Evaluation of the properties of Bungarus caeruleus venom and checking the efficacy of antivenom used in Bangladesh for its bite treatment.

As a disaster-prone country with unique geographical features, snake biting is a major public health concern in Bangladesh. The primary reasons of mortality from snakebite include late presentation to the hospital, low efficacy of antivenom, and a lack of adequate management facilities. Because snake venom characteristics vary depending on geographical location, antivenom should be manufactured from snakes native to the region in which it would be administered. Bungarus caeruleus is a highly venomous snake contributing to the major snakebite issue in Bangladesh. Therefore, the neutralization efficacy of the antivenom against B. caeruleus venom was evaluated in the current study along with the characterization of venom. For biological characterization of venom, RP-HPLC and SDS-PAGE profiling, hemolytic activity, hemorrhagic activity, phospholipases A2 (PLA2) activity, edema inducing activity and histopathological observations were carried out following standard protocol. LD50 of the venom was calculated along with neutralization potency of Incepta antivenom through probit analysis. Results showed that venom possesses phospholipase A2 activity, hemolytic activity and edema inducing activity while hemorrhagic activity was absent in the skin of envenomed mice. Histopathological alterations including necrosis, congestion and infiltrations were observed in envenomed mice organs after hematoxylin and eosin staining. Neutralization study showed that Incepta polyvalent antivenom could neutralize (potency 0.53 mg/ml) the lethal effect in in vitro study on mice. Further investigation on snakebite epidemiology and clinical observations of the envenomed patients will help in combating the snakebite problem more efficiently.

Bioremediation potential of hydrocarbon degrading bacteria: isolation, characterization, and assessment.

Oil contamination is a worldwide concern now. However, oil contaminated environment is enriched with microorganisms that can utilize petroleum oil and use hydrocarbon for their growth, nutrition and metabolic activities. In the present study, bacteria present in the oil contaminated soil were isolated by enrichment culture technique using Minimal Salt (MS) media supplemented with diesel oil and burned engine oil as a sole carbon source. The isolated bacteria were characterized by morphological and biochemical tests and identified by molecular tool through cycle sequencing method. Three isolates were morphologically characterized as gram-negative, cocci shaped and 16S rRNA sequence analysis revealed that the isolates are closely related to Pseudomonas sp., Acinetobacter sp., and Enterobacter sp. respectively. Growth condition was optimized at pH 7.0 and temperature 37 °C. All the isolates were susceptible to several antibiotics and they have no antagonistic effect with soil beneficial bacteria. Three isolates were grown in two different concentrations of diesel oil and burned engine oil (4% v/v and 8%, v/v) respectively. Study revealed that with increasing the concentration of diesel oil in the media the growth rate of all the isolates were decreased. In contrast, the growth rates of all the three isolates were increased, with increasing concentration of burned engine oil. In our study, all the isolates showed their degradation efficacy in 4% v/v diesel oil and in 8% v/v burned engine oil. So, our research clearly shows that the isolates could be potentially used for bioremediation purposes for cleaning up petroleum polluted area.

Diesel degradation efficiency of Enterobacter sp., Acinetobacter sp., and Cedecea sp. isolated from petroleum waste dumping site: a bioremediation view point.

Bioremediation through biodegradation is applied for cleaning up several environmental pollutions including petroleum oil spill containing petrol, diesel, mobil, kerosene, lubricating, etc. which have devastated several endangered terrestrial and aquatic ecosystems. Therefore, the current research was aimed to isolate and identify diesel degrading bacteria from the petroleum waste dumping site and determined their degrading efficiency. The bacterial strains were isolated through a minimum salt medium supplemented with 2% diesel as the sole carbon source. The bacteria were identified by morphological, biochemical characterization, and 16S rRNA gene sequencing. The optimized growth pattern was evaluated by utilization of a wide range of temperatures (25, 30, 35, and 40 °C) and pH (5,6,7 and 8) as well as different concentrations of diesel (2, 3, 5and 7%). Finally, the degradation rate was determined by measuring the residual diesel after 7, 14, and 21 days of incubation. The study isolated Enterobacter ludwigii, Enterobacter mori, Acinetobacter baumannii, and Cedecea davisae where all are gram-negative rod-shaped bacilli. All the bacterial strains utilized the diesel at their best at 30 °C and pH 7, among them, A. baumannii and C. davisae exhibited the best degrading efficiency at all applied concentrations. Finally, the determination of degradation rate (%) through gravimetrical analysis has confirmed the potency of A. Baumannii and C. davisae where the degradation rate was around 61 and 52% respectively after 21 days of incubation period with 10% diesel. The study concludes that all of those isolated bacterial consortiums, especially A. baumannii and C. davisae could be allocated as active agents used for bioremediation to detoxify the diesel-containing contaminated sites in a cost-effective and eco-friendly way.

