Potential Deleterious Effects of L-Citrulline Supplementation in Isoproterenol-Induced Myocardial Infarction: Focus on Nitrosative Stress

L-Citrulline shows potential activity as a supplement to prevent myocardial infarction through vasodilative and possible antioxidative effects but may be deleterious by causing nitrosative stress. This study determined the potentially deleterious effects of L-citrulline supplementation in isoproterenol-induced myocardial infarction with a focus on nitrosative stress. L-Citrulline supplementation was given orally at dosages of 300 or 600mg/kg body weight daily for 6 days. Myocardial infarction was induced in Wistar rats via subcutaneous injection of isoproterenol (85 mg/kg body weight (BW)) on day 4 and 5. Blood pressure was measured at the end of the study (day 6) and rats were sacrificed to collect heart tissue samples for a histopathological evaluation. The histopathological evaluation was done using hematoxylin and eosin staining for the myocardial damage evaluation and immunohistochemical (IHC) staining of arginase-2, inducible nitric oxide synthase (iNOS), and 3-nitrotyrosine to evaluate nitrosative stress. L-Citrulline supplementation failed to show a significant protective effect on blood pressure and exacerbated the decrease of diastolic blood pressure. Both low and high dose L-citrulline supplementation had a significant protective effect on myocardial damage compared to the isoproterenol group (p<0.01). L-Citrulline also caused increased nitrosative stress as shown by increased expression of arginase-2 and 3-nitrotyrosine on IHC staining but tended to show an ameliorative effect on iNOS expression. A significant increase in arginase-2 expression was detected between the high dose group and the other groups (p<0.01 vs. normal and isoproterenol groups; p<0.05 vs. low dose group). L-Citrulline supplementation increased 3-nitrotyrosine expression in a dose-dependent manner, which was significantly different compared to the normal group (low dose: p<0.013; high dose: p<0.003). L-Citrulline increased the production of nitrosative stress but resulted in less myocardial damage through its other effects

FTIR Method for Peptide Content Estimation and Degradation Kinetic Study of Canarium Nut Protein

Quantitative analysis of bioactive peptide mostly conducted by measuring the activity. While the determination of peptide content in natural sources has been conducted using various instruments, vibrational spectroscopy remains underutilized. Here, we attempted new developed method in peptide quantification and degradation kinetic analysis using Fourier-Transform Infrared Spectroscopy. Bovine Serum Albumin was used as standard protein in method development and validation. Peptide content was estimated by converting peak area to concentration. The method was used to estimate peptide content in Canarium nut protein and its hydrolysates, which potentially hold biological activity. Kinetic study was conducted with microwave as an accelerator for hydrolysis, an apparatus rarely used in peptide study. Amide I band on wavenumber range of 1724.05-1619.91 cm-1 was selected for analysis, considering its selectivity and linearity. The method also met other validation requirement, including accuracy and precision. When applied in quantitative analysis, the method was able to calculate peptide content decrease in Canarium nut protein after hydrolysis using papain (38.24%), pepsin (33.67%) and alkaline reagent (28.53%). In kinetic study, microwave-assisted peptide degradation exhibited logarithmic profile with the equation of y=-0.148ln(x)+0.9591 and R² value of 0.963. Based on these results, FTIR is useful in estimating peptide content and in analyzing degradation kinetic profile.

The Effect of Brazilin from Caesalpinia sappan on Cell Cycle and Modulation and Cell Senescence in T47D cells

Ethanolic extract and brazilein-containing fraction of Caesalpinia sappan L., has been reported to inhibit cell proliferation in T47D (ER+ PR+/- cell, Luminal A subtype model). The Luminal A subtype is the most common subtype of breast cancer in Indonesian women. In this study, we explored the activity of the reduced form of brazilein, i.e. brazilin, in T47D cells proliferation and the mechanism that involved. The cytotoxicity activity of brazilin was observed using MTT assay. While the cell cycle modulation analysis was done by using flowcytometry, and the senescence assay was observed using S-A-β-galactosidase assay. The results showed that brazilin inhibited cell growth in a dose-dependent manner with an IC50 value of 50μM (or 14.3μg/mL). That was higher than a brazilein-containing fraction, which was reported previously by our group to have an IC50 value of 68μg/mL against the same cell. Cell cycle analysis showed that cells treated with brazilin were accumulated at the G2/M phase in a dose-dependent manner. Furthermore, cells treated with a combination of brazilin and doxorubicin was accumulated at the G2/M phase and sub G1 phase. Cells accumulation at sub G1 phase indicates that the cells undergo apoptosis. Our data of S-A-β-galactosidase assay showed that cells treated with 1/4IC50, 1/2IC50, and IC50 brazilin had lower senescent cells compared to the untreated cells. The morphology of cells treated with IC50 (50μM) brazilin changed. The cells shape became rounded, cells were shrinkage and detached from the well plate, indicating that cells may undergo apoptosis. These results suggested that brazilin was cytotoxic towards T47D cells and its combination with dox potentially induced apoptosis and decreased cell senescence. The ability of brazilin to decrease cell senescence provides new insight of utilization of C. sappan or its constituents, particularly brazilin, as anti-ageing

