Isolation and characterization of antihypertensive peptides from soy bean protein

Proteins and peptides are the most diverse biomolecules found in nature and make our interest due to their wide applications in food and pharmaceutical industry. Angiotensin Converting Enzyme (ACE) plays a major role in controlling blood pressure. The inhibition of ACE with peptides is a main target in the regulation of hypertension. The objective of the present study was to investigate the therapeutic potential of soy bean. This was accomplished by isolation of ACE inhibitory peptides using response surface methodology (RSM) and characterization of these bioactive peptides by mass spectrometry. 31 hydrolyzed fractions were isolated and evaluated for their ACE inhibition potential. Hydrolyzed fraction having highest ACE inhibitory activity was characterized by liquid chromatography-mass spectrometry (LC-MS) technique. RSM results showed maximum ACE inhibition potential (64%) by hydrolyzate was obtained at 45 ºC temperature, pH 8.0, E/S 0.2 in 2 hours hydrolysis time. Results of LC-MS analysis revealed Ser-Gly, Ser-Pro, Met-Ala, His-Ala, Lys-Pro, Phe-Thr, Met-Leu, Pro-Arg, Ala-Pro-Val, Pro-Ala-Leu, Val-Met-Gly, Pro-Leu-Val, Pro-Pro-Gln, His-Arg-Gly, Ser-Phe-Val-Leu, Ala-Val-His-Try, Arg-Thr-Val-Arg, His-His-Tyr-Leu-Val, Asp-Gly-Ala-Cys-Ser-Ala-Asn and MetVal-Thr-Gly-Pro-Gly-Cys-His bioactive peptides in hydrolyzed fraction of soy bean. Our data provide evidence that response surface methodology is a good approach for isolation of antihypertensive bioactive peptides with more potent activity as nutraceuticals or pharmaceuticals. Therefore soy bean can be use for industrial production of pharmaceutical grade natural medicines for handling high blood pressure.

Enhanced Production Of Streptokinase By Chemical Mutagenesis Of Streptococcus agalactiae EBL-20

Abstract Streptokinase (SK) is an enzyme that is used for the treatment of cardiovascular diseases. The current study focused on the enhanced production of SK by inducing mutation in Streptococcus agalactiae EBL-20 and optimization of medium components and culture conditions for the maximum growth of mutant derived strain. S. agalactiae EBL-32 was selected as a potent mutant after exposure of S. agalactiae EBL-20 to EMS for 180 minutes. SK activity obtained from mutant derived strain was found to be 1.6 fold higher as compared to the activity achieved by wild strain. Nutritional requirements of the mutated strain were optimized by single factor analysis method suggesting glucose as the optimum carbon source; yeast extract and peptone as a suitable nitrogen sources and corn steep liquor (CSL) as an appropriate substrate for the maximum SK production. The culture conditions determined by response surface methodology (RSM) suggested that a temperature value of 37.5⁰C and pH 7 of the fermentation medium with 2.50 mL inoculum size for 36 hours of incubation was optimum for maximum yield of SK. Hence the optimization studies resulted into 1.92 fold increase in the yield of SK suggesting the new isolate suitable for commercial scale production of SK.

Pars plana vitrectomy in vitreous hemorrhage with or without Intravitreal Bevacizumab: a comparative overview

Objective: To compare the success in patients having vitreous hemorrhage undergoing pars plana vitrectomy with or without preoperative intravitreal injection of Bevacizumab

Methods: This Randomized controlled trial was conducted at Department of Ophthalmology, Jinnah Postgraduate Medical Centre. Karachi. Duration of study was six months from January 2010 to June 2010. In this study 56 patients of advanced diabetic eye disease were divided into two groups. Patients in Group-A underwent three ports pars plana vitrectomy with preoperative intravitreal injection of Bevacizumab [Avastin] 1.25mg/0.05ml, 3.5mm from the limbus seven days before surgery and in Group-B patients underwent vitrectomy without preoperative intravitreal Bevacizumab [Avastin]. Intraoperative bleeding was monitored in both groups and was graded as no bleeding, mild bleeding and severe bleeding. The results were statistically analyzed through computer software SPSS 17