A comparative study of biological potentiality and EAC cell growth inhibition activity of Phyllanthus acidus (L.) fruit pulp and seed in Bangladesh.

Medicinal plant-derived bioactive compounds have recently gained more interest in biological research as an important source of novel drug candidates. Phyllanthus acidus (L.) is a widely distributed herbal medicinal plant naturally used in Ayurvedic medicine in Bangladesh. The present study focused on exploring the biological potential as well as the inhibitory effect of EAC cell growth with a comparative analysis between Phyllanthus acidus fruit pulp and seed. Crude methanol extract of P. acidus (MEPA) fruit pulp and seed was assessed as DPPH and NO free radical scavengers. While Brine Shrimp lethality bioassay, the standard protocol of phytochemical screening and hemagglutination assay were performed successively to determine the toxic effect on normal cells, the identification of some crucial phytochemicals, and the existence of lectin protein. EAC (Ehrlich's Ascites Carcinoma) cell growth inhibition was determined by hemocytometer and morphological changes of EAC cells were observed by a fluorescence microscope using Swiss albino mice. The IC50 value of MEPA fruit pulp and seed was obtained as 57.159 µg/ml and 288.743 µg/ml respectively where minimal toxic effects on Brine Shrimp nauplii demonstrates that it is a good source of natural antioxidant compounds. Again, MEPA fruit pulp and seed-mediated effective agglutination of mouse blood erythrocyte strongly support the presence of lectin protein. Furthermore, MEPA fruit pulp and seed extract-treated EAC cells showed 65.71% and 28.57% growth inhibition respectively. The fluorescent microscopic examination of EAC cells treated with MEPA fruit pulp has shown more remarkable structural changes in the nucleus than that of seed. Based on the above findings, the present study reveals that MEPA fruit pulp can be considered as a novel biological candidate for the treatment of fatal diseases shortly.

Deciphering the molecular pathways of apoptosis using purified fractions from leaf extract of Basella alba through studying the regulation of apoptosis related genes.

Apoptosis plays a pivotal role in the exclusion of abnormal cells without any ruin of surrounding healthy cells. Generally, it occurs through an orderly and autonomously process which is controlled by proper function of various genes. Therefore, the current experiments detect the expression level/pattern of those genes to confirm the involvement of extrinsic and intrinsic pathway using Basella alba leaf (BAL). Several fractions after gel filtration chromatography of BAL extract have been pooled to evaluates its apoptosis induction potentiality on Ehrlich's Ascites Carcinoma (EAC) cells through conducting a number of bio-assays such as cell growth inhibition assay, fluorescence and optical microscopy, DNA fragmentation assay and gene expression analysis etc. The pooled fractions of BAL showed 12-56% inhibitory effect on EAC cell line at the concentration range of 25-400 µg/ml that was determined by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) assay. They also exhibited excellent cell growth inhibition at in vivo and in vitro condition when treated with 10, 20 and 40 mg/kg day. After administration of six consequent days, significant morphological features of apoptosis were observed in EAC cells under both fluorescence and optical microscope which was further supported by DNA fragmentation assay. The polymerase chain reaction amplification of bax, bcl-2 (B-cell lymphoma 2), p53, tumor necrosis factor-α, Fas, NF-kß (Nuclear factor-Kappa-B), PARP-1 (Poly (ADP-ribose) polymerase), Cyt-c cas-8, cas-9 and cas-3 revealed that the experimental sample able to induce apoptosis in both extrinsic and intrinsic pathways through altering the gene expression. The current findings suggest that sample from BAL occupy wonderful competence to induce cell apoptosis and become an ideal resource for cancer treatment.

