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   2020| October-December  | Volume 19 | Issue 4  
    Online since January 4, 2021

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Isolation, characterization, and screening of actinomycetes producing bioactive compounds from Egyptian soil
Ghadir E Daigham, Amira Y Mahfouz
October-December 2020, 19(4):381-390
Background Unsuitable prescription and the inappropriate use of antibiotics have led to the development of antibiotic resistance in many pathogenic bacteria. So, there is a need to screen for alternative sources, such as actinomycetes, a potent antibiotic producer for antimicrobial activity, with the hope of discovering novel strains proficient to produce antibiotics against the resistant bacteria. Materials and methods In the current work, six actinomycetes were isolated from Egyptian soil, characterized, and then identified by microscopic and macroscopic observations. The bioactive substances were extracted by the solvent extraction method using ethyl acetate. Antimicrobial activity of the obtained extracts was evaluated against some gram-positive and gram-negative bacteria. Fourier-transform infrared spectroscopy analysis and cytotoxic activity of the most active extracts were carried out. Results and conclusion Crude extract of A2 showed 30±0.10-mm mean inhibition zones against gram-positive bacteria Bacillus megaterium. Significantly, the isolate A6 showed the highest mean zone of inhibition of 20.0±0.1, 19.4±0.05, and 19.0±0.10 mm against Pseudomonas aeruginosa, Klebsiella oxytoca, and Escherichia coli, respectively. The isolate A5 only showed the largest antibacterial activity against B. megaterium, with inhibition zone of 69±0.28 mm. Only two isolates (A2 and A6) were chosen for further study based on broad-spectrum activity in comparison with other isolates. The two isolates A2 and A6 were identified as Streptomyces enissocaesilis MT658130 and Streptomyces atrovirens MT658195, respectively, using 16S rRNA. Fourier-transform infrared analysis of the extract of two strains reported the existence of OH, C=C, C-O, S=O, N=C=S, and C-Br as the most efficient groups. Cytotoxic activity of S. enissocaesilis A2 and S. atrovirens A6 extracts against hepatocellular carcinoma (HepG2) cell line showed good cytotoxicity, with 2.97 and 1.48 µg/ml IC50 value, respectively. The results evoked that actinomycete isolates under study have potent biological activities.
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Evaluation of antioxidant and antiapoptogenic effects of Sumach seed extract on liver injury induced by amethopterin in male rat model
Shiva Roshankhah, Yadollah Bahrami, Amir Abdolmaleki, Mohammad R Salahshoor
October-December 2020, 19(4):391-400
Background and objective Amethopterin (AM), like most chemotherapy medications, causes cellular generation of oxidative stress. In Iranian traditional medicine, Sumach is widely used owing to its antioxidant properties. In this study, the authors aimed to assess the probable antioxidant, antiapoptogenic, and therapeutic effects of Sumach extract on liver tissue and hepatocytes against hepatic degeneration induced by AM as a chemotherapy drug. Materials and methods A total of 64 rats were grouped (n=8), including (1) control, (2) AM (20 mg/kg), (3–5) Sumach (75, 100, 150 μg/ml), and (6–8)- AM+Sumach. Hydroalcoholic extract of Sumach seeds was prepared. Treatments with Sumach extract were applied intraperitoneally daily for 28 consecutive days. Nitrite oxide was measured by Griess assay. Ferric reducing ability of plasma (FRAP) and malonaldehyde test were used to measure total antioxidant and lipid peroxidation levels, respectively. Hepatocyte apoptotic index was analyzed via tunnel test. The expression level of p53 and Bax genes was measured by real-time PCR. Furthermore, liver enzymatic function, histopathological changes, and morphometric alterations were examined. Results and conclusion Antioxidant values, p53 and Bax genes’ expression, nitrite oxide, enzymes, and morphometrical features were increased significantly (except FRAP levels, decreased) in AM group as compared with the control group (P<0.01). Besides, the evaluated parameters were all significantly reduced (except insulin and FRAP levels and Bcl2 gene expression, which were increased) in SMGC and diabetic+SMGC groups in comparison with the diabetic group (P<0.05). Moreover, evaluated parameters were significantly reduced in Sumach and Sumach+AM groups (except FRAP level, which is increased) compared with the AM group (P<0.01). According to the acquired data, Sumach through its antiapoptogenic and antioxidant features is able to eliminate hepatotoxicity induced by AM administration, which leads to restoration of histopathological liver changes.