Results: Twenty eight patients in Group-A who were given an injection of intravitreal Bevacizumab [Avastin] before surgery, intraoperative bleeding monitored was ,no bleeding in 17 cases [60.7%], mild was observed in 6 cases [21.4%] and severe bleeding requiring diathermy to stop was observed in only 5 cases [17.9%]. 28 patients in Group-B that underwent surgery without Avastin no bleeding was observed in only 2 cases [7.1%], mild in 6 cases [21.4%] and severe in 20 cases [71.4%]

Conclusions: Intravitreal injection of Bevacizumab [Avastin] was effective before vitrectomy in the surgical management of Advanced Diabetic Eye disease

Identification and lead-in characterization of novel B3 metallo-[beta]-lactamases

Metallo-[beta]-lactamases [MBLs] are zinc ion dependent enzymes that are responsible for the emergence and spread of [beta]-lactam resistance among bacterial pathogens. There are uncharacterized putative MBLs in the environment and their emergence is major interference in the generation of universal MBL inhibitors so it is important to identify and characterize novel MBLs. In this study two novel MBLs from Luteimonas sp. J29 and Pseudoxanthomonas mexicana were identified using B3 MBLs as query in BLAST database search. 3D models of putative MBLs generated by SWISSMODEL server taking AIM-1 as a structural template were verified using web based structure assessment and validation programs. Multiple sequence alignment revealed that residues important for substrate binding were conserved and loop region residues [156-162 and 223-230] important for catalysis are variable in these novel MBLs. Homology models showed typical MBL [alpha]/[beta]/[beta]/[alpha] sandwich fold containing six [alpha] helices, twelve [beta] strands and metal interacting residues are conserved in similar way as with other B3 MBLs. We report promising putative B3 MBLs with some variations and substrate docking studies revealed that novel MBLs have attributes close to acquired B3 MBLs

Dietary supplementation of bitter gourd reduces the risk of hypercholesterolemia in cholesterol fed sprague dawley rats

Functional and health endorsing benefits of various foods are often attributed to their phytochemistry. The bitter gourd holds potential in improving the health of the individuals owing to its incredible versatility in phytochemistry. However, the efficacy of different parts of bitter gourd needs attention of the researchers. In the current exploration, different parts of bitter gourd were evaluated for their cholesterol lowering potential in cholesterol fed Sprague dawley rats. For the purpose, four types of bitter gourd part i.e. whole fruit, seedless fruit, seeds, and seed extracts were used and compared with placebo in hypercholesterolemic rats. In placebo, momentous increase in serum cholesterol, triglycerides and LDL levels was observed. All parts attenuate the cholesterol 18.79 to 40.17% triglycerides 25.97 to 37.01% and LDL 14.49 to 26.09%. However, 1% extract powder was most effective in reducing the cholesterol and triglycerides. From the present study, it is deduced that bitter gourd extract can be supplemented in food products for the management of hypercholesterolemia. However, future studies in human subjects needs to be conducted for meticulousness of the present findings

Multiple pesticide residues; in spinach and associated health risk

In past, no work has been done regarding the pesticide residues in spinach targeting peri-urban area of district Faisalabad. Keeping in view the persistent nature and cumulative behavior as well potential toxic effects of pesticides as a result of consumption of vegetable this is planned. Food safety is a major public concern worldwide. This is serious problem in the whole world especially in economically poor countries like Pakistan. The production of safe food is an important aspect of food quality assurance as well as human health. The people are demanding fresh, hygienic and quality food. To study the level of pesticide residues in spinach grown in peri-urban area of district Faisalabad and compare the level of contaminants respective their maximum residues limits [MRLS]. Ten farmers were selected randomly for spinach in the peri-urban area of district Faisalabad and vegetable samples were collected at optimum maturity in triplicate and transported to the laboratory of National Institute of Food Science and Technology, University of Agriculture, Faisalabad for further processing. The residues of pesticide were determined by GC-ECD. The results regarding pesticide residues in different spinach samples showed variations and residues varied as deltamethrin 0.024 to 0.143 mg Kg[-1], permethrin 0.159 to 0.573 mg Kg[-1], cypermethrin 0.098 to 0.503 mg Kg[-1], profenofos 0.016 to 0.425 mg Kg[-1], chlorpyrifos 0.009 to 0.212 mg Kg[-1], bifenthrin 0.012 to 0.623 mg Kg[-1], endosulfan 0.043 to 0.103 mg Kg[-1], dimethoate 0.077 to 0.515 mg Kg[-1], respectively. The chlorpyriphos has no residues in any sample of spinach. The spinach samples are found to be contaminated with a number of different pesticides and these pesticides are not applied judiciously according to good agricultural practices and recommended doses. The farmers are unaware of pesticide use and other agricultural practices due to lack of extension as result causes of serious problems to all community

Improved streptokinase production; unirradiation of streptococcus equisimilis

Several strains of beta-hemolytic Streptococci produce streptokinase enzyme that can bind and activate human plasminogen to plasmin. Streptokinase degrades the fibrin lump by its explicit lysine joining site and so it is applied as a remedy in thrombolytic therapy. The purpose of the study was to subject wild strain of Streptococcus equisimilis to strain development technique, using random mutagenesis by UV irradiation for enhanced production of streptokinase. To evaluate the hyper production of streptokinase after mutagenesis of wild Streptococcus equisimilis by means of UV irradiation. Randomized study. 2012-2014. Enzyme Biotechnology Laboratory, Department of Biochemistry, University of Agriculture, Faisalabad-Pakistan. UV lamp [TUP 40w lamp which has about 90% of its radiation at 2540-2550 A[0]] was used for the mutation of Streptococcus equisimilis cells [1x 10[7] cells mL[-1]] for enhanced production of streptokinase. 10 mL fresh inoculum was transferred to sterile petri plates, which were exposed to UV light for 30, 60, 90, 120, 150, 180, 210, 240 and 270 minutes. The exposure was carried out at distance of 20cm from the centre of lamp. A dose producing 87% killing was selected as optimum dose, after preparing kill curve. The kill/ survival curve was prepared and time of exposure giving [210 minutes] 3 log kill was selected for mutation of the Streptococcus equisimilis for hyper production of streptokinase enzyme. Enzyme assay was performed for both wild and mutant strains. Dose of 210 minutes was selected as best dose which was followed by the selection using triton X-100. Finally the selected strain S. equisimilis EBL-UV-210 showed 480 U mL[-1] of streptokinase activity in quantitative blood clot liquefaction test, which is quite higher than wild strain [370 U mL[-1]]. This maximum yield of streptokinase was obtained after 24h, at CSL 4%, pH 7.5, 37[degree sign] C, KH[2]PO[4] 0.04%, K[2]HPO[4] 0.05%, MgSO[4]. 7H[2]O 0.04%, NaHCO[3] 0.15%, CaCO[3] 0.004%, CH[3]COONa. CH[3]CO 0.10%, FeSO[4]. 7H[2]O 0.04%, MnCl[2]. 4H[2]O 0.02%, glucose 2%, yeast extract 3% and 5% inoculum size in liquid state fermentation. Results showed that mutated strain gave enhanced streptokinase activity in comparison to the wild strain. Our current study focused on streptokinase production from this UV mutated streptococcus equisimilis species and purification of this enzyme by ammonium sulfate precipitation, Ion exchange and gel filtration chromatography. The activity of streptokinase was determined by using quantitative blood clot liquefaction method

Evaluation of Antimicrobial Activity of Glucose Oxidase from Aspergillus niger EBL-A and Penicillium notatum

This work aimed to study the production and purification of glucose oxidase by Aspergillus niger and Penicillium notatum using corn steep liquor as the substrate and evaluate its antimicrobial activity for use in pharmaceutical and food industries. The enzyme was purified by ammonium sulfate precipitation (60-85%), DEAE-cellulose ion exchange and Sephadex G-200 size exclusion chromatography. The crude enzyme extracts of A. niger and P. notatum showed 2.32 and 5.53 U mg-1 specific activities, respectively, which after desalting was 15.52 and 12.05 U mg-1, and after ion exchange and gel filtration chromatography was 29.09 - 62 and 25.72 - 59.37 U mg-1 for A. niger and P. notatum, respectively. The antimicrobial activity was determined by disc diffusion method against selected microbial strains where glucose oxidase from A. niger showed anti-bacterial activity, while no fungicidal effects were shown by both A. niger and P. notatum glucose oxidases.

Kinetic characterization of glucose aerodehydrogenase from Aspergillus niger EMS-150-F after optimizing the dose of mutagen for enhanced production of enzyme

In the present study enhanced production of glucose aerodehydrogenase from Aspergillus niger has been achieved after optimizing the dose of chemical mutagen ethyl methane sulfonate (EMS) that has not been reported earlier. Different doses of mutagen were applied and a strain was developed basing upon the best production. The selected strain Aspergillus niger EMS-150-F was optimized for nutrient requirements in order to produce enzyme through fermentation and the results showed the best yield at 2% corn steep liquor (CSL), 36 hours fermentation time, pH 5, 30°C temperature, 0.3% KH2PO4, 0.3% urea and 0.06% CaCO3. The enzyme was then purified and resulted in 57.88 fold purification with 52.12% recovery. On kinetic characterization, the enzyme showed optimum activity at pH 6 and temperature 30°C. The Michaelis-Menton constants (Km, Vmax, Kcat and Kcat/Km) were 20 mM, 45.87 U mL-1, 1118.81 s-1 and 55.94 s-1 mM-1, respectively. The enzyme was found to be thermaly stable and the enthalpy and free energy showed an increase with increase in temperature and ΔS* was highly negative proving the enzyme from A. niger EMS-150-F resistant to temperature and showing a very little disorderliness.

Chronic viral hepatitis c infection; frequency of impaired glucose tolerance in patients

Objective: The objective of the study was to measure the frequency of impaired glucose tolerance [IGT] in patients having chronic hepatitis C at medical outpatient department [OPD] of Allied hospital Faisalabad

Sample technique: Non probability consecutive sampling

Duration of study: Duration of study was 6 months with first patient enrolled on 3.02.2010 and last patient enrolled on 07-08-2010

Setting: Medical [OPD] Outpatient Department of Allied Hospital, Faisalabad

Subjects: 375 patients with chronic hepatitis C, coming in Medical OPD of Allied hospital Faisalabad were enrolled

Methods: 375 patients diagnosed as having chronic hepatitis C coming in Medical OPD of Allied hospital Faisalabad, were included in the study. Fasting and two hours after glucose load blood sample were collected in sterilized syringes and was tested for fasting blood glucose [FBS] and random blood glucose level [RBS]

Results: In this study 375 patients having HCV positive on Elisa were enrolled. Out of 375 patients with chronic hepatitis C, 143 [38.1%] were found to have IGT while 232 [61.9%] were having no IGT. There was significant association between chronic hepatitis C and development of IGT

Conclusions: It is concluded that there is close association in the development of IGT in patients with chronic hepatitis C

Production of rabbit antibodies against purified Glucose oxidase

Glucose oxidase is an active oxygen species generating enzyme produced from Aspergillus niger grown in submerged fermentation. Disintegration of the mycelium resulted in high glucose oxidase activity that was subjected to ammonium sulfate precipitation at 60-85% saturation rates that resulted to 6.14 U mg -1 specific activity. Purification of enzyme by anion exchange column (DEAE-Cellulose) resulted into 22.53 U mg-1 specific activity and 10.27 fold purification. This was applied to sephadex G-200 column for gel filtration chromatography. It was observed that enzyme achieved 59.37 U mg-1of specific activity with 27.08 fold purity and 64.36% recovery. Purified glucose oxidase was injected into rabbits through intravenous route, to raise the glucose oxidase antibodies. After 30 days incubation period, the rabbits were slaughtered and serum was separated from blood. The antibodies were isolated by ammonium sulfate precipitation and confirmed by agar gel precipitation test. This could be a convenient and low cost alternate assay for the estimation of glucose oxidase in biological fluids. Moreover, such antibodies against the said enzyme could be used in various therapeutic and diagnostic applications.

