EVALUATION OF α-AMYLASE, α-GLUCOSIDASE AND ALDOSE REDUCTASE INHIBITORS IN ETHYL ACETATE EXTRACTS OF ENDOPHYTIC FUNGI ISOLATED FROM ANTI-DIABETIC MEDICINAL PLANTSHTML Full Text
EVALUATION OF α-AMYLASE, α-GLUCOSIDASE AND ALDOSE REDUCTASE INHIBITORS IN ETHYL ACETATE EXTRACTS OF ENDOPHYTIC FUNGI ISOLATED FROM ANTI-DIABETIC MEDICINAL PLANTS
N. Pavithra1, L. Sathish1, Nagasai, Babu2, V. Venkatarathanamma2, H. Pushpalatha3, G. Bhanuprakash Reddy3and K. Ananda *1
Biological Sciences1, Poornaprajna Institute of Scientific Research, Devanahalli, Bangalore 562110, India.
Department of Zoology & Biotechnology2, Acharya Nagarjuna University Nagarjunanagar-522510, A.P, India
Biochemistry Division3, National Institute of Nutrition, Hyderabad, India.
ABSTRACT:Twenty two endophytic fungi were isolated from anti-diabetic plants Momordica charantia and Trigonella foenum-graceum. Ethyl acetate extracts of all fungal isolates were tested for inhibition of aldose reductase and carbohydrate metabolizing enzymes. Ethyl acetate extracts of nine endophytic fungi were found to be positive for α- amylase and α- glucosidase inhibitors. Crude extracts of fungal isolates PTFL005 and PTFL006 showed promising inhibition activity on α- amylase with an IC 50 value of 15.48 and 13.48μg/mL respectively. Whereas, the control acarbose had 22.38 μg/mL of IC 50 value for α- amylase at similar experimental condition. Fungal isolates PTFL006 and PTFL011 were found to have potent α- glucosidase inhibitors with an IC 50 value of 17.37 and 10.71μg/mL which was close to the standard acarbose (6.53μg/mL). The only one fungal isolate PMCF003 showed reasonably better inhibition for aldose reductase with an IC50 value of 40μg/mL as compared to standard control quercetin (15μg/mL). All the active endophytic fungal isolates were identified by DNA sequencing method. The most active isolates identified asTrichoderma atroviride (PMCF003) and Stemphylium globuliferum (PTFL005, PTFL006 and PTFL011) were reported for the first time as endophytes in these plants from India.
INTRODUCTION: Hyperglycemia defined as a heterogeneous disease caused due to deficiency of insulin secretion and action1. International Diabetes Federation has reported that 8.3 % (366 million) of adults were infected with diabetes in the year 2011 and this number may rise up to 552 million people by the year 2030 2. Post prandial hyperglycemia is caused due to action of two enteric enzymes (α-amylase and α-glucosidase) that are attached to the brush border of the intestinal cells.
Therefore inhibition of these two enzymes leads to slow rate of carbohydrate digestion and glucose absorption3, 4. The known glucosidase inhibitors commercially available are acarbose and miglitol that competitively and reversibly inhibits α-glucosidase enzyme from intestine as well as pancreas, but these synthetic drugs possess side effects such as liver disorders, flatulence, abdominal pain, renal tumours, hepatic injury, acute hepatitis, abdominal fullness and diarrhoea5-7.
About 26.1 % of Indian population are found to be suffering from diabetic neuropathy which is an important complication of diabetes mellitus 8. Aldose reductase is the first and rate-limiting enzyme in the polyol pathway that reduces glucose to sorbitol, inhibition of aldose reductase would delay or substantially prevent these secondary diabetic complications as studied in animal models and also evaluated in clinical trials 9. High levels of aldose reductase have been detected in the cornea, lens, kidney and myelin sheath 10.
The medicinal plants such as Momordica charantia (MC) commonly known as Bitter gourd and Trigonella foenum-graceum (TF) commonly known as Menthya has been used in Indian herbal medicine for diabetic control.