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Effect of sage oil on tamoxifen-induced hepatotoxicity and nephrotoxicity in female rats
Hagar H Mourad, Amira N Abd-El Razik, Eman N Hosny, Mayada M El-Gizawy
October-December 2020, 19(4):350-360
Background and objective Sage oil has anti-inflammatory and antioxidant activities. The current study was designed to evaluate the efficacy of sage oil at two different doses against hepatotoxicity and nephrotoxicity induced by tamoxifen in female rats. Materials and methods A total of 56 female Wistar albino rats were divided into control, sage control, tamoxifen-treated rats (45 mg/kg body weight), and rats treated with tamoxifen along with sage oil (0.2 ml/kg or 0.4 ml/kg). Each dose of sage oil was used as a protective (before tamoxifen) and as a therapeutic (after tamoxifen) treatment. At the end of the experiment, serum levels of tumor necrosis factor-alpha, as well as liver and kidney function biomarkers, were measured. Levels of lipid peroxidation (malondialdehyde), nitric oxide, and reduced glutathione and the activities of glutathione peroxidase and Na+/K+-ATPase were measured in liver and kidney homogenates. Sections of liver and kidney were examined for histopathological changes. Results and conclusion Tamoxifen-induced hepatic and renal impairment was evident from the significant elevation in alkaline phosphatase, aspartate aminotransferase, and alanine aminotransferase activities and bilirubin, urea, and creatinine levels. Furthermore, tamoxifen significantly increased serum tumor necrosis factor-alpha, as well as hepatic and renal malondialdehyde and nitric oxide levels. This was accompanied by a significant decrease in the levels of glutathione and the activities of glutathione peroxidase and Na+/K+-ATPase in liver and kidney. Treatment with sage oil at high dose (0.4 ml/kg) ameliorated almost all the biochemical and histopathological changes induced by tamoxifen in liver and kidney. In conclusion, sage oil has the ability to attenuate hepatotoxicity and nephrotoxicity induced by tamoxifen.
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New approach for production of saturated fatty acids in cassava cell cultures as antibreast cancer agent
Nermeen M Arafa, Usama I Aly, Mahmoud Moawad Elsayed, Mohamed K El-Bahr
October-December 2020, 19(4):361-370
Background and objective Cassava plant is one of the major economical crops, involved in many industrial applications and therapeutic purposes such as suppression of cancer cell activity. The present work targeted to assess saturated fatty acids and their derivatives in cassava cell cultures. Materials and methods Stem explants of in vitro growing plantlets were induced for calli on MS-medium supplemented with 1 mg/l naphthaleneacetic acid+0.5 mg/l benzylaminopurine. Medium containing 5 mg/l 2,4 dichlorophenoxyacetic acid (2.4-D) and 0.2 mg/l benzylaminopurine was selected for callus productivity. The extraction adequacy of aprotic polar (ethylacetate) and nonpolar (chloroform, n-hexane) solvents was analyzed and evaluated by gas chromatography–mass spectrometry. Results and conclusion Chloroform callus extract contained mostly fatty acid methyl esters and fatty acid propyl esters. In contrast, n-hexane extract contained higher amounts of fatty acid constituents in free form, such as palmitic acid (23.55%). Ethylacetate extract included the highest value of lauric acid (28.34%) in free form as well other fatty acids such as caprylic acid (14.525%), capric acid (2.53%), and enanthic acid (6.41%). Ethylacetate extract conferred the optimal efficiency to suppress the breast cancer cell prevalence (2.63 μg IC50), followed by hexanoic extract (3.44 μg IC50), and then chloroformic extract (6 μg IC50) recording the least value for cancer cell propagation. Potential of cassava as one of the medicinal valuable plants should be promoted for health-boosting purposes.