Effect of captopril glycation inhibition in normal and diabetic plasma by captopril

To investigate the inhibitory effect of Captopril on level of glycation [in vivo]. [2] To study glycation inhibition in vivo. Study Case study. Period: Sep. 2006 to March. 2008. One year seven months. Department of Biochemistry University of Agriculture, Faisalabad. Different parameters like fluorescence, total proteins, TBA [thiobarbituric acid] method, periodate borohydride assay were used to check the effect of inhibitor on glycation. Thirty two combinations were made and all these combinations were placed at 37°C, at same time for five weeks. 3mL of blood sample was drawn after 1st, 3rd and 5th week of incubation to perform the experiments for glycation and glycation inhibition. Along with the same temperature [37°C], different combinations of glucose and inhibitor were used. Effective concentration of inhibitor helped to decrease the level of glycation. All concentrations of glucose [G, G and G] showed 1 2 3 glycation with protein. The inhibitor Captopril [all concentrations] showed variations in inhibition of glycation at one temperature [37°C with different parameters [Fluorescence, TBA and Periodate] but the most effective concentration of inhibitors at each condition is I [1mM] but I [10 3 1 mM] and I [5 mM] were also equally effective after I. Periodate borohydride Assay is more effective for glycation determination than 2 3 thiobarbituric acid assay. Captopril can be used as glycation inhibitor in future. As it enhances the activity of transketolase, it can produce 3DG compound which can block the AGEs. However, more experimentations should be done on animal or on large scale before its application in diabetic patients

Recent ethical and technical issues of cadaveric human bodies and tissues for educational purposes in pakistan: initiation of a discussion for making a policy

OBJECTIVE: To initiate a discussion, for devising a standardized nationwide policy for procurement and use of Cadaveric human bodies and tissues for educational purposes and then respectful disposition of the remains

DESIGN: Cohort design

PLACE: A Public Medical College

DURATION: 2006 through 2011

METHODS: Available data was grouped into two chronological equal halves, Group-A 2006 to 2008, Group-B 2009 to 2011. The data was analyzed for age, sex, Known identity, Unknown Identity, later identified-claimed and remaining unidentified-unclaimed dead bodies. Studies related to policies for cadaveric procurement, transport, handling, utilization and disposal are presented

RESULTS: A total of 1605 records were analyzed. 54.9 % Cadavers were from 2 and 3 decade of life. The males in Group-A were 86.32% and in Group-B were 81.82%

DISCUSSION: About 41.55 % cadavers brought to Anatomy department were with Unknown Identity. But 34.93 % were later identified and claimed. Hence the police department of the region and the Anatomy Department of the institution with the help of print media are providing a great service to the society

The remaining unclaimed bodies are also utilized for dissection or in case of putrefied bodies bones were used for medical education. Even after exhaustive search no published data was available for cadavers brought to a public mortuary in Pakistan, as is presented here

CONCLUSION: Pakistan, being a developing nation, is quite vulnerable to many hazards of the increasing demands of cadavers in the medical education business. This is high time to start a discussion for making a comprehensive policy for caring and utilizing dead bodies for medical education. In this respect the anatomist, Islamic scholars and legal experts can collaborate with 'organ transplant' legislation bodies for devising policies

Production of extracellular lipases by Rhizopus oligosporus in a stirred fermentor