Recent studies had shown that endophytic fungi which are living inside the plants have the ability to produce identical or similar bioactive compounds as their host plants 11, 12. Endophytic fungi are found to be promising sources of secondary metabolites and they also play an important role in drug discovery13. In recent years endophytes are considered as outstanding source of secondary metabolites with pharmaceutically important bioactive compounds. The aim of present study was to isolate endophytic fungi from medicinal plants MC and TF to evaluate the antidiabetic activity of secondary metabolite.
MATERIALS AND METHODS:
Isolation of Endophytic fungi
Fresh samples of Momordica charantia (fruit) and Trigonella foenum-graceum (leaf and twig) were collected from the nearby market located at Devanahalli, Bangalore, India. Plant materials were collected in a sterile polythene bag and processed within one hour of sample collection to avoid contamination. Samples were thoroughly washed under running tap water to remove debris and cut into 1 cm long.
In case of bittergourd, seeds were removed and pericarp is used for the isolation. Surface of samples sterilized sequentiallywith70%ethanolfor1.5min followed by 1.0 %sodiumhypochlorite(NaOCl)(v/v)for2minandfurther cleaned bypassingthroughtwosetsofsteriledistilledwater. Thesterilesampleswereplacedonpotatodextroseagar(PDA)media containing200mg/Lofstreptomycin antibiotic in petriplate. Theparafilmwrappedpetriplateswereincubatedat room temperaturetillthefungalmyceliastartsgrowingfromthesample.The isolated endophyticfungiwastransferredintoanewagarslantsandstoredat4°Cforthefurtherstudies 14.
Fermentation and extraction of secondary metabolites
Two or three discs of fresh mycelium grownfrom mother culture was inoculated into 500 mL Erlenmeyer flask containing 300 mL YPF(Yeast extract- 3g, Peptone- 10g, Fructose- 20g, Distilled water -1000mL, pH-6) and incubated at room temperature for 21 days under stationary conditions with intermittent shaking. After incubation, ethyl acetate (1:1 ratio) was added to flask containing media and kept on shaker (150 rpm) for overnight. The mycelial mass was separated from the broth by filtration and macerated with ethyl acetate to get intracellular secondary metabolites. The organic phase of extraction mixture was separated and evaporated to dryness before storing at 4°C for further analysis.
α-amylase inhibition Assay
100 μL of the porcine pancreatic amylase (PPA) solution (1.1 U) was added to 100 μL of test substance and incubated at room temperature (RT) for 20 minutes. The reaction was initiated by the addition of 100 μL of 1% soluble starch solution and incubated at 37°C. The reaction was arrested after 10 min of incubation by the addition of 200 μL of dinitrosalicylic acid colour reagent. The tubes were kept in a boiling water bath for 10 minutes, cooled and diluted with distilled water and absorbance was measured at 470 nm 15. Acarbose was used as the standard alpha-amylase inhibitor and vehicle was used as negative controls. All the assays were carried out in triplicates and average percent inhibition of enzymes by the fungal extract was calculated using the following formula.
12% Inhibition=Control OD-Test ODControl ODÃ—100">
α-Glucosidase inhibition Assay
Alpha glucosidase inhibition activity of ethyl acetate extract of fungal isolates were measured in microplate wells using the method explained by Suneel et al 16. α –glucosidase enzyme solution was prepared in PBS (pH 7.0) and diluted to 0.4 U/mL of working solution. The test solution was prepared by mixing 50μL of enzyme with 10μL of fungal extract and diluted to 150μL with 100mM phosphate buffer pH 7.0 and mixed well. The reaction mixture was incubated at RT, after 5 min 50μL of substrate p-nitro phenyl α-D-glucopyranose (20mM) was added. The reaction mixture was mixed well before incubating for 15 min at RT, later 30μL of sodium carbonate solution (200mM) was added to stop the reaction. The optical density of a yellow coloured product measure at 405nm using a microplate reader. Control and test blank OD’s were obtained by replacing enzyme with buffer17.