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Hepatoprotective agents as calcium channel blockers in paracetamol-induced and alcohol-induced hepatotoxicity models (curative method) in rats
Kola Venu, Errabelli Sujitha, Neha Samreen, Chamakuri S Rao, Munazza Fathima, Ahad H Syed, Vineela Yarasingi, Tula Kavya
October-December 2020, 19(4):371-380
Background It has been reported that accumulation of Ca++ ions in the cells causes depletion of mitochondrial respiration and ATP, synthesis leading to cell death. Furthermore, it has also been reported that paracetamol and alcohol produce hepatotoxicity by facilitating accumulation of excess Ca++ in hepatocytes. Ca++ channel blockers block the entry of Ca++ into the cells. Hence, the present study is planned to evaluate hepatoprotective activity of Ca++ channel blockers in the aforementioned models. Objective The aim was to study the hepatoprotective activity of three Ca++ channel blockers, namely, lercanidipine 1/4th TD (0.09 mg), ½ TD (0.18 mg) and 1TD (0.36 mg); felodipine 1/4 TD (0.045 mg), ½ TD (0.09 mg), and 1TD (0.18 mg); and isradipine ¼ TD (0.0225 mg), medium 1/2 TD (0.045 mg), and 1TD (0.09 mg), with three selected doses in paracetamol-induced and alcohol-induced hepatotoxicity in rats with curative aspects. Materials and methods The hepatoprotective activity of Ca++ channel blockers is evaluated in PCM- (2 g/kg) and ALC-(3.76 g/kg) induced hepatotoxic models in rats. The study recorded thiopental-induced sleeping time; physical parameters like liver weight and liver volume; and biochemical parameters, like alanine transaminase, aspartate aminotransferase, alkaline phosphatase, direct bilirubin, total bilirubin, albumin, and total protein, which were estimated by using a semiauto analyzer. Silymarin is used as a standard reference drug in both the paracetamol-induced and alcohol-induced hepatotoxic models. Standard drug silymarin produced a significant hepatoprotective activity. Results Ca++ channel blockers too at three different doses as mentioned before with curative aspect produced a significant hepatoprotective activity in paracetamol-induced and alcohol-induced hepatotoxic models. Ca++ channel blockers cured the hepatotoxic effects of both paracetamol and alcohol in rats and offered hepatoprotective activity. Conclusion Lercanidipine, felodipine, and isradipine exhibited a significant hepatoprotective activity in both paracetamol-induced and alcohol-induced hepatotoxic models in rats.
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Eco-friendly and superficial approach for synthesis of silver nanoparticles using aqueous extract of Nigella sativa and Piper nigrum L seeds for evaluation of their antibacterial, antiviral, and anticancer activities a focus study on its impact on seed germination and seedling growth of Vicia faba and Zea mays
Amira Y Mahfouz, Ghadir E Daigham, Asmaa M Radwan, Abeer A Mohamed
October-December 2020, 19(4):401-413
Background Silver nanoparticles (AgNPs) have fascinated extensive interest owing to their potential applications, especially in biomedicine. Materials and methods Through this investigation, AgNPs were green synthesized from AgNO3 using Nigella sativa and Piper nigrum aqueous seed extracts as a reducing agent. Biosynthesized AgNPs were characterized using transmission electron microscopy, Fourier-transform infrared spectroscopy, zeta potential, and dynamic light scattering. Antibacterial, antiviral, and anticancer activities of AgNPs were studied. Furthermore, the effect of biosynthesized AgNPs on seed germination and seedling growth of Vicia faba and Zea mays was determined. Results and conclusion Results evoked the ability of N. sativa and P. nigrum aqueous seed extracts to build up AgNPs in spherical shape with average size of 20 and 50 nm, respectively, by using transmission electron microscopy analysis. The results indicate the ability of N. sativa and P. nigrum AgNPs to inhibit both tested gram-positive and gram-negative bacteria. It is conceivable from the results that AgNPs of the N. sativa and P. nigrum are markedly effective against herpes simplex virus-1 in terms of decreased viral load and mortality, exhibiting 83.23 and 94.54% of antiviral activity, respectively. Furthermore, in-vitro studies of N. sativa and P. nigrum AgNPs against hepatocellular carcinoma have shown good cytotoxic effect, with IC50 values equal to 7.12 and 4.98 µg/ml, respectively. AgNPs exert a promoting action on seed germination percentage and seedling growth of both Vicia faba and Zea mays. This study indicated an economical, simple, and efficient ecofriendly technique using aqueous seed extracts of N. sativa and P. nigrum L. for synthesis of AgNPs and confirmed that green AgNPs are safer.
