Exposure to aflatoxin B1 and associated risk factors in hepatitis C patients in cosmopolitan city of Pakistan: facility-based study.

INTRODUCTION: population-based follow-up study has been designed to investigate the contributing factors to high exposure to Aflatoxin B1 (AFB1) and the subsequent associated risk factors among hepatitis C-infected patients at a referral centre, Karachi. Pakistan. Hepatitis C infection affects millions of individuals worldwide and confers high morbidity and mortality, especially in lower middle-income countries (LMICs) including Pakistan. A literature review of recent studies has revealed that a number of hepatocellular carcinomas (HCC) cases are markedly increased in Pakistan, where one of the potential causes of HCC is hepatitis C virus. The objectives of this study were to determine frequency of Aflatoxin B1 (AFB1) exposure and other associated characteristics among hepatitis C patients at a referral centre, Karachi, Pakistan. METHODS: a semi-structured pre-coded pro forma designed to collect socio-demographic, Pharmacological, biochemical and clinical information from patients and hospital records. Patient´s pre and post polymerase chain reaction (PCR), serum alanine aminotransferase (ALT) levels and other blood parameters were analysed. AFB1 exposure was determined using an ELISA kit and validated through high-performance liquid chromatography (HPLC). RESULTS: AFB1 exposure was found in 30 (34%) patients. Post treatment responders were 49 (55.6%). More than 37% of study participants had a family history of hepatitis C. About 74% had a history of surgical procedure, and around 36% of study participants had a blood transfusion history. Up to 36% participants were fond of spicy food and around 25% study participants were eating roadside food on daily basis. CONCLUSION: high frequency of AFB1 exposure due to risky dietary habits, low level of formal education and awareness are contributing factors may be responsible for high exposure of AFB1. Effective and multidimensional strategies are needed to prevent advance stage progression of disease and associated complications.

Combined pretreatment of sugarcane bagasse using alkali and ionic liquid to increase hemicellulose content and xylanase production.

BACKGROUND: Lignin in sugarcane bagasse (SB) hinders its utilization by microorganism, therefore, pretreatment methods are employed to make fermentable components accessible to the microbes. Multivariate analysis of different chemical pretreatment methods can aid to select the most appropriate strategy to valorize a particular biomass. RESULTS: Amongst methods tested, the pretreatment by using sodium hydroxide in combination with methyltrioctylammonium chloride, an ionic liquid, (NaOH+IL) was the most significant for xylanase production by Bacillus aestuarii UE25. Investigation of optimal levels of five significant variables by adopting Box-Behnken design (BBD) predicted 20 IU mL- 1 of xylanase and experimentally, a titer of 17.77 IU mL- 1 was obtained which indicated the validity of the model. The production kinetics showed that volumetric productivity of xylanase was much higher after 24 h (833.33 IU L- 1 h- 1) than after 48 h (567.08 IU L- 1 h- 1). The extracted xylan from SB induced more xylanase in the fermentation medium than pretreated SB or commercially purified xylan. Nuclear Magnetic Resonance, Fourier transform infrared spectroscopy and scanning electron microscopy of SB indicated removal of lignin and changes in the structure of SB after NaOH+IL pretreatment and fermentation. CONCLUSION: Combined pretreatment of SB with alkali and methyltrioctylammonium chloride appeared better than other chemical methods for bacterial xylanase production and for the extraction of xylan form SB.

Cellulose extraction from methyltrioctylammonium chloride pretreated sugarcane bagasse and its application.

Cellulose, the most abundant feedstock of chemicals and energy is extracted from various agro-industrial wastes, such as sugarcane bagasse (SB). Pretreatment of SB with ionic liquids improves extraction of cellulose, yet the use of ionic liquid is hindered by its high cost. In this study, cellulose was extracted from SB pretreated with methyltrioctylammonium chloride under relatively mild conditions. The extracted cellulose from pretreated SB (PTB) and untreated SB (UTB) was characterized by scanning electron microscopy and FTIR. Fermentation of cellulose extracted from PTB by a thermophilic bacterium, Bacillus aestuarii UE25, yielded 245.16% higher titers of cellulase than cellulose extracted from UTB. The recyclability of the IL was assessed to make the pretreatment process cost effective and was monitored through TLC and FTIR. The results of this research demonstrated the potential of ionic liquid pretreated SB for cellulose extraction and for its subsequent utilization in thermostable cellulase production.

Utilization of methyltrioctylammonium chloride as new ionic liquid in pretreatment of sugarcane bagasse for production of cellulase by novel thermophilic bacteria.

