This document describes a study that characterized the physicochemical properties of indole acetic acid oxidase (IAA oxidase) from Alternaria cepulae. The researchers found that the optimum pH for IAA oxidase activity was 5.5, and the optimum temperature was 40°C. Gel chromatography determined the molecular weight of IAA oxidase to be 30,000 daltons. The study provides information on the purification and characterization of IAA oxidase from A. cepulae which is involved in leaf blight disease of onions.
Kinetic study of free and immobilized protease from Aspergillus sp.IOSR Journals
In the present investigation partially purified alkaline protease from Aspergillus sp. As#6 and As#7 strains were entrapped in calcium alginate beads and characterized using casein as a substrