Effect of Storage Temperature on Bio efficacy of Aqueous Extract of Ganoderma lucidum

Stability testing is key requirement in the drug development process along with storage temperature which may deteriorate bio-constituents and efficacy of natural products. Aqueous extract of Ganoderma lucidum has revealed pharmacological effects against high altitude stressors and has potential for mitigating high altitude maladies. In the present study, the extract of Ganoderma lucidum was stored at different storage conditions such as room temperature, 4 °C and -20 °C for two year and qualitative and quantitative analysis of bio-constituents and bio-efficacy was carried out. No significant change was observed in any extract kept in different temperature conditions in terms of its polysaccharide, phenolic and flavonoids content. The extract kept at room temperature absorbed slight moisture in few samples but no change in overall polysaccharide, phenolic and flavonoids content was recorded. The moisture absorption problem was not observed in extracts stored at 4 °C and -20 °C. The bio-efficacy of the extract at room temperature, 4 °C or -20 °C were comparable to the freshly prepared extract and the data from the studies suggest that extract has good shelf life up to two year without loss of bio-efficacy. Overall, the extract retained its bio-efficacy for two years at different temperature storage conditions

Neuroprotective effect of peroxiredoxin 6 against hypoxia-induced retinal ganglion cell damage

Abstract Background The ability to respond to changes in the extra-intracellular environment is prerequisite for cell survival. Cellular responses to the environment include elevating defense systems, such as the antioxidant defense system. Hypoxia-evoked reactive oxygen species (ROS)-driven oxidative stress is an underlying mechanism of retinal ganglion cell (RGC) death that leads to blinding disorders. The protein peroxiredoxin 6 (PRDX6) plays a pleiotropic role in negatively regulating death signaling in response to stressors, and thereby stabilizes cellular homeostasis. Results We have shown that RGCs exposed to hypoxia (1%) or hypoxia mimetic cobalt chloride display reduced expression of PRDX6 with higher ROS expression and activation of NF-κB. These cells undergo apoptosis, while cells with over-expression of PRDX6 demonstrate resistance against hypoxia-driven RGC death. The RGCs exposed to hypoxia either with 1% oxygen or cobalt chloride (0-400 μM), revealed ~30%-70% apoptotic cell death after 48 and 72 h of exposure. Western analysis and real-time PCR showed elevated expression of PRDX6 during hypoxia at 24 h, while PRDX6 protein and mRNA expression declined from 48 h onwards following hypoxia exposure. Concomitant with this, RGCs showed increased ROS expression and activation of NF-κB with IkB phosphorylation/degradation, as examined with H2DCF-DA and transactivation assays. These hypoxia-induced adverse reactions could be reversed by over-expression of PRDX6. Conclusion Because an abundance of PRDX6 in cells was able to attenuate hypoxia-induced RGC death, the protein could possibly be developed as a novel therapeutic agent acting to postpone RGC injury and delay the progression of glaucoma and other disorders caused by the increased-ROS-generated death signaling related to hypoxia.Peer Reviewe

Survival response of hippocampal neurons under low oxygen conditions induced by Hippophae rhamnoides is associated with JAK/STAT signaling.

Janus activated kinase/signal transducers and activators of transcription (JAK/STATs) pathway are associated with various neuronal functions including cell survival and inflammation. In the present study, it is hypothesized that protective action of aqueous extract of Hippophae rhamnoides in hippocampal neurons against hypoxia is mediated via JAK/STATs. Neuronal cells exposed to hypoxia (0.5% O2) display higher reactive oxygen species with compromised antioxidant status compared to unexposed control cells. Further, these cells had elevated levels of pro-inflammatory cytokines; tumor necrosis factor α and interleukin 6 and nuclear factor κappa B. Moreover, the expression of JAK1 was found to be highly expressed with phosphorylation of STAT3 and STAT5. Cells treated with JAK1, STAT3 and STAT5 specific inhibitors resulted in more cell death compared to hypoxic cells. Treatment of cells with extract prevented oxidative stress and inflammatory response associated with hypoxia. The extract treated cells had more cell survival than hypoxic cells with induction of JAK1 and STAT5b. Cells treated with extract having suppressed JAK1 or STAT3 or STAT5 expression showed reduced cell viability than the cell treated with extract alone. Overall, the findings from these studies indicate that the aqueous extract of Hippophae rhamnoides treatment inhibited hypoxia induced oxidative stress by altering cellular JAK1, STAT3 and STAT5 levels thereby enhancing cellular survival response to hypoxia and provide a basis for possible use of aqueous extract of Hippophae rhamnoides in facilitating tolerance to hypoxia

