Search results for: Enzymes

Authors: Abou-Baker Salim, A. A. K. Abou-Arab, Sherif R. Mohamed, T. A. Eldesouky

Abstract:

Pomegranate (Punica granatum L.) is an ancient fruit of great medical interest and rich source of antioxidants. Pesticides as dimethoate play a crucial role in the occurrence many diseases in plants, animal and human. Therefore the ability of Pomegranate (Punica granatum L.) to alleviate hepatotoxicity induced by organophosphate pesticide dimethoate was investigated. Albino male rats were divided randomly into 4 groups and kept at 7 animals per group in an environmentally controlled condition for 6 weeks. The first group was served as a control group (basal diet), the second group fed on basal diet supplemented with 5% freeze dried pomegranate seeds, the third group fed on 20 ppm dimethoate contaminated diet and the last group fed on dimethoate contaminated diet supplemented with 5% freeze dried pomegranate seeds. The results revealed that administration of dimethoate caused high significant increased in liver functions: alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP) activities as well as lipid peroxide (malonaldhyde, MDA); on the other hand high significant decreased on glutathione (GSH), glutathione peroxidase (GPx), albumin and total protein were observed. However addition of 5% freeze dried pomegranate seeds significantly improved all previously mentioned parameters. These results indicate the dimethoate induced hepatotoxicity and highlight the protective effect of pomegranate seeds as a potential protective agent against dimethoate induced hepatotoxicity. This may be attributed to the powerful antioxidants (polyphenols, total phenols, and total flavonoids) which present in high levels in pomegranate as well as improving the immunity by activation of antioxidant enzymes GSH and GPx.

Keywords: Pomegranate, organophosphate pesticides, dimethoate, liver toxicity, free radicals, antioxidants.

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Authors: Gnanaprakash K, Madhusudhana Chetty C, Ramkanth S, Alagusundaram M, Tiruvengadarajan VS, Angala Parameswari S, Mohamed Saleem TS

Abstract:

Carbon tetrachloride (CCl4) is a well-known hepatotoxin and exposure to this chemical is known to induce oxidative stress and causes liver injury by the formation of free radicals. Flacourtia indica commonly known as 'Baichi' has been reported as an effective remedy for the treatment of a variety of diseases. The objective of this study was to investigate the hepatoprotective activity of aqueous extract of leaves of Flacourtia indica against CCl4 induced hepatotoxicity. Animals were pretreated with the aqueous extract of Flacourtia indica (250 & 500 mg/kg body weight) for one week and then challenged with CCl4 (1.5 ml/kg bw) in olive oil (1:1, v/v) on 7th day. Serum marker enzymes (ALP, AST, ALT, Total Protein & Total Bilirubin) and TBARS level (Marker for oxidative stress) were estimated in all the study groups. Alteration in the levels of biochemical markers of hepatic damage like AST, ALT, ALP, Total Protein, Total Bilirubin and lipid peroxides (TBARS) were tested in both CCl4 treated and extract treated groups. CCl4 has enhanced the AST, ALT, ALP and the Lipid peroxides (TBARS) in liver. Treatment of aqueous extract of Flacourtia indica leaves (250 & 500 mg/kg) exhibited a significant protective effect by altering the serum levels of AST, ALT, ALP, Total Protein, Total Bilirubin and liver TBARS. These biochemical observations were supported by histopathological study of liver sections. From this preliminary study it has been concluded that the aqueous extract of the leaves of Flacourtia indica protects liver against oxidative damages and could be used as an effective protector against CCl4 induced hepatic damage. Our findings suggested that Flacourtia indica possessed good hepatoprotective activity

Keywords: Carbon Tetrachloride, Flacourtia indica, Hepatoprotective activity, Oxidative stress

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Authors: K. Gauthaman, T.S. Mohamed Saleem, V. Ravi, Sita Sharan Patel, S. Niranjali Devaraj

Abstract:

The present study was designed to investigate the cardio protective role of chronic oral administration of alcoholic extract of Terminalia arjuna in in-vivo ischemic reperfusion injury and the induction of HSP72. Rabbits, divided into three groups, and were administered with the alcoholic extract of the bark powder of Terminalia arjuna (TAAE) by oral gavage [6.75mg/kg: (T1) and 9.75mg/kg: (T2), 6 days /week for 12 weeks]. In open-chest Ketamine pentobarbitone anaesthetized rabbits, the left anterior descending coronary artery was occluded for 15 min of ischemia followed by 60 min of reperfusion. In the vehicle-treated group, ischemic-reperfusion injury (IRI) was evidenced by depression of global hemodynamic function (MAP, HR, LVEDP, peak LV (+) & (- ) (dP/dt) along with depletion of HEP compounds. Oxidative stress in IRI was evidenced by, raised levels of myocardial TBARS and depletion of endogenous myocardial antioxidants GSH, SOD and catalase. Western blot analysis showed a single band corresponding to 72 kDa in homogenates of hearts from rabbits treated with both the doses. In the alcoholic extract of the bark powder of Terminalia arjuna treatment groups, both the doses had better recovery of myocardial hemodynamic function, with significant reduction in TBARS, and rise in SOD, GSH, catalase were observed. The results of the present study suggest that the alcoholic extract of the bark powder of Terminalia arjuna in rabbit induces myocardial HSP 72 and augments myocardial endogenous antioxidants, without causing any cellular injury and offered better cardioprotection against oxidative stress associated with myocardial IR injury.

Keywords: Antioxidants, HSP72, Ischemic reperfusion injury, Terminalia arjuna.

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Authors: Ching-Wei Tsai, Chih-I Liu, Ruoh-Chyu Ruaana

Abstract:

A new strategy for oriented immobilization of proteins was proposed. The strategy contains two steps. The first step is to search for a docking site away from the active site on the protein surface. The second step is trying to find a ligand that is able to grasp the targeted site of the protein. To avoid ligand binding to the active site of protein, the targeted docking site is selected to own opposite charges to those near the active site. To enhance the ligand-protein binding, both hydrophobic and electrostatic interactions need to be included. The targeted docking site should therefore contain hydrophobic amino acids. The ligand is then selected through the help of molecular docking simulations. The enzyme α-amylase derived from Aspergillus oryzae (TAKA) was taken as an example for oriented immobilization. The active site of TAKA is surrounded by negatively charged amino acids. All the possible hydrophobic sites on the surface of TAKA were evaluated by the free energy estimation through benzene docking. A hydrophobic site on the opposite side of TAKA-s active site was found to be positive in net charges. A possible ligand, 3,3-,4,4- – Biphenyltetra- carboxylic acid (BPTA), was found to catch TAKA by the designated docking site. Then, the BPTA molecules were grafted onto silica gels and measured the affinity of TAKA adsorption and the specific activity of thereby immobilized enzymes. It was found that TAKA had a dissociation constant as low as 7.0×10-6 M toward the ligand BPTA on silica gel. The increase in ionic strength has little effect on the adsorption of TAKA, which indicated the existence of hydrophobic interaction between ligands and proteins. The specific activity of the immobilized TAKA was compared with the randomly adsorbed TAKA on primary amine containing silica gel. It was found that the orderly immobilized TAKA owns a specific activity twice as high as the one randomly adsorbed by ionic interaction.

Keywords: Protein Oriented immobilization, Molecular docking, ligand design, surface modification.

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Authors: Monisha Isaac

Abstract:

Biotechnology is a discipline which explains the use of living organisms and systems to construct a product, or we can define it as an application or technology developed to use biological systems and organisms processes for a specific use. Particularly, it includes cells and its components use for new technologies and inventions. The tools developed can be further used in diverse fields such as agriculture, industry, research and hospitals etc. The 21^st century has seen a drastic development and advancement in biotechnology in India. Significant increase in Government of India’s outlays for biotechnology over the past decade has been observed. A sectoral break up of biotechnology-based companies in India shows that most of the companies are agriculture-based companies having interests ranging from tissue culture to biopesticides. Major attention has been given by the companies in health related activities and in environmental biotechnology. The biopharmaceutical, which comprises of vaccines, diagnostic, and recombinant products is the most reliable and largest segment of the Indian Biotech industry. India has developed its vaccine markets and supplies them to various countries. Then there are the bio-services, which mainly comprise of contract researches and manufacturing services. India has made noticeable developments in the field of bio industries including manufacturing of enzymes, biofuels and biopolymers. Biotechnology is also playing a crucial and significant role in the field of agriculture. Traditional methods have been replaced by new technologies that mainly focus on GM crops, marker assisted technologies and the use of biotechnological tools to improve the quality of fertilizers and soil. It may only be a small contributor but has shown to have huge potential for growth. Bioinformatics is a computational method which helps to store, manage, arrange and design tools to interpret the extensive data gathered through experimental trials, making it important in the design of drugs.

Keywords: Biotechnology, advancement, agriculture, bio-services, bio-industries, bio-pharmaceuticals.