Preparation of rat lens aldose reductase (ALR2)
Crude aldose reductase was prepared from 2-3 month old WNIN male rate lens by dissecting its eye balls. The dissected lenses were homogenized and diluted 10 times using 100 mM Phosphate buffer pH6.2. the homogenized sample was centrifuged at 15000 x g for 30 min at 4°C and the supernatant was used as enzyme source (ALR2)18. The protocol was followed as per the animal ethical committee approval in NIN, Hyderabad, India.
Aldose reductase enzyme assay
Aldose reductase enzyme activity in the lens extract was carried out as described by Hayman and Kinoshita 19. The 1 mL of assay mixture contained 50 µmol of potassium phosphate buffer pH 6.2, 0.2 M lithium sulfate, 5 µmol β-mercapto ethanol, 10 μmol DL-glyceraldehyde and enzyme preparation (rat lens). The assay mixture was incubated at 37oC and initiated reaction by the addition of 0.1 μmol of NADPH. The rate of oxidation of NADPH was measured at 340nm using a spectrophotometer (Lamda 35, Perkin-Elmer, Shelton, U.S.A.). Specific activity of ALR2 was expressed in µmoles of NADPH oxidized /h/100 mg protein.
Aldose reductase inhibition assay
For inhibition studies concentrated stocks of crude extract was prepared in DMSO were used and the ﬁnal concentration of DMSO was not more than 1%. Various concentrations of extracts were added to assay mixtures of ALR2 and incubated for 5 min before initiating the reaction by NADPH as described above. The percentage inhibition was calculated considering the activity in the absence of extract as 100%. The IC50 values were determined by non-linear regression analysis of the plot of percentage inhibition versus log concentration 20.
Identification of Endophytic fungi:
Endophytic fungi were transferred to potato dextrose broth and incubated at 30°C for 7 days. After 7 days of incubation, mycelium was collected by filtration and DNA was extracted as the protocol described by Lu Y et al21. Purified DNA was subjected to PCR amplification using primers ITS1 and ITS4. PCR was done in 25µl reaction containing 50ng genomic DNA, 10 x PCR buffer (2.5μL) , 10 mM dNTP (0.5μL) , 25 mM MgCl2 (3.5μL), 10 µM each primer (0.5μL), 250 unit Taq DNA polymerase (0.2μL). PCR was carried out in a thermal cycler (TECHNE) with an initial denaturation of 95º C for 5 min, followed by 35 cycles of 95º C for 30 s, 52º C for 45 s, 72º C for 90 s and a final extension at 72º C for 10 min.
Amplicon DNA was purified using Qiagen columns and submitted to Eurofins, Bangalore, India. Automated DNA sequencing was performed using an ABI 3730xl 96-capillary DNA Analyzer Machine. The sequence data was analyzed using the BLAST software (BLASTN) available at the National Center of Biotechnolgy information (NCBI) web site (http://wwwncbinihgov/) to determine the identity of the endophyte. Gene sequences were submitted to genebank and accession numbers were obtained.22
Isolation of endophytic fungi and extraction of secondary metabolites
A total of 11 endophytic fungi were isolated from bitter gourd fruits and 11 from menthya leaves. All the isolates of menthya are from leaves and none from any of the twigs part. All the endophytic fungi are subcultured in to new agar plates and stored in multiple methods such as agar plates, agar slants and sterile distilled water. Twenty two fungal isolates were grown in large scale separately and extraction of secondary metabolites in ethyl acetate solvent yield about 0.5 to 1.0gm/L of dry powder per fungi.
Alpha-amylase and α-glucosidase inhibition assay:
All the 22 endophytic fungal extracts were tested for PPA inhibitors and only 9 of them have shown significant inhibition for both enzymes. Percentage inhibitions of PPA by different fungal isolates at various concentrations was calculated and plotted in the Figure 1 for both MC and TF. Crude extract of nine endophytic fungi isolate PTFL005 and PTFL006 have IC50 values of 15.48 and 13.48μg/mL respectively. Both highl active isolates are isolated from the leaf part of the menthya plant. The IC50 value of these two fungal extract was less than IC50 value of standard drug acarbose (22.38μg/mL) when assayed under similar condition. The IC50 values for all the fungal extract was calculated along with the standard anti-diabetic drug acarbose as control and shown in the Figure 2.
FIGURE 1: PERCENT INHIBITION OF α-AMYLASE WITH DIFFERENT CONCENTRATION OF FUNGAL EXTRACT.
Fungal isolates PMCF003, PMCF011 and PTLF001 to PTLF011 compared with standard Acarbose
FIGURE 2: IC50 VALUES OF FUNGAL EXTRACT AGAINST α AMYLASE ENZYME.
Fungal isolates PMCF003, PMCF011 and PTFL001 to PTFL011 compared with standard Acarbose.
The same set of 22 endophytic fungal extracts tested for α-amylase inhibitors were also tested for α-glucosidase inhibitors. Ethyl acetate extracts of nine isolates had shown very good inhibition activity for α-glucosidase enzyme. The percentage inhibition of glucosidase enzyme by the different concentrations of crude extract was measured and plotted against the concentration as shown in Figure 3.
FIGURE 3: PERCENT INHIBITION OF α -GLUCOSIDASE WITH DIFFERENT CONCENTRATION OF FUNGAL EXTRACT.
Fungal isolates PMCF003, PMCF011 and PTLF001 to PTLF011 compared with standard Acarbose
Two isolates PTFL006 and PTFL011 had IC50 values of 17.37 and 10.71μg/mL respectively when compared to acarbose IC50 value (6.53μg/mL) under similar experimental condition. These two most active isolates were also from leaves of the menthya plant. The IC50 values calculated for all the active fungal extracts along with the standard acarbose and plotted against concentration of crude extracts as shown in the Figure 4.
FIGURE 4: IC50 VALUES OF FUNGAL EXTRACT AGAINST α-GLUCOSIDASE ENZYME.
Fungal isolates PMCF003, PMCF011 and PTFL001 to PTFL011 compared with standard Acarbose
Aldose reductase inhibition assay
Out of 22 endophytic fungi isolated, ethyl acetate extracts of ten endophytic fungi (PMCF001, PMCF002, PMCF003, PMCF006, PMCF008, PMCF011, PTFL005, PTFL006 and PTFL011) were tested for aldose reductase inhibitors (ALR2). Only three extracts (PMCF001, PMCF003 and PTFL006) were found to be positive for aldose reductase inhibitors. Percentage inhibition of rat lens aldose reductase enzyme by fungal extract at different concentrations estimated and plotted against concentration along with the quercetin as standard (Figure 5).
FIGURE 5: PERCENT INHIBITION OF ALDOSE REDUCTASE ENZYME WITH DIFFERENT CONCENTRATION OF FUNGAL EXTRACT.
Fungal isolates PMCF001, PMCF003 and PTLF006 compared with standard Quercetin
The IC50 values for these endophytic fungi against ALR2 was determined and represented in the Figure 6. PTFL006, PMCF001 and PMCF003 showed aldose reductase inhibition with an IC50 value of 100, 78 and 40 μg/ml respectively. PTFL005, PTFLl006 and PTFL011 have shown percentage inhibition against aldose reductase at the concentration range 0-100 μg/mL with 46, 51 and 35 % respectively.
FIGURE 6: IC50 VALUES OF FUNGAL EXTRACT AGAINST ALDOSE REDUCTASE ENZYME.
