokinase and decreased levels of reduced glutathione (GSH) and antioxidant enzymes (superoxide dismutase, SOD and glutathione peroxidase, GPx). Lead treatment did not alter humoral immunity, but inhibited DTH response when compared to the control. Lead administration also increased DNA fragmentation in liver. Oral administration of Kombucha tea to rats exposed to lead decreased lipid peroxidation and DNA damage with a concomitant increase in the reduced glutathione level and GPx activity. Kombucha tea supplementation relieved the lead induced immunosuppression to appreciable levels., Conclusion: The results suggest that K-tea has potent antioxidant and immunomodulating properties.

At high altitudes, the reactive oxygen species are continuously generated as a consequence of low oxygen partial pressure (hypoxia), which causes tissue damage. The body's defence system to combat the oxidative stress (e.g., anti-oxidant enzymes, free radical scavengers such as vitamin C, vitamin E, beta-carotene, reduced glutathione and minerals such as selenium, etc.) may diminish. In the present study, the antioxidant effect of selenium (Se) in reducing the hypoxia-induced oxidative stress was evaluated by exposing male albino rats to hypoxic stress in a decompression chamber. Exposure to hypoxia resulted in an increase in malondialdehyde (MDA) levels in plasma and tissues and a concurrent decrease in blood glutathione (GSH), glutathione peroxidase (GPx), plasma protein and plasma selenium content when compared with controls. Haemoglobin concentration (Hb%), red blood corpuscles (RBC) and white blood corpuscles (WBC) count were also increased in the hypoxia-exposed group. Selenium supplementation to animals reversed the trend. There was a significant decrease (P < 0.001) in MDA and subsequent increase in plasma and tissue GSH levels. Similarly the blood and tissue GPx and plasma protein also increased significantly in the Se supplemented animals compared with control animals. The Hb%, RBC and WBC counts showed no significant difference between Se-fed and control rats. These results suggest that selenium may help in reducing the lipid peroxidation during hypoxia.

The fruits extracts of Emblica officinalis (Amla) has been reported to have strong anti-oxidant properties. There is a paucity of studies on the immunomodulatory properties of fruit extracts of Amla in immuno-compromised states, with the emphasis on lymphocytes. Therefore, the aim of the study was to determine the anti-oxidant and immunomodulatory properties of Amla using chromium (VI) as an immunosuppressive agent. Chromium (Cr) treatment results in enhanced cytotoxicity, free radical production, lipid peroxidation and decreased glutathione peroxidase (GPx) activity and diminished glutathione (GSH) levels. There was a significant inhibition of both lipopolysaccharide and concanavalin-A-stimulated lymphocyte proliferation. Chromium also inhibited Con A stimulated interleukin-2 and gamma-interferon production significantly. Further, there was enhanced apoptosis and DNA fragmentation in the presence of Cr. Amla significantly inhibited Cr-induced free radical production and restored the anti-oxidant status back to control level. Amla also inhibited apoptosis and DNA fragmentation induced by Cr. Interestingly, Amla relieved the immunosuppressive effects of Cr on lymphocyte proliferation and even restored the IL-2 and gamma-IFN production considerably.

Hypoxia is known to induce oxidative stress in organisms leading to tissue injury. In the present study beta-carotene (BC) given at 10 mg/kg body weight (BW) in reducing the oxidative stress induced by hypoxia was evaluated on male albino rats. Hypoxia exposure caused an increase in malondialdehyde (MDA) levels in plasma and tissues, a concurrent decrease in blood glutathione (GSH), glutathione peroxidase (GPx), plasma protein and plasma BC content. Hemoglobin concentration, Red blood corpuscles (RBC) and White blood corpuscles (WBC) count were also increased under hypoxia. BC supplementation reversed the trend, inducing a significant decrease (P<0.05) in MDA and subsequent increase in plasma and tissue GSH levels in animals exposed to hypoxia. Blood GPx and plasma protein also increased significantly in BC supplemented animals. BC supplementation did not alter the changes in Hb concentration, RBC and WBC count. BC has potent antioxidant activities in reducing the oxidative stress induced by hypobaric hypoxia.

