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Saleem F.,Lahore College for Women University | Aslam R.,Lahore College for Women University | Aslam R.,Food and Biotechnology Research Center | Saleem Y.,Food and Biotechnology Research Center | And 5 more authors.
Pakistan Journal of Zoology

Synthetic polymers obtained from petrol causes air pollution only because they are not degraded in soil for a long time. For this reason, a microbial plastic poly-β-hydroxybutyrate (PHB) has gained importance since it can be easily degraded in nature. PHB is a widely distributed intracellular reserve substance typical of prokaryotes. PHB exists in the cytoplasmic fluid in the form of crystalline granules about 0.5 μm in diameter and can be isolated as native granules or by solvent extraction. The study aimed at screening of PHB producing strain and optimizing media parameters for increased PHB production by a Bacillus thuringiensis strain CMBL-BT-6, which was identified as PHB producing strain by staining with Sudan Black B dye. Conditions for best PHB production were optimized. Maltose was found to be best carbon source and ammonium nitrate the best nitrogen source producing 0.867g/L and 0.953g/L of PHB, respectively. Glucose was used as a substrate and 4% glucose concentration (0.953g/L) was found to be best for PHB production. The data further indicated that CMBL-BT-6 produced 2.75g/L PHB at pH 7.0, when incubated at 37°C for 24 h. © 2014 Zoological Society of Pakistan. Source

Samra Z.Q.,University of Punjab | Shabir S.,University of Punjab | Rehmat Z.,University of Punjab | Zaman M.,University of Punjab | And 3 more authors.
Applied Biochemistry and Biotechnology

Human serum paraoxonase 1 (PON1) is known as an antioxidant and is also involved in the detoxification of many compounds. In this study, a novel purification strategy was employed to purify the PON1 by using cholesterol-conjugated magnetic nanoparticles. Magnetic nanoparticles were synthesized and conjugated with cholesterol through diazotized p-aminohippuric acid. In Fourier transform infrared spectrum of cholesterol-p-aminohippuric acid-Fe3O4 nanoparticles, the appearance of peaks at 3,358.3, 1,645 cm-1, and at 2,334.9 cm-1 confirmed the conjugation. The molecular weight of purified PON1 was nearly 45 kDa on sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE), and isoelectric point was 5.3. The specific activity was 438 U mg-1 protein, and the purification fold was 515 with 73% yield. The Km values were 1.3 and 0.74 mM with paraoxon and phenyl acetate, respectively. Western blot of 2D-PAGE confirmed the homogeneity and stability of the enzyme. Mg+2, Mn +2, glycerol, (NH4)2SO4, PEG 6000, Triton X-100, and phenylmethylsulfonyl fluoride did not show any effect on activity. Pb+2, Co+2, Zn2+, ethanol, β-mercaptoethanol, and acetone reduced the activity while Ni2+, Cd2+, Cu2+, iodoacetic acid, SDS, dimethylformamide, DMSO inhibited the activity. In vitro enzyme activity was slightly reduced by acetyl salicylic and acetaminophen and reduced 50% with amino glycosides and ampicillin antibiotics at concentrations of 0.6 and 30 mg ml-1, respectively. This is the first report for the synthesis of cholesterol-conjugated magnetic nanoparticles for simple purification of PON1 enzyme. © Humana Press 2009. Source

Dar N.,Jinnah Degree College for Women | Javed M.,University of Punjab | Samra Z.Q.,University of Punjab
Archives of Biological Sciences

Enterotoxins secreted by S. aureus are known as a food-poisoning agent that is associated with various gastrointestinal pathological conditions. In this study, a one-step immunodetection method was devised for routine checking of SEB in milk and fruit juices available locally. Antibodies against recombinant SEB were raised, purified, and cross reactivity was checked against clinically important bacteria (Shigella flexneri, Streptococci, Salmonella typhi, Klebsiella and Bacillus subtilis). Purified anti-SEB antibodies were conjugated with gold nanoparticles (Ab-GNPs) for direct detection of SEB in samples. SEB (33%, 4.76% and 15%) was found in non-sterilized milk (118), sterilized milk (42) and juices (60), respectively. Coagulase, MSA tests and PCR amplification of 725 bp of the SEB gene confirmed the presence of S. aureus in the collected samples positive for SEB. Immunoassay is easy, reliable and less time consuming and will be helpful to detect the SEB in food samples at local level. Source

Pervaiz S.,University of Punjab | Shaheen T.,University of Punjab | Shaheen S.,University of Punjab | Dar N.,Jinnah Degree College for Women | Samra Z.Q.,University of Punjab
Journal of Applied Microbiology

Aim: S-layer proteins are considered as a good nanocarrier due to their binding and self-assembled properties. These can be used to prepare the immunomatrixes for the removal of toxins from the samples. Methods and Results: Two S-layer proteins 70 and 40 kDa of thermophilic Thermobifida fusca were extracted with guanidine hydrochloride and purified. Antibodies against S-layer proteins were developed, and their monospecificity was checked. Immunogold labelling indicated that these are surface proteins. Immunomatrixes (70-SLIM, 40 SLIM) were prepared by covalently immobilizing S-layer proteins in microwell and further conjugated with anti- Staphylococcus aureus enterotoxin B (SEB) antibodies. The binding of 70 and 40 kDa proteins was observed nearly 7·0 μg cm-1 to binding area, and the conjugation with anti-SEB antibodies was found 1·22 μg μg-1 of 70 kDa and 0·875 μg μg-1 of 40 kDa. The average binding and elution of pure SEB toxin on 70-SLIM and 40-SLIM was 5·0 μg SEB toxin. The SEB toxin in milk samples was also removed on immunomatrixes successfully. Conclusion: It is the first report, and this study shows that the thermophilic S-layer proteins can be used to prepare the immunomatrixes. Significance and Impact of Study: Information in this study can be used to design the strategies for the removal of biologically important materials or toxins from samples. Copyright © 2013. Source

Samra Z.Q.,University of Punjab | Sana A.,University of Punjab | Bano S.,University of Punjab | Farooq M.,University of Punjab | And 2 more authors.
Journal of Clinical Laboratory Analysis

Cardiac diseases are the major cause of death. Paraoxonase1 (PON1) is known as free radicals scavenger/anti-atherosclerosis, whereas xanthine oxidase (XO) is a free radicals generator. This study was undertaken to determine and compare the Paraoxonase and arylesterase activities of PON1 enzyme and activity of XO enzyme. The concentration of XO and PON1 enzymes along with lipid profile, lipid peroxides, and thiol level in plasma of cardiac patients (n = 200) and healthy persons (n = 200) of Lahore metropolitan, Pakistan was also determined. Anti-PON1 and anti-XO antibodies were developed, purified, and used to measure the concentration of PON1 and XO by competitive ELISA. It is observed that low paraoxonase (P = 0.0073)/arylesterase activity (P = 0.0038) of PON1 enzyme and its low concentration (P = 0.0049) were observed in cardiac patients, whereas elevated level of XO activity (P = 0.0129) and its concentration (P = 0.0097) was observed in cardiac patients as compared with healthy persons. Low levels of HDL (P = 0.0013), thiol (P = 0.0014) and high level of cholesterol (P = 0.0025), triglycerides (P = 0.0018), LPO (P = 0.0014), and LDL level (P = 0.05) were observed in cardiac patients admitted in intensive care unit as compared with hypertensive patients and control subjects. It is concluded that overall low PON1 and high XO activities do cause imbalance of free radical system which ultimately leads to or enhance the cardiac pathological conditions. © 2010 Wiley-Liss, Inc. Source

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