QuEChERS and C18 as solid phase extraction sorbent - ultra-high performance liquid chromatography -ultraviolet-visible method for determination of parabens in cosmetics products

Concerns are growing about human exposure to endocrine-disrupting chemicals (EDCs), especially during the preadolescent development stage. Parabens are prevalent EDCs widely used as additives in cosmetics. So, the determination of parabens in such products is important. In this study, we developed a reliable and sensitive method to determine simultaneously nine common parabens (methylparaben, ethylparaben, phenylparaben, benzylparaben, penthylparaben, and two groups of isomeric compounds include propylparaben, isopropyl paraben, and butylparaben, isobutylparaben) in cosmetics products. The QuEChERS and solid-phase extraction techniques are used for extraction parabens from non-surfactant cosmetics (perfume, mouth wash solution) and surfactant cosmetics (shampoo, cream, gel), respectively and quantified by using ultra-performance liquid chromatography coupled with the ultraviolet-visible detector. All nine compounds showed good linearity with regression coefficients predominantly above 0.990. The LOD and LOQ of parabens were 0.07 µg/mL; 0.2 µg/mL, respectively. The recoveries ranged from 80 to 110% with the relative standard deviations below 8%. The developed method was successfully applied to determine parabens in various commercial cosmetic products from a local supermarket and the total parabens concentrations are in a wide-ranged from 2.0 to 1270 mg/kg

The effect of combination of active fraction Andrographis paniculata (Burm.f) Ness and Centella asiatica (i) Urban on the alpha glucocidase inhibitor and antioxidant activities

The Andrographis panicullata and Centella asistica extract have been reported that had a anti-diabetic effect. However, the specific mechanism and the effect combination of both were not yet reported. This study was purposed to determine the potency of extract, fractions and the combination of Andrographis panicullata (AP) and Centella asistica (CA) active fraction to inhibit alpha glucosidase enzymeand its ability to reduce DPPH radical.AP and CA were extracted using 50% ethanol then fractionated with solvents under different polarity levels. The inhibiting activity to alpha glucosidase enzyme and antioxidant activity of each fractions was tested. The most active fractions from AP and CA were then combined and re-tested for activity. The results result reported that both of AP and CA had inhibition of alpha glucosidase activity and antioxidant activity. Based on calculation combination index (CI) of of active fraction of AP and CA showing in alpha glucocidase activity had a antagonist action and antioxidant had a sinergic action. Therefore, combination of AP and CA not has not recomended for alpha glucocidase inhibitor but the combination has ability to reduce DPPH radical

Effect of Growth Factor In Callus Induction and Bioactive Compounds In Seed Explant of Kaffir Lime (Citrus hystrix DC.)

Our previous study showed that kaffir lime leaf extracts may have anti-cancer properties. However, production of  bioactive compounds is affected by environmental factors. Here, we present a method to control environmental conditions using in vitro culture techniques such as callus induction. Calluses were induced from seed embryo explants of kaffir lime on MS medium with combinations of 2,4-D and BAP at concentrations 1:0.5; 1:1; and 2:1, respectively. Fourty and 60 days-old calluses were extracted using chloroform and ethyl acetate and analyzed by GC-MS. Results showed all combinations of 2,4-D and BAP were able to induce callogenesis from seed embryo explants of kaffir lime with no significant differences of callus initiation time, biomass, morphology and growth rates. However differences were detected in the bioactive compound profiles. In kaffir lime callus, both fatty acids and secondary metabolites were detected. Specifically, in 40 days-old calluses (exponential growth phase) we detected α-pinene and 1.8–cineole in plants treated with 2,4-D: BAP at concentration 1:0.5 and 2:1. In 60 days-old calluses (stationary phase) we detected a number of compounds in plants treated with 2,4-D:BAP at concentrations of 1:0.5 and 2:1, including caryophyllene, linoleoyl chloride, thiogeraniol, stigmasterol, clianosterol, citronellal, neo-isopulegol, citronellol, geraniol, eugenol, cyclopropane, pristane, elemol and farneso

Antidiabetic Activity of Okra Fruit (Abelmoschus esculentus (L) Moench) Extract and Fractions in Two Conditions of Diabetic Rats