Objective: The objective of the study was to evaluate toxicity, anti-stress activity and hepato-protective properties of Kombucha tea., Method: Kombucha tea was fed orally for 15 days using three different doses i.e. normal dose, five and ten times the dose. Rats were then sacrificed and various biochemical, and histological parameters were estimated. Anti-stress activity was evaluated either by 1) by exposing animals to cold and hypoxia and estimating the levels of malondialdehyde and reduced glutathione in plasma/blood or 2) by subjecting the animals to restraint stress and recording faecal output. Hepato-toxicity was induced by challenging the animals to an acute dose of paracetamol (1 gm/kg) orally and determining the plasma levels of SGPT, SGOT and MDA., Results: The effect of oral administration of different doses of K-tea to albino rats was examined and the results indicate that K-tea has no significant toxicity as revealed by various biochemical and histopathological parameters. K-tea has been found to prevent lipid peroxidation and fall in reduced glutathione level when rats were exposed to cold and hypoxia in simulated chamber. Further, K-tea has also been found to decrease the Wrap-restraint faecal pellet output in rats. K-tea has also been found to decrease paracetamol induced hepatotoxicity significantly., Conclusion: The study shows that K-tea has anti-stress and hepato-protective activities.

The effect of Kombucha tea (KT) on oxidative stress induced changes in rats subjected to chromate treatment are reported. KT feeding alone did not show any significant change in malondialdehyde (MDA) and reduced glutathione (GSH) levels, but did enhance humoral response and delayed type of hypersensitivity (DTH) response appreciably over control animals. Chromate treatment significantly enhanced plasma and tissue MDA levels, decreased DTH response considerably, enhanced glutathione peroxidase and catalase activities; however, no change in GSH, superoxide dismutase and antibody titres was noticed. KT feeding completely reversed the chromate-induced changes. These results show that Kombucha tea has potent anti-oxidant and immunopotentiating activities.

Gamma interferon (IFN-gamma)-secreting CD4+ T cells have long been established as an essential component of the protective immune response against Mycobacterium tuberculosis. It is now becoming evident from studies with the murine model of tuberculosis that an important role also exists for major histocompatibility complex (MHC) class I-restricted CD8+ T cells. These cells are capable of acting as both IFN-gamma secretors and cytotoxic T lymphocyte (CTL) effectors; however, their exact role in immunity against tuberculosis remains unclear. This study demonstrates the presence of Mycobacterium bovis BCG-reactive CD8+ T cells in healthy BCG-vaccinated donors and that these CD8+ T cells are potent cytokine producers as well as cytotoxic effector cells. Using FACScan analysis, we have shown that restimulation with live M. bovis BCG induced more CD8+-T-cell activation than the soluble antigen purified protein derivative and that these cells are actively producing the type 1 cytokines IFN-gamma and tumor necrosis factor alpha (TNF-alpha). These CD8+ T cells also contain the cytolytic granule perforin and are capable of acting as potent CTLs against M. bovis BCG-infected macrophages. The mycobacterial antigens 85A and B (Ag85A and Ag85B, respectively), and to a lesser extent the 19- and 38-kDa proteins, are major antigenic targets for these mycobacterium-specific CD8+ T cells, while whole-M. bovis BCG activated effector cells from these BCG-vaccinated donors, as expected, failed to recognize the 6-kDa ESAT-6 protein. The use of metabolic inhibitors and blocking antibodies revealed that the CD8+ T cells recognize antigen processed and presented via the classical MHC class I pathway. These data suggest that CD8+ T cells may play a critical role in the human immune response to tuberculosis infection.

The effect of exposure to hypoxia on macrophage activity, lymphocyte function and oxidative stress was investigated. Hypoxia enhanced peritoneal macrophage activity as revealed by enhanced phagocytosis and free radical production. There was no significant change in antibody titres to sheep red blood cells in either serum or spleen during hypoxia. However, there was a considerable reduction in the delayed-type hypersensitivity response to sheep red blood cells, indicating the impairment of T-cell activity. Hypoxia decreased the blood glutathione (reduced) level and increased plasma malondialdehyde by a factor of about 2. It is therefore speculated that hypoxia imposes an oxidative stress leading to decreased T-cell acivity.