Fungal isolates PMCF001, PMCF003 and PTLF006 compared with standard Quercetin
Molecular identification of endophytic fungi
The endophytic fungal isolates which showed inhibition for at least one of the three enzymes α-amylase, α- glucosidase or aldose reductase inhibitors were identified by rDNA molecular method. Conserved DNA region for all these fungal isolates was purified, amplified and sequenced for their identification. The sequence were submitted to the Gene Bank and obtained Gene Bank accession number. Based on the sequence similarity closest matching in the blast search was selected and identified the fungal isolates. The identified species, gene bank accession number and their percent nucleotide homology with the existing data base are represented in the Table 1.
TABLE 1: LIST OF ENDOPHYTIC ISOLATES IDENTIFIED BY rDNA METHOD
|Sl No.||Endophytic fungi code||Identified
|Gene bank accession number||Molecular Identity
(closest match in GenBank)
(closest match in GenBank)
|Nucletide homology (%)|
|2||PMCF003||Trichoderma atroviride||KC702784||Trichoderma atroviride strainTUCIM N154||JN387049.1||99|
|3||PMCF011||Trichoderma atroviride||KF911101||Trichoderma atroviride T-26||KC884783.1||100|
|4||PTFL001||Stemphylium lycopersici||KF911102||Stemphylium lycopersici||AB704312.1||99|
|5||PTFL002||Stemphylium globuliferum||KF911103||Stemphylium globuliferum||KF479193.1||100|
|6||PTFL003||Alternaria sp PEGT001||KF911104||Alternaria sp PEGT001||KC707558.1||100|
|7||PTFL004||Stemphylium lycopersici||KF911105||Stemphylium lycopersici||AB704312.1||97|
|8||PTFL005||Stemphylium globuliferum||KF911106||Stemphylium globuliferum||KF479193.1||100|
|9||PTFL006||Stemphylium globuliferum||KF911107||Stemphylium globuliferum||KF479193.1||100|
|10||PTFL011||Stemphylium globuliferum||KF911108||Stemphylium globuliferum||KF479193.1||100|
All the three endophytic isolates PMCF001, PMCF003 and PMCF011 from M. charantia were identified as Trichoderma atroviride with 99-100% sequence homology. The fungal isolates PTFL002, PTFL005, PTFL006 and PTFL011 from Menthya plant are identified as Stemphylium globuliferum having sequence similarity of 99-100% with the existing genebank data. The isolate PTFL003 which was identified as Alternaria sp. showed 100% sequence homology. PTFL001 and PTFL004 identified as Stemphylium lycopersici with 99% and 97% sequence homology respectively.
DISCUSSION: Several studies have reported the use of herbal extracts as antidiabetic, however these extracts have not proved to be successful as medicines due to lack of specific standards being prescribed for herbal medicines and supportive animal/clinical trials 23. Aqueous methanol extracts of M.charantia seeds have shownan α-glucosidase inhibitory activity 24. There was a significant decrease (p<0.01) in the fasting blood glucose levels in streptozotocin (STZ) induced diabetic rats treated with the MC extract, as compared to the untreated diabetic rats.
M. charantia extract known to exhibit anti-hyperglycemic effect in the streptozotocin or alloxan-induced diabetic rats 25 26. The compound 4-hydroxyleucine present in the seeds of TF have proven to stimulate insulin production 27. It has been reported that there was a significant reduction in blood glucose level in alloxan-diabetic rats treated with aqueous extract of fenugreek leaf as compared to healthy rats 28.