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Optimization of riboflavin production by Rhodotorula glutinis using statistical design
Hassan S Hassan, Yousseria M Shetaia, Mona S Shafei, Heba A Elrefai, Mahmoud M Elaasser
October-December 2020, 19(4):312-320
Background and objective Riboflavin is considered a significant nutritional and growth factor in humans, animals, plants, and microorganisms. This water-soluble vitamin is an intrinsic component of the basic metabolic processes and is a precursor of the oxidation-reduction coenzymes, flavin mononucleotide and flavin adenine dinucleotide. Riboflavin was synthesized by different microorganisms such as Ashbya gossypii, Eremothecium ashbyii, Candida famata, and Bacillus subtilis. The factors controlling riboflavin production, such as environmental and nutritional factors, were implemented to increase the yield of riboflavin. A statistical experimental design, such as the Plackett–Burman, enables the finding out of the most effective factors that increase the yield of riboflavin. Materials and methods One-factor-at-a-time approach was used to evaluate the effect of different nitrogen and carbon sources for riboflavin production by a local yeast isolate Rhodotorula glutinis. To optimize the riboflavin production by R. glutinis, the combined effects of seven variables (lactose, yeast extract, KH2PO4, MgSO4, time, pH, and rpm) were assessed using the Plackett–Burman design. Results and conclusion A preliminary medium formulation trial suggested that yeast extract and lactose were the appropriate nitrogen and carbon sources, respectively. The two-level Plackett–Burman design was implemented to study the nutritional and environmental factors for riboflavin production. Among the seven variables tested, lactose, KH2PO4, and rpm were identified as the most significant factors. The optimal levels of the three variables were determined by response surface methodology based on the Box–Behnken design. The validity of the model developed was verified, and the maximum riboflavin concentration was 88.25 μg/ml, representing 1.27 folds higher in the improved medium.
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Optimization and molecular characterization of novel Aspergillus spp. producing invertase enzyme
Rasha G Salim, Nivien A Abo-Sereh, Bigad E Khalil
October-December 2020, 19(4):321-329
Background and objective Invertase, one of the most important enzymes used in industrial and pharmaceutical applications, has been produced from different Aspergillus spp. Statistical and mathematical tools [response surface methodology (RSM)] have been carried out to optimize culture conditions to facilitate maximum enzyme production. This investigation aims to screen the ability of some Aspergillus spp. to produce invertase enzyme, to optimize the culture parameters, and to detect the genetic diversity between them. Materials and methods A total of five fungal strains were tested for the production of invertase. The most promising strain was selected for optimization. Optimization experiments were carried out to obtain maximum enzyme production, and a molecular marker (RAPD-PCR) was used to differentiate between these strains. Results and conclusion Some Aspergillus spp. were examined to produced invertase enzyme after 2 days of cultivation. Aspergillus niger recorded the highest enzyme production (6.8 U/ml). Ten RAPD-PCR primers were used to differentiate between Aspergillus spp. A total of 131 bands were amplified in the five strains of Aspergillus spp. Based on RAPD-PCR analysis, the highest similarities obtained between A. parasiticus and A. niger showed 75%. Moreover, the RAPD-PCR technique is a good tool for differentiation between the Aspergillus spp. To optimize culture condition, RSM was used to increase invertase production by A. niger. Temperature, pH, and incubation time dramatically affect invertase yield. A combination of these factors (pH 5, 35°C, and incubation for 72 h) was optimum for maximum production of invertase (8.2 U/ml). Our results employed RSM to elucidate the combination of effective factors such as temperature, pH, and incubation time to obtain a high yield of an important enzyme, invertase, from A. niger, used as a promising candidate for industrial applications.