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Authors: Hanaa M. M. El-Khayat, Hoda Abdel-Hamid, Kadria M. A. Mahmoud, Hanan S. Gaber, Hoda, M. A. Abu Taleb, Hassan E. Flefel

Abstract:

Snails are considered as suitable diagnostic organisms for heavy metal–contaminated sites. Biomphalaria alexandrina snails are used in this work as pollution bioindicators after exposure to chemical mixtures consisted of heavy metals (HM); zinc (Zn), copper (Cu) and lead (Pb); and persistent organic pollutants; Decabromodiphenyl ether 98% (D) and Aroclor 1254 (A). The impacts of these tested chemicals, individual and mixtures, on liver and kidney functions, antioxidant enzymes, complete blood picture, and tissue histology were studied. Results showed that Cu was proved to be the highly toxic against snails than Zn and Pb where LC₅₀ values were 1.362, 213.198 and 277.396 ppm, respectively. Also, B. alexandrina snails exposed to the mixture of HM (¼ LC₅ Cu, Pb and Zn) showed the highest bioaccumulation of Cu and Zn in their whole tissue, the most significant increase in AST, ALT & ALP activities and the highest significant levels of total protein, albumin and globulin. Results showed significant alterations in CAT activity in snail tissue extracts while snail samples exposed to most experimental tests showed significant increase in GST activity. Snail samples that exposed to HM mixtures showed a significant decrease in total hemocytes count while snail samples that exposed to mixtures containing A & D showed a significant increase in total hemocytes and Hyalinocytes. Histopathological alterations in snail samples exposed to individual HM and their mixtures for 4 weeks showed degeneration, edema, hyper trophy and vaculation in head-foot muscle, degeneration and necrotic changes in the digestive gland and accumulation in most tested organs. Also, the hermaphrodite gland showed mature ova with irregular shape and reduction in sperm number. In conclusion, the resulted damage and alterations in B. alexandrina studied parameters can be used as bioindicators to the presence of pollutants in its habitats.

Keywords: Biomphalaria, Zn, Cu, Pb, AST, ALT, ALP, total protein albumin, globulin, CAT and Histopathology.

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Authors: Anna Pick Kiong Ling, Jia May Chin, Rhun Yian Koh, Ying Pei Wong

Abstract:

Background: Uncontrolled inflammation may cause serious inflammatory diseases if left untreated. Non-steroidal anti-inflammatory drug (NSAIDs) is commonly used to inhibit pro-inflammatory enzymes, thus, reduce inflammation. However, long term administration of NSAIDs leads to various complications. Medicinal plants are getting more attention as it is believed to be more compatible with human body. One of them is a flavonoid-containing medicinal plants, Strobilanthes crispus which has been traditionally claimed to possess anti-inflammatory and antioxidant activities. Nevertheless, its anti-inflammatory activities are yet to be scientifically documented. Objectives: This study aimed to examine the anti-inflammatory activity of S. crispus by investigating its effects on intracellular oxidative stress and prostaglandin E₂ (PGE₂) levels. Materials and Methods: In this study, the Maximum Non-toxic Dose (MNTD) of methanol extract of both leaves and stems of S. crispus was first determined using 3-(4,5-dimethylthiazolyl-2)-2,5-diphenytetrazolium Bromide (MTT) assay. The effects of S. crispus extracts at MNTD and half MNTD (½MNTD) on intracellular ROS as well as PGE₂ levels in 1.0 µg/mL LPS-stimulated RAW 264.7 macrophages were then be measured using DCFH-DA and a competitive enzyme immunoassay kit, respectively. Results: The MNTD of leaf extract was determined as 700µg/mL while for stem was as low as 1.4µg/mL. When LPS-stimulated RAW 264.7 macrophages were subjected to the MNTD of S. crispus leaf extract, both intracellular ROS and PGE₂ levels were significantly reduced. In contrast, stem extract at both MNTD and ½MNTD did not significantly reduce the PGE₂ level, but significantly increased the intracellular ROS level. Conclusion: The methanol leaf extract of S. crispus may possess anti-inflammatory properties as it is able to significantly reduce the intracellular ROS and PGE₂ levels of LPS-stimulated cells. Nevertheless, further studies such as investigating the interleukin, nitric oxide and cytokine tumor necrosis factor-α (TNFα) levels has to be conducted to further confirm the anti-inflammatory properties of S. crispus.

Keywords: Anti-inflammatory, natural products, prostaglandin E2, reactive oxygen species.