Differential expression of Bax-Bcl-2 and PARP-1 confirms apoptosis of EAC cells in Swiss albino mice by Morus laevigata.

A safer natural alternative to treat neoplastic cells by inducing apoptosis is a prime requisite. Therefore, the current study was to evaluate the antiproliferative activity of Morus laevigata, a wild-type Mulberry species. Antioxidant and cytotoxic activity of aqueous extracts of M. laevigata leaf (MLL) and M. laevigata bark (MLB) were evaluated. The in vivo cell growth inhibition was assessed on Ehrlich's ascites carcinoma (EAC) bearing mice model. Fluorescent microscopy and expression of PARP-1, Bax, and Bcl-2 through qPCR were performed to evaluate apoptosis. MLL and MLB extracts show promising antioxidant property with an IC50 value of 186.76 µg/ml and 352.97 µg/ml, respectively, with a decent LD50 value of 99.16 µg/ml and 92.54 µg/ml for MLL and MLB extract, respectively, indicated notable cytotoxicity. Cell growth inhibition was observed using MLL and MLB extracts were 68.33% and 48.66%, respectively. The morphological alteration, DNA fragmentation, and differential expression of Bax, Bcl-2, and PARP-1 confirm the induction of the intrinsic pathway of apoptosis. PRACTICAL APPLICATIONS: Plant-based medicine always plays a tremendous role in preventing several fatal diseases like cancer. The study evaluated the anticancer activity of a wild-type mulberry. Moreover, the potent antioxidant activity of the plant makes it possible to be a great candidate for cancer remedy. Besides, the molecular expression of the genes related to apoptosis confirms the plant's bioactive compounds could be a drug lead to neoplastic cells in the future. Presences of an immense antioxidant properties urge that they can be contribute in cancer treatment through the cell death pathways.

Methanolic Bark Extract of Abroma augusta (L.) Induces Apoptosis in EAC Cells through Altered Expression of Apoptosis Regulatory Genes.

Abroma augusta (L.), one of the herbal medicinal plants, is widely used for treatment of various maladies. The present study was initiated to determine the antioxidant, hemolytic, cytotoxicity, and anticancer activities of methanolic extract from the bark of the plant. The phytochemical screening was done by analyzing different phytochemicals present in the extract. We observed the presence of alkaloids, steroids, terpenoids, flavonoids, reducing sugars, and glycosides in the bark extract which showed the highest antioxidant capacity. Antioxidant potential of the methanolic extract was evaluated in vitro by DPPH (2,2-diphenyl-1-picrylhydrazyl) scavenging assay method. This extract showed prominent scavenging activity with IC50 value of 38.65 µg/ml. The hemolytic activity of the extract was evaluated at concentrations ranging from 250 to 1000 µg/ml. It was observed that the extract induced hemolysis percentage of 9.41% to 4.1%, which implies that the extract has no potent hemolytic activity. Cytotoxicity and anticancer activities were observed on Ehrlich ascites carcinoma (EAC) cells. In addition, the bark showed promising cytotoxicity with IC50 value of 329.41 µg/ml, and the study indicated that the extract was capable of inhibiting EAC cell growth by 75.5% when administered at 100 mg/kg/day body weight intraperitoneally for five consecutive days to Swiss albino mice. Morphological change of apoptotic cell was determined by fluorescence and optical microscopy. DNA fragmentation is another marker for apoptosis, and the bark extract-treated EAC cells showed smeared and fragmented DNA bands. Apoptosis correlated well with the upregulation of p53 and Bax and also with the downregulation of NF-κB and Bcl-2. Furthermore, activity and interaction of two A. augusta compounds were tested through molecular docking simulation study. In conclusion, our results suggest that A. augusta bark has the potential to be considered as an anticancer agent.

In-vivo antiproliferative activity of Morus latifolia leaf and bark extracts against Ehrlich's ascites carcinoma.