In the present study we have isolated endophytic fungi from these two plants and evaluated their antidiabetic activity. Endophytic fungi have the capability to produce bioactive compounds such as alkaloids, terpenoids, steroids, quinones, lignans, phenols and lactones 29. In a study six endophytic fungi were isolated from Swietenia macrophylla King seeds and five ethyl acetate extracts had showed better glucosidase inhibition activity than acarbose with lowest IC50 values was 73.64 µg/mL. These α-glucosidase inhibitor was found to be belongs to flavonoid group 30. Structure activity relationship studies have proved that ganoderma acids isolated from Ganoderma lingzhi as a potentα-glucosidase inhibitor 31. In the present study crude ethyl acetate extract of two isolates belonging to Stemphylium globuliferum species isolated from Menthya leaf showed highest PPA inhibitory activity with an IC50 values of 15.48 and 13.48μg/mL. These two crude extracts IC50 values are far below the IC50 value of standard drug acarbose (22.38 µg/mL) under similar experimental condition, which indicates these isolates has higher potential for production of pharmaceutically important drug if purified further. There are three isolates belonging to the Stemphylium globuliferumspecies which also showed very good α-glucosidase inhibition activity. The IC50 value for these are ranging from 10-18 µg/mL which is very near to the standard acarbose (6.53 µg/mL) tested under similar in-vitro condition.
Endophytic fungus B.Os.1F isolated from Kumis kucing (Orthosiphon spicatus BBS) showed highest inhibition percentage with 93.91% from filtrate extract and 89.01% from biomass extract. Endophytic fungi isolated from Mengkudu (Morinda citrifolia L.), Sirih Merah (Piper crocatum L.), Sirih Hitam (Piper ornatum L.) have shown alpha glucosidase inhibition activity 32. Aqueous extracts of bitter gourd and fenugreek have shown aldose reductase inhibitory activities with IC50 values <0.5 mg/mL33.
As per the proposal of this study, the fungi isolated from Bitter gourd have shown the best aldose reductase inhibition activity at crude extract level. The best aldose reductase inhibition was shown by the endophytic fungi Trichoderma atroviride isolated from Bittergourd.
To our present knowledge there are no reports on Stemphylium sp having α-amylase and α-glucosidase inhibitors. This is the first study on Stemphylium sp isolated from menthya and proved to have a potent antidiabetic agent. Trichoderma sp isolated from bittergourd found to be the potent aldose reductase inhibitors with an IC50 value of 40 μg/mL. The endophytic microorganisms are excellent sources of bioactive natural products having great demands in health care sector34. Therefore exploration of these endophytic fungi having the capability to produce antidiabetic compounds will lead to revolution in biotechnology industry.
CONCLUSIONS: Present study concludes that Stemphylium globuliferum is one of the best endophytic fungi having both α-amylase inhibition and α-glucosidase inhibition activity and this is the first report from India. The crude extract having the α-amylase inhibition activity more than the standard available drug acarbose is of very interesting and promising result. The α-glucosidase inhibition activity of the same fungal species were also very near to the standard acarbose value indicates that there is a better hope in optimization of secondary metabolite production and purification of compounds for the pharmaceutical applications.
ACKNOWLEDGEMENTS: Authors PN, SL and AK would like to thank Admar Mutt Education Foundation for the financial support in conducting this research. SL and PN are thankful to Manipal University for their PhD registration.
- Tundis R, Loizzo M, Menichini F. Natural products as α-amylase and α-glucosidase inhibitors and their hypoglycaemic potential in the treatment of diabetes: an update. Mini reviews in medicinal chemistry. 2010;10(4):315-331.
- Federation ID. IDF diabetes atlas: International Diabetes Federation, Executive Office; 2011.
- Jing M, Rayner C, Jones K, Horowitz M. Insulin secretion in healthy subjects and patients with Type 2 diabetes-role of the gastrointestinal tract. Best Practice & Research Clinical Endocrinology & Metabolism. 2009;23(4):413-424.
- Powers AC, D’alessio D. Endocrine pancreas and pharmacotherapy of diabetes mellitus and hypoglycemia. Goodman & Gillman’s the pharmacological basis of therapeutics. 12th ed. McGraw Hill. 2011:1237-1273.
- El-Kaissi S, Sherbeeni S. Pharmacological management of type 2 diabetes mellitus: An update. Current diabetes reviews. 2011;7(6):392-405.