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Authors: Norsakinah Mohammad Osman, Ali Etemad, Patimah Ismail

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Type 2 diabetes mellitus (T2DM) is a complex metabolic disorder that characterized by the presence of high glucose in blood that cause from insulin resistance and insufficiency due to deterioration β-cell Langerhans functions. T2DM is commonly caused by the combination of inherited genetic variations as well as our own lifestyle. Metallothionein (MT) is a known cysteine-rich protein responsible in helping zinc homeostasis which is important in insulin signaling and secretion as well as protection our body from reactive oxygen species (ROS). MT scavenged ROS and free radicals in our body happen to be one of the reasons of T2DM and its complications. The objective of this study was to investigate the association of MT1A and MT2A polymorphisms between T2DM and control subjects among Malay populations. This study involved 150 T2DM and 120 Healthy individuals of Malay ethnic with mixed genders. The genomic DNA was extracted from buccal cells and amplified for MT1A and MT2A loci; the 347bp and 238bp banding patterns were respectively produced by mean of the Polymerase Chain Reaction (PCR). The PCR products were digested with Mlucl and Tsp451 restriction enzymes respectively and producing fragments lengths of (158/189/347bp) and (103/135/238bp) respectively. The ANOVA test was conducted and it shown that there was a significant difference between diabetic and control subjects for age, BMI, WHR, SBP, FPG, HBA1C, LDL, TG, TC and family history with (P<0.05). While the HDL, CVD risk ratio and DBP does not show any significant difference with (P>0.05). The genotype frequency for AA, AG and GG of MT1A polymorphisms was 72.7%, 22.7% and 4.7% in cases and 15%, 55% and 30% in control respectively. As for MT2A, genotype frequency of GG, GC and CC was 42.7%, 27.3% and 30% in case and 5%, 40% and 55% for control respectively. Both polymorphisms show significant difference between two investigated groups with (P=0.000). The Post hoc test was conducted and shows a significant difference between the genotypes within each polymorphism (P=0. 000). The MT1A and MT2A polymorphisms were believed to be the reliable molecular markers to distinguish the T2DM subjects from healthy individuals in Malay populations.

Keywords: Type 2 Diabetes Mellitus (T2DM), Metallothionein (MT), MT1A (rs11076161), MT2A (rs10636), Malay, Genetic Polymorphism.

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Authors: Siti Aisya Gany, Swee Ching Tan, Sook Yee Gan

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Diminished antioxidant defense or increased production of reactive oxygen species in the biological system can result in oxidative stress which may lead to various neurodegenerative diseases including Alzheimer’s disease (AD). Microglial activation also contributes to the progression of AD by producing several proinflammatory cytokines, nitric oxide (NO) and prostaglandin E2 (PGE2). Oxidative stress and inflammation have been reported to be possible pathophysiological mechanisms underlying AD. In addition, the cholinergic hypothesis postulates that memory impairment in patient with AD is also associated with the deficit of cholinergic function in the brain. Although a number of drugs have been approved for the treatment of AD, most of these synthetic drugs have diverse side effects and yield relatively modest benefits. Marine algae have great potential in pharmaceutical and biomedical applications as they are valuable sources of bioactive properties such as anticoagulation, antimicrobial, antioxidative, anticancer and anti-inflammatory. Hence, this study aimed to provide an overview of the properties of Malaysian seaweeds (Padina australis, Sargassum polycystum and Caulerpa racemosa) in inhibiting oxidative stress, neuroinflammation and cholinesterase enzymes. These seaweeds significantly exhibited potent DPPH and moderate superoxide anion radical scavenging ability (P<0.05). Hexane and methanol extracts of S. polycystum exhibited the most potent radical scavenging ability with IC50 values of 0.157±0.004mg/ml and 0.849±0.02mg/ml for DPPH and ABTS assays, respectively. Hexane extract of C. racemosa gave the strongest superoxide radical inhibitory effect (IC50 of 0.386±0.01mg/ml). Most seaweed extracts significantly inhibited the production of cytokine (IL-6, IL-1 β, TNFα) and NO in a concentration-dependent manner without causing significant cytotoxicity to the lipopolysaccharide (LPS)-stimulated microglia cells (P<0.05). All extracts suppressed cytokine and NO level by more than 50% at the concentration of 0.4mg/ml. In addition, C. racemosa and S. polycystum also showed anti-acetylcholinesterase activities with the IC50 values ranging from 0.086-0.115 mg/ml. Moreover, C. racemosa and P. australis were also found to be active against butyrylcholinesterase with IC50 values ranging from 0.118- 0.287 mg/ml.