Cancer is the second death causing disease all over the world and until today 100 different types of cancer have been identified whose treatment methods consist of serious side effects on human body. To reduce the frequency of adverse effects of cancer treatment, nowadays plant derived natural components are getting priority. The plant Morus latifolia is widely available in northern part of Bangladesh. The earlier researches suggested that popular varieties of some Morus sp. like Morus alba, Morus indica etc. have good anti-proliferative activity. Hence, this study was designed to evaluate the anti-proliferative activity of leaf and bark extracts of M. latifolia against Ehrlich's ascites carcinoma (EAC) in vivo. The leaf and bark extracts of M. latifolia were used in several bioassays including Brine shrimp lethality test, hemagglutination activity test, antioxidant activity test, and cell growth inhibition test. Besides, fluorescence microscopy was performed to study apoptotic features in EAC cells, and molecular analysis like real-time PCR were also conducted. The results of Brine shrimp lethality test, hemagglutination activity test, and antioxidant activity assay supported the cell growth inhibition capability of leaf and bark extracts which was confirmed by in vivo cell growth inhibition bioassay. Moreover, the experimental extracts were able to induce cell apoptotis through altering the expression pattern of Bcl-2 and Bax genes. All of the results of this study suggest that several noble compounds are present in M. latifolia plant extracts which are capable for healing cancer cells.

Effect of Citrus macroptera Fruit Pulp Juice on Alteration of Caspase Pathway Rendering Anti-Proliferative Activity against Ehrlich's Ascites Carcinoma in Mice.

Citrus macroptera (Rutaceae) has long been used in folk medicine in Bangladesh. Considering the folkloric context, this study was aimed to scrutinize anti-proliferative activity of C. macroptera fruit pulp juice (CMFPJ) against Ehrlich's ascites carcinoma (EAC). The anti-proliferative capacity of CMFPJ was investigated and confirmed primarily using MTT assay. In vivo anti-proliferative aptitude of CMFPJ was investigated with 25, 50, and 100 mg/kg/day intraperitoneal (i.p.) treatment. Anti-proliferative efficacy of CMFPJ was assessed based on EAC growth inhibition. CMFPJ inhibited EAC growth in vitro in a dose-dependent manner. And the percentages of in vivo EAC growth inhibition were 19.53, 49.2, and 68.9% at 25, 50, and 100 mg/kg CMFPJ respectively. CMFPJ significantly induced expression of apoptosis regulatory genes caspase-8, caspase-9, cytochrome-c, and caspase-3. This considerable anti-cancer activity was perhaps due to combinatorial effect of lectin, polyphenols, and flavonoids present in CMFPJ.

In Vivo Anticancer Activity of Basella alba Leaf and Seed Extracts against Ehrlich's Ascites Carcinoma (EAC) Cell Line.

Cancer is a class of diseases characterized by uncontrolled cell growth. The current treatment options of cancer are radiotherapy, chemotherapy, hormone therapy, and surgery, where all of them have unpleasant side effects. Due to their adverse side effects, it is challenging to develop new drug for cancer treatment. Hence, the scientists are trying to seek for noble compounds from natural sources to treat cancer. Therefore, in the present investigation, a widely consumable vegetable Basella alba was subjected to evaluate its antiproliferative effect along with molecular signaling of apoptosis in Ehrlich ascites carcinoma (EAC) cell line. Cell growth inhibition was determined by haemocytometer whereas apoptosis of cancer cells were studied by florescence microscope using Hoechst-33342 stain and result was supported by DNA fragmentation and certain cancer related genes expression through PCR analysis. B. alba leaf and seed extract exhibit a considerable scavenging activity in comparison to a standard antioxidant BHT. Moreover, the leaf and seed extracts were able to agglutinate 2% RBC of goat blood at minimum 12.5µg/ml and 50.0µg/ml concentration, respectively. A significant cytotoxic activity was also found in both leaf and seed extract. In haemocytometic observation, the leaf and seed extracts exhibit about 62.54±2.41% and 53.96±2.34% cell growth inhibition, respectively, whereas standard anticancer drug Bleomycin showed 79.43±1.92% growth inhibition. Morphological alteration under fluorescence microscope showed nuclear condensation and fragmentation which is the sign of apoptosis. Apoptosis induction was also confirmed by DNA laddering in leaf and seed treated EAC cells. Upregulation of the tumor suppressor gene P53 and downregulation of antiapoptotic gene Bcl-2 enumerate apoptosis induction. Therefore, current study manifested that leaf and seed extracts of B. alba have antiproliferative activity against EAC cell line and can be a potent source of anticancer agents to treat cancer.