Okra (Abelmoschus esculentus (L) Moench) fruit is empirically used in type 2 diabetes mellitus treatment. This research aims to know the antihyperglycemic activity of okra fruit extract and fractions in streptozotocin-nicotinamide (STZ-NA) induced as well as in insulin resistance diabetic rats, the effect on pancreatic cells regeneration, and the effect on immunohistochemical expression of glucose transporter-4. This study used a group of 35 male Wistar rats for STZ-NA induced diabetic model and another group of 35 rats for insulin resistance diabetic model. Gliclazide (0.72mg/kg BW) and metformin (45mg/kg BW) were used as drug control in STZ-NA induced and insulin resistance diabetes, respectively. Okra fruit ethanol extract, n-hexane, ethyl acetate, and water fraction were orally administered with dose of 200; 107; 6 and 86mg/kg BW, respectively, for 28 days after diabetic condition was obtained. Blood glucose level was measured every week. Hematoxylin-eosin staining was used to evaluate the pancreatic cells regeneration, while immunohistochemistry was used to evaluate the expression of glucose transporter-4 in muscle membrane cells, at the end of the treatment. The results revealed that ethyl acetate fraction was the most effective in lowering blood glucose level in both condition of diabetes. Ethyl acetate fraction decreased the necrosis of pancreatic cells in STZ-NA induced diabetic rats and increased the expression of glucose transporter-4 in muscle cell of insulin resistance diabetic rats

Cytotoxic Potential of Arthrospira platensis Extract on Cervical Cancer Cells Line Hela: Study on Antiproliferative, Cell Cycle, Apoptosis Induction and Anti Metastasis

Cervical cancer can be treated conventionally with chemotherapy agents, but its use has side effects and complications in the form of damage to normal cells. This study aims to determine the potential of A. platensis as an alternative anticancer agent that is selective towards normal cells. Based on TLC analysis, A. platensis contains antioxidant compounds such as β-carotene, flavonoids, and terpenoids which are able to inhibit proliferation and trigger apoptosis of cancer cells. The study was conducted using cervical cancer cells HeLa and normal cells HDFa. A. platensis macerated with 96% ethanol at a ratio of 1:4. Based on probit analysis, it is known that ethanol extract of A. platensis has a cytotoxic effect on HeLa cells with IC50 values of 260.444μg/mL and index selectivity towards HDFa cells of 7.931. The mechanism of cytotoxic activity of ethanol extract of A. platensis is related to its ability to extend the doubling time, increase the induction of apoptosis, and reduce the rate of cells migration. Ethanol extract of A. platensis can also increase cells accumulation in the S phase to prevent cells from entering the G2/M phase

In Vitro Antiplasmodial Activity and Cytotoxicity of Active Subfractions of Harmsiopanax aculeatus Leaves

Harmsiopanax aculeatus leaves, a medicinal plant with locally named kapur, have been used traditionally to treat malaria in Maluku, Indonesia. However, the scientific information of this plant is still limited. In our previous study, the methanol extract of this plant leaves have been proven to possess in vitro antiplasmodial activity. This study was conducted to evaluate in vitro antiplasmodial activity and cytotoxicity of  subfractions of the plant leaves. Fractionation was performed using a column chromatography with Sephadex LH-20 as the  stationary phase and methanol as the mobile phase. The subfractions obtained were then tested for in vitro antiplasmodial activity on a chloroquine-resistant FCR3 strain of Plasmodium falciparum using a visual method. Cytotoxicity was evaluated by using MTT assay. The in vitro antiplasmodial activity and cytotoxicity were expressed as IC50, calculated using probit analysis with SPSS 16 for windows. The results showed that the four subfractions tested have a high antiplasmodial activity with IC50 values of 0.09; 0.18; 0.01; and 0.77 µg.mL-1, respectively. In addition, these subfractions had IC50 values of >400 µg.mL-1 against Vero cells indicating that they were non-toxic. In conclusion, the subfractions of H. aculeatus leaves are very active and selective against P. falciparum. Further study will be conducted to isolate the active compounds

In Vitro Study: Effect of Cobalt(II) Chloride Against Dengue Virus Type 1 in Vero Cells

Dengue virus (DENV) serotypes DENV-1 to DENV-4 are enveloped viruses that belong to the genus Flavivirus of the Flaviviridae. Dengue vaccine or antiviral has not yet been clinically approved for humans, even though there have been great efforts toward this end. Antiviral activity against DENV is needed to develop to be an alternative drug for DENV virus. Cobalt(II) chloride have been used in the treatment and prevention of diseases of humans since ancient times. The aim of this study is to investigate the antiviral effects and Cytotoxicity of Cobalt(II) chloride. This compound was further investigated for its inhibitory effect on the replication of DENV-1 in Vero cells. Antiviral activity and Cytotoxicity measured by WST-1 assay. The IC50 value of the Cobalt(II) chloride for DENV-1 was 0.38 μg/ml. The cytotoxicity of Cobalt(II) chloride to Vero cell suggest that the CC50 value was 2.91 µg/ml The results of this study demonstrate the anti-dengue serotype 1 inhibitory activity of Cobalt(II) chloride was a high toxic compound