Protective efficacy of <i>Hippophae rhamnoides</i> extract on hypoxia induced cell death.

<p>(A) effect of extract on hypoxia induced on cell death (B) effect of extract on hypoxia induced reactive oxygen species generation (c) photomicrograph of HT22 cells following hypoxia exposure with or without extract, 40x magnification (d) active caspase 3 expression in HT22 cells following hypoxia exposure with or without extract (e) lactate dehydrogenase leakage in HT22 cells following hypoxia exposure with or without extract. *p<0.05; * vs control; ^# vs hypoxia.</p

Effect of trolox and quercetin on sulfur mustard-induced cytotoxicity in human peripheral blood lymphocytes

Objective: To evaluate the protective activity of antioxidants, viz. trolox and quercetin, against sulfur mustard (SM)-induced cytotoxicity. Materials and Methods: Cytotoxicity of various concentrations (20-640 µM) of SM, in the presence or absence of 10 µM trolox or quercetin (-0.5, 0, or +0.5 h) was determined in human peripheral blood lymphocytes after 6-h exposure. Cell viability was measured by Trypan blue dye exclusion (TBDE). Further, a cytotoxic concentration of SM (80 µM) was challenged by the two antidotes (-0.5 h) and cell viability was measured by TBDE and leakage of intracellular lactate dehydrogenase (LDH). Mitochondrial integrity and peroxide levels were measured by 3-4,5-dimethyl thiazol- Z -yl)-2,5-diphenyltetrazolium bromide and 2′,7′-dichlorofluoroscin diacetate assay, respectively. Morphological changes of cells exposed to 320 µM SM (with or without antidotes) were also visualized under light microscope. Results: On the basis of TBDE , SM caused cell death of approximately 50% at 80 µM and 100% at 640 µM, respectively. Pretreatment of trolox conferred significant protection compared with quercetin. Also, pretreatment of trolox significantly reduced cell death and LDH leakage caused by 80 µM SM but did not prevent the loss of mitochondrial integrity. Trolox significantly reduced the levels of peroxides generated by SM. The better protection offered by trolox was evidenced in cell morphology studies too. Conclusion: Pretreatment (-0.5 h) of trolox afforded significant protection against SM-induced cytotoxicity in human lymphocytes. The protection was related to the antioxidant property of trolox, a water soluble analog of a-tocopherol

PCR primer sequences and annealing temperature.

<p>PCR primer sequences and annealing temperature.</p

Depicts total phenols and flavonoid content (A), antioxidant activity (B) and RP-HPLC chromatogram (C) of aqueous extract of <i>Hippophae rhamnoides</i>.

<p>Depicts total phenols and flavonoid content (A), antioxidant activity (B) and RP-HPLC chromatogram (C) of aqueous extract of <i>Hippophae rhamnoides</i>.</p

mRNA expressions of JAK1, STAT3, STAT5a, STAT5b, TNFα, IL6 and β-actin in HT22 cells following hypoxia exposure with or without extract.

<p>*p<0.05; * vs control; ^# vs hypoxia.</p

Effects of extract treatment on active caspase 3 activity (A) and lactate dehydrogenase leakage into culture medium (B) in HT22 cells following hypoxia exposure.

<p>*p<0.05; * vs control; ^# vs hypoxia.</p