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Effect of Annona squamosa Linn against aluminium chloride induced Alzheimer’s disease in rats
Kanala Somasekhar Reddy, Bhupalam Pradeepkumar, Akkiraju Sudheer, Mallela Vijayajyothi
October-December 2020, 19(4):330-337
Background and objective Alzheimer’s disease (AD) is an age-related neurodegenerative disorder characterized by memory deficits. Various studies have been conducted to find therapeutic approaches for AD. However, a suitable treatment alternative is still not available. The present study is aimed to investigate the polyphenolic fraction of Annona squamosa Linn (PFAS) against aluminium chloride (AlCl3)-induced AD in rats. Materials and methods Wistar rats were divided into group I (normal control), which received distilled water; group II, which received AlCl3 (100 mg/kg, intraperitoneal route); groups III and IV received PFAS (200 and 400 mg/kg, oral, respectively) and inducing agent (AlCl3 100 mg/kg, oral); and group V received donepezil (1 mg/kg, oral) and inducing agent (AlCl3 100 mg/kg, oral). The rats were given respective treatment for 28 days, and behavioral parameters were determined on first day, 15th, and 28th day. After 28th day, rats were killed, and antioxidant parameters and brain acetylcholinesterase content were determined, and histopathological studies were done. Results and conclusion The PFAS showed dose-dependent protective effect against AD by significant improvement in locomotor activity, motor coordination, spatial memory, and conditioned avoidance response; significant decrease in lipid peroxidation and acetyl cholinesterase; and increase in antioxidants compared with AlCl3-treated rats. PFAS mitigated the AlCl3-induced histological changes by dose dependently. PFAS showed potent neuroprotective effect against AlCl3-induced oxidative stress in rats. Hence, it would be promising compound to treat AD.
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Enhanced production, partial purification, and characterization of alkaline thermophilic protease from the endophytic fungus Aspergillus ochraceus BT21
Eman W Elgammal, Mohamed I El-Khonezy, Eman F Ahmed, Ahmed M Abd-Elaziz
October-December 2020, 19(4):338-349
Backgroundand objective Endophytic fungi are thought to be a potential source for biologically active compounds such as enzymes, especially proteases which find their application in modern biochemical industries. The aim of this study was to optimize the production conditions of protease from Aspergillus ochraceus BT21, which was previously isolated from the Egyptian medicinal plant Ruprechita salicifolia. The produced protease was optimized, partially purified and characterized. Materials and methods A. ochraceus BT21 was identified by 18 S rRNA under the accession number of MN564896 in gene bank. The physicochemical parameters of the fermentation medium were optimized. Furthermore, the harvested protease was concentrated and partially purified by ethanol fractionation and then characterized to detect the enzyme identity. Results and conclusion The protease production increased by about 7.5-fold (3644.9 U/mg) after applying the final optimized fermentation medium, which contains dextrin 30, peptone 2, K2HPO4 1, MgSO4 0.5, KCl 0.5, and FeSO4 0.01(g/l) at 35°C, pH 8.0 and 150 rpm using 6% inoculum size after 6 days of incubation. The purification results showed that the highly recovered fraction was at 60% ethanol concentration with a purification fold of 4.3 and enzyme recovery of 36.5%. The enzyme was thermotolerant with an optimum temperature of 50°C and optimum pH of 8.0. Furthermore, it was observed that there was a reverse relationship between the metal ion concentration and the enzyme relative activity. Finally, the data indicated that casein and human blood were the most suitable substrates for this enzyme, indicating that the enzyme can work in alkaline conditions and has thermotolerance properties with high affinity toward the blood substrate, which makes it a potential candidate for detergent formulation to facilitate blood stain removal.
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Slime fungi: the brainless super microorganism
Marwa O Elnahas, Waill A Elkhateeb, Ghoson M Daba
October-December 2020, 19(4):307-311
Myxomycetes, commonly known as plasmodial slime molds, are unique organisms which take an intermediate position between plant and animal kingdoms. They can form a jelly-like plasmodia which feed on bacteria and are able to move by a synchronized perpendicular flow of their protoplasm. Slime molds can be classified into three groups: cellular, plasmodial, and net slime molds. These interesting organisms are novel sources of several bioactive secondary metabolites with anticancer, antioxidant, and antimicrobial activities. In this review, the ecology, occurrence, and secondary metabolites secreted by these unique organisms are highlighted.
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