Keywords: Anticholinesterase, antioxidative, neuroinflammation, seaweeds.

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Authors: Chlopicka Joanna, Barton Henryk, Kryczyk Jadwiga, Francik Renata

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Recent studies demonstrated that high-fat diet increases oxidative stress in plasma and in a variety of tissues. Many researchers have been looking for natural products, which can reverse the effect of high fat diet. Recently, buckwheat is becoming common ingredient in functional food because of it properties. In study on buckwheat, it is known that, this plant plays roles as anti-oxidative, anti-inflammatory and anti-hypertensive. Nevertheless still little is known about buckwheat groats. The aim of this study was to investigate the effects of addition of buckwheat groats to the fat diet (30% lard), on some antioxidant and oxidant stress parameters in plasma and selected tissues in Wistar rats. The experiment was carried out with three months old male Wistar rats ca. 250g of body weight fed for 5 weeks with either a high-fat (30% of lard) diet or control diet, with or without addition of buckwheat groats. In plasma biochemistry and the activities of the antioxidant enzymes were measured selected tissues: glutathione peroxidase (GPX), catalase (CAT) and the levels of total and reduced glutathione (GSH), free thiol groups (pSH), antioxidant potential of plasma (FRAP) and oxidant stress indices - proteins carbonyl groups (CO) and malonyldialdehyde concentration (MDA). Activity of catalase (CAT) in plasma of rats was significantly increased in buckwheat groats groups and activity of GPx3 in plasma of rats was decreased in buckwheat groups as compared to control group. The reduced glutathione (GSH) in plasma of rats was significantly increased and protein CO was significantly decreased in buckwheat groups as compared to controls. The lowered concentration of GSH was found in serum of rats fed buckwheat groats addition but it accompanied in 7-fold increase in reduced-to-oxidized glutatione ratio, significant increase in HDL and decrease in nonHDL concentration. Conclusions: Buckwheat groats indicate a beneficial effect in inhibiting protein and lipid peroxidation in rats and improved lipid profile. These results suggest that buckwheat groats exert a significant antioxidant potential and may be used as normal food constituent to ameliorate the oxidant-induced damage in organism. 

Keywords: Antioxidant, buckwheat, high-fat diet, rats.

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Authors: Roya Bakhtiar, Alireza Abdolmohammadi, Hadi Hajarian, Zahra Nikousefat, Davood, Kalantar-Neyestanaki

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The objective of the present study was to determine the polymorphism in the leptin (332G>A) and its association with biometric traits in Sanjabi sheep. For this purpose, blood samples from 96 rams were taken, and tail length, width tail, circumference tail, body length, body width, and height were simultaneously recorded. PCR was performed using specific primer to amplify 463 bp fragment including exon 3 of leptin gene, and PCR products were digested by Cail restriction enzymes. The 332G>A (at 332th nucleotide of exon 3 leptin gene) that caused an amino acid change from Arg to Gln was detected by Cail (CAGNNNCTG) endonuclease, as the endonuclease cannot cut this region if G nucleotide is located in this position. Three genotypes including GG (463), GA (463, 360and 103 bp) and GG (360 bp and 103 bp) were identified after digestion by enzyme. The estimated frequencies of three genotypes including GG, GA, and AA for 332G>A locus were 0.68, 0.29 and 0.03 and those were 0.18 and 0.82 for A and G alleles, respectively. In the current study, chi-square test indicated that 332G>A positions did not deviate from the Hardy–Weinberg (HW) equilibrium. The most important reason to show HW equation was that samples used in this study belong to three large local herds with a traditional breeding system having random mating and without selection. Shannon index amount was calculated which represent an average genetic variation in Sanjabi rams. Also, heterozygosity estimated by Nei index indicated that genetic diversity of mutation in the leptin gene is moderate. Leptin gene polymorphism in the 332G>A had significant effect on body length (P<0.05) trait, and individuals with GA genotype had significantly the higher body length compared to other individuals. Although animals with GA genotype had higher body width, this difference was not statistically significant (P>0.05). This non-synonymous SNP resulted in different amino acid changes at codon positions111(R/Q). As leptin activity is localized, at least in part, in domains between amino acid residues 106-1406, it is speculated that the detected SNP at position 332 may affect the activity of leptin and may lead to different biological functions. Based to our results, due to significant effect of leptin gene polymorphism on body size traits, this gene may be used a candidate gene for improving these traits.

Keywords: Body size, Leptin gene, PCR-RFLP, Sanjabi sheep.

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