- T. Fujisawa HI, K. Inoue, Y. Kawabata, and T.Ogihara, . “Eﬀect of two alpha-glucosidase inhibitors, voglibose and acarbose, on postprandial hyperglycemia correlates with subjective abdominal symptoms,” Metabolism. 2005;54:387-390
- S.K.Singh PKR, D.Jaiswal,andG.Watal. “Evidence based critical evaluation of glycemic potential of Cynodon dactylon,”. Evidence-Based Complementary and Alternative Medicine. 2007; 6( 4): 415–420.
- Pradeepa R, Rema M, Vignesh J, Deepa M, Deepa R, Mohan V. Prevalence and risk factors for diabetic neuropathy in an urban south Indian population: the Chennai Urban Rural Epidemiology Study (CURES‐55). Diabetic Medicine. 2008;25(4):407-412.
- Reddy GB, Muthenna P, Akileshwari C, Saraswat M, Petrash JM. Inhibition of aldose reductase and sorbitol accumulation by dietary rutin. Current Science. 2011;101(9):1191-1197.
- Kim HM, Lee JM, Choi K, Park KW. Articles: Inhibition of Aldose Reductase from Rat Lenses by Methanol Extracts from Korean Folk Plants. Natural Product Sciences. 2010;16(4):285-290.
- Yin H, Sun Y. Vincamine-producing endophytic fungus isolated from Vinca minor. Phytomedicine: international journal of phytotherapy and phytopharmacology. 2011;18(8-9):802-805.
- Cui Y, Yi D, Bai X, Sun B, Zhao Y, Zhang Y. Ginkgolide B produced endophytic fungus (Fusarium oxysporum) isolated from Ginkgo biloba. Fitoterapia. 2012;83(5):913-920.
- Rateb ME, Ebel R. Secondary metabolites of fungi from marine habitats. Natural Product Reports. 2011;28(2):290-344.
- Zhang Q, Wei X, Wang J. Phillyrin produced by Colletotrichum gloeosporioides, an endophytic fungus isolated from Forsythia suspensa. Fitoterapia. 2012;83(8):1500-1505.
- Thalapaneni NR, Chidambaram KA, Ellappan T, Sabapathi ML, Mandal SC. Inhibition of carbohydrate digestive enzymes by Talinum portulacifolium (Forssk) leaf extract. Journal of Complementary and Integrative Medicine. 2008;5(1):1-10
- Kumar AS, V.Venkatarathanamma, K.Suneeta, Kumari BS. Comparative In vitro screening of α-Amylase and α-Glucosidase enzyme Inhibitory studies in leaves of Annona species. Journal of Pharmacy Research. 2011;4(12):4431-4434.
- Tadera K, Minami Y, Takamatsu K, Matsuoka T. Inhibition of alpha-glucosidase and alpha-amylase by flavonoids. J Nutr Sci Vitaminol (Tokyo). 2006;52(2):149-153.
- Suryanarayana P, Kumar PA, Saraswat M, Petrash JM, Reddy GB. Inhibition of aldose reductase by tannoid principles of Emblica officinalis: implications for the prevention of sugar cataract. Mol Vis. 2004;10(2):148-154.
- Hayman S, Kinoshita JH. Isolation and properties of lens aldose reductase. Journal of Biological Chemistry. 1965;240(2):877-882.
- Muthenna P, Suryanarayana P, Gunda SK, Petrash JM, Reddy GB. Inhibition of aldose reductase by dietary antioxidant curcumin: mechanism of inhibition, specificity and significance. FEBS Lett. 2009;583(22):3637-3642.
- Lu Y, Chen C, Chen H, Zhang J, Chen W. Isolation and Identification of Endophytic Fungi from Actinidia macrosperma and Investigation of Their Bioactivities. Evid Based Complement Alternat Med. 2012;2012:382742.
- Lu YC, C.Chen, H.Zhang, J.Chen, W. Isolation and Identification of Endophytic Fungi from Actinidia macrosperma and Investigation of Their Bioactivities. Evid Based Complement Alternat Med. 2012;2012:382742.