The present investigation deals with the kinetics of submerged extracellular lipases fermentation by both wild and mutant strains of Rhizopus oligosporus var. microsporus in a laboratory scale stirred fermentor. Other parameters studied were inoculum size, pH, agitation and rate of aeration. It was found that the growth and lipases production was increased gradually and reached its maximum 9.07± 0.42ª U mL-1 (W) and 42.49 ± 3.91ª U mL-1 (M) after 30h of fermentation for both wild and mutant strain. There is overall increase of 109 percent (W) and 124 percent (M) in the production of extracellular lipases as compared to shake flask. Another significant finding of the present study is that the fermentation period is reduced to 30 h in case of wild and 23 h in case of mutant from 48 h in shake flask studies. The specific productivity of mutant strain (qp = 377.3 U/g cells/h) was several folds higher than wild strain. The specific production rate and growth coefficient revealed the hyperproducibility of extracellular lipases using mutant IIB-63NTG-7.

Anticoccidial activity of Curcuma longa L. in broilers

Comparative efficacy of turmeric (Curcuma longa L.) crude powder and salinomycin sodium on the occurrence of coccidiosis and growth performance of broiler was evaluated. A total of 90, day-old chicks were randomly divided into six groups. From first day onward, ration was supplemented with 1, 2 and 3 percent turmeric powder in groups 2, 3 and 4, respectively, group 1 received salinomycin sodium @ 12 g 50 kg-1 feed while groups 5 and 6 were kept as infected un-medicated and uninfected un-medicated controls. First five groups were infected with Eimeria tenella sporulated oocysts @ 1,00,000/chick at the age of 20 days. Body weight gain, feed consumption, feed conversion ratio were investigated throughout the experimental period, and bloody diarrhea and oocysts excretions were investigated at the first and the second week after infection. Maximum coccidiostatic effect was observed with turmeric (3 percent) showing mild bloody diarrhea as compared to other infected groups receiving turmeric containing rations. This effect was comparable with a standard coccidiostat i.e., salinomycin sodium. Similarly, the weight gain in the groups treated with salinomycin sodium (2280g) and 3 percent turmeric (2293g) were also significantly higher (p < 0.05) than that of infected control group (1955g). In the groups treated with ration supplemented with 3 percent turmeric powder and salinomycin sodium, the peak excretion of oocysts was delayed about 1 or 2 days relative to the control infected group. Concentration-dependent coccidiostatic effect of turmeric suggested that further studies should be carried out to determine the possible maximum safe levels of turmeric with least toxic effects to be used as coccidiostat.

effect of size of the needle on the rate of pneumothorax after ct guided transthoracic needle aspiration biopsy of lung masses

Purpose of Study: To analyze the rate of Pneumothorax after CT guided TNAB of lung masses, using multiple needle sizes

Patients and Method: Seventy patients underwent CT guided FNA of the chest masses

Results: Out of seventy, 18 patients i.e.; 26 % developed Pneumothorax. With 18G needle 24 patients underwent FNA and seven of them i.e: 25% developed Pneumothorax. With 20G needle, 26% developed pneumothorax and with the 22G needle 25% developed Pneumothorax

Conclusion: The study shows that the rate of Pneumothorax after CT guided TNAB of Lung Masses remains almost the same regardless of the size of the needle

Retained sponge after abdominal surgery

To study the cause and clinical presentation along with clinical course [coperative and postoperative] of retained abdominal sponge. Design: Observational case series. Place and Duration of Study: Surgical Department, Bahawal Victoria Hospital, Bahawalpur, from April 1, 2000 to August 31, 2002. Materials and The study included 12 patients. All the patients, who presented to surgical department during the stated period with retained abdominal sponge postoperatively, were included in the study. Retained abdominal foreign bodies, other than the surgical sponge, were excluded. A total of 12 patients presented with retained abdominal sponge. Out of these, 33.33% were males and 66.66% females. Modes of presentation were intestinal obstruction, 58.33%, discharging sinus, 41.67%, intraabdominal abscess,16.67%, peritonitis, 16.67% and mass abdomen, 8.33%. One death followed despite provision of due care. Retained surgical sponge is a potentially lethal condition and all preventive measures should be taken to avoid this condition