Fermentation of carbohydrates present in lignocellulosic (LC) biomass is facilitated by lignin removal, which is usually achieved by adopting various pretreatment methods to provide the enzymes proper access to their respective substrates. Pretreatment using ionic liquid (IL) is relatively recent advancement and considered as mild and green process. ILs can dissolve extensive quantities of biomass and depolymerize the cellulose. In this context, an abundantly available LC biomass, sugarcane bagasse (SB), was pretreated using alkali or with an IL, methyltrioctylammonium chloride, and was used for cellulase production from thermophilic bacteria. In all, 26 indigenously isolated thermophilic bacterial strains were quantitatively screened for cellulase production. 16S rDNA sequences of the promising isolates UE10 and UE27 revealed relatedness with Brevibacillus borstelensis, while the strain UE1 belonged to Aneurinibacillus thermoaerophilus. Cellulase production was compared by utilizing alkali pretreated and IL pretreated SB and the later was found more appropriate. UE1, UE10 and UE27 yielded 22.2, 22.18 and 33.3 IU mL-1 of endoglucanase, respectively, by fermenting IL pretreated SB. The changes in SB structure after pretreatment were evaluated by scanning electron microscopy. This study demonstrated the potential of novel thermophilic bacterial strains to utilize IL pretreated SB for production of industrially important enzyme, cellulase.

Methyltrioctylammonium chloride mediated removal of lignin from sugarcane bagasse for themostable cellulase production.

The present study was aimed to evaluate the Methyltrioctylammonium Chloride (IL) and Sodium hydroxide effect on sugarcane bagasse (SB) structure and its subsequent utilization to produce cellulase from a thermophilic bacterium Bacillus aestuarii UE25. The strain was isolated from a crocodile pond of Manghopir, Karachi. Ten different factors affecting IL pretreatment of SB and cellulase production by UE25 were evaluated by Plackett-Burman design and three significant factors were optimized by employing Box Behnken design. Under optimum conditions, the strain produced 118.4 IU mL-1 of EG and 75.01 IU mL-1 of BGL that corroborated well with the predicted values by the model. Scanning electron microscopy, gravimetric analysis, Fourier transform infrared spectroscopy and NMR of SB revealed removal of lignin, decrease in cellulose content and structural changes in the SB after pretreatment and fermentation. The data provide prospects of utilizing this IL in comparison to imidazolium based IL for pretreatment of biomass.

Synthesis and bio-molecular study of (+)-N-Acetyl-α-amino acid dehydroabietylamine derivative for the selective therapy of hepatocellular carcinoma.

BACKGROUND: The purpose of present work is to synthesize novel (+)-Dehydroabietylamine derivatives (DAAD) using N-acetyl-α-amino acid conjugates and determine its cytotoxic effects on hepatocellular carcinoma cells. METHODS: An analytical study was conducted to explore cytotoxic activity of DAAD on hepatocellular carcinoma cell lines. The cytotoxicity effect was recorded using sulforhodamine B technique. Cell cycle analysis was performed using Propidium Iodide (PI) staining. Based on cell morphology, anti growth activity and microarray findings of DAAD2 treatment, Comet assay, Annexin V/PI staining, Immunoperoxidase assay and western blots were performed accoringly. RESULTS: Hep3B cells were found to be the most sensitive with IC50 of 2.00 ± 0.4 µM against (+)-N-(N-Acetyl-L-Cysteine)-dehydroabietylamine as DAAD2. In compliance to time dependent morphological changes of low cellular confluence, detachment and rounding of DAAD2 treated cells; noticeable changes in G2/M phase were recorded may be leading to cell cycle cessation. Up-regulation (5folds) of TUBA1A gene in Hep3B cells was determined in microarray experiments. Apoptotic mode of cell death was evaluated using standardized staining procedures including comet assay and annexin V/PI staining, Immuno-peroxidase assay. Using western blotting technique, caspase dependant apoptotic mode of cell death was recorded against Hep3B cell line. CONCLUSION: It is concluded that a novel DAAD2 with IC50 values less than 8 µM can induce massive cell attenuation following caspase dependent apoptotic cell death in Hep3B cells. Moreover, the corelation study indicated that DAAD2 may have vital influence on cell prolifration properties.

Comparative evaluation of in vitro cytotoxic effects among parent abietyl alcohol and novel fatty acid ester derivatives against MCF7 and hepatocellular carcinoma cell lines.

Synthesis of twelve hitherto unreported esters of abietyl alcohol and screening of these esters against four cancer cell lines including one breast cancer line MCF7 and four hepatocellular carcinoma cell lines (HCC) Huh7, Hep3B, Snu449 and Plc has been determined using SRB assay. The Cell cycle progression showed changes in cellular behaviour after 48 and 72 hours in MCF7 and Huh7 cell lines. Abietyl alcohol was obtained from the reduction of abietic acid, a tricyclic diterpene, isolated from oleoresin of Pinus longifolia Roxberghii.

Synthesis and characterization of amino acid conjugates of oleanolic acid and their in vitro cytotoxic effect on HCC cell lines.

Oleanolic acid (3ß-hydroxy-olean-12-en-28-oic acid; OA-01), a pentacyclic triterpene, exhibit a wide range of pharmacological and biological activities. We have isolated oleanolic acid from methanolic extract of Periploca aphylla, collected from surroundings of Karachi in the month of February. Furthermore, four known and two new C-28 amino acid conjugates of oleanolic acid were prepared to explore potential of these compounds on HCCs and one breast cancer cell line. Cytotoxic effects revealed that as compare to parent compound (OA-01), two derivatives OA-04 (p<0.0001) and OA-06 (p<0.01) showed significantly increased/higher inhibition rates.