- Gupta S. Prospects and perspectives of natural plants products in medicine. Pharmacology. 1994;26(1):1-12
- Matsuur H, Asakawa C, Kurimoto M, Mizutani J. Alpha-glucosidase inhibitor from the seeds of balsam pear (Momordica charantia) and the fruit bodies of Grifola frondosa. Biosci Biotechnol Biochem. 2002;66(7):1576-1578.
- Chandra A, Mahdi AA, Ahmad S, Singh RK. Indian herbs result in hypoglycemic responses in streptozotocin-induced diabetic rats. Nutrition research. 2007;27(3):161-168.
- Reyes B, Bautista N, Tanquilut N, Anunciado R, Leung A, Sanchez G, Magtoto R, Castronuevo P, Tsukamura H, Maeda K. Anti-diabetic potentials of Momordica charantia and Andrographis paniculata and their effects on estrous cyclicity of alloxan-induced diabetic rats. Journal of ethnopharmacology. 2006;105(1-2):196.
- Palanuvej C, Hokputsa S, Tunsaringkarn T, Ruangrungsi N. In vitro glucose entrapment and alpha-glucosidase inhibition of mucilaginous substances from selected Thai medicinal plants. Sci Pharm. 2009;77:837-849.
- Ali L AKA, Hassan Z, et al. 1995; 61:358-360. Characterization of the hypoglycaemic effects of Trigonella foenum graecum seed. . Planta Med 1995;61:358-360.
- Li-jian X, Li-gang Z, Jiang-lin Z, Wei-bo J. Recent Studies on the Antimicrobial Compounds Produced by Plant Endophytic Fungi. Natural Product Research & Development. 2008;20(4).
- Ramdanis R, Soemiati A, Abdul M. Isolation and α-Glucosidase inhibitory activity of endophytic fungi from mahogany (Swietenia macrophylla King) seeds. International Journal of Medicinal and Aromatic Plants. 2012;2(3):447-452.
- Fatmawati S, Kondo R, Shimizu K. Structure-activity relationships of lanostane-type triterpenoids from Ganoderma lingzhi as alpha-glucosidase inhibitors. Bioorg Med Chem Lett. 2013;23(21):5900-5903.
- Edward J. Dompeipen YS, Wahyudi Priyono Suharso, Herry Cahyana and Partomuan Simanjuntak. Potential Endophytic Microbes Selection for Antidiabetic Bioactive Compounds Production. Asian Journal of Biochemistry, . 2011;6:465-471.
- Saraswat M, Muthenna P, Suryanarayana P, Petrash JM, Reddy GB. Dietary sources of aldose reductase inhibitors: prospects for alleviating diabetic complications. Asia Pacific journal of clinical nutrition. 2008;17(4):558.
- Ruby Erach Jalgaonwala BVM, Raghunath Totaram Mahajan. A review: Natural products from plant associated endophytic fungi J. Microbiol. Biotech. Res. 2011;1(2):21-32.
How to cite this article:
PavithraN, Sathish L., Babu N, VenkatarathanammaV, Pushpalatha H, Reddy GBand Ananda K: Evaluation of α-Amylase, α--Glucosidase And Aldose Reductase Inhibitors in Ethyl Acetate Extracts of Endophytic Fungi Isolated from Anti-Diabetic Medicinal Plants .Int J Pharm Sci Res2014; 5(12): 5334-41.doi: 10.13040/IJPSR.0975-8232.5 (12).5334-41.
All © 2014 are reserved by International Journal of Pharmaceutical Sciences and Research. This Journal licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 3.0 Unported License.
N. Pavithra , L. Sathish , Nagasai, Babu , V. Venkatarathanamma , H. Pushpalatha , G. Bhanuprakash Reddy and K. Ananda *
Assistant Professor Poornaprajna Institute of Scientific Research, Bidalur Post, Devanahalli Bangalore-562110, India
07 May, 2014
14 July, 2014
18 August, 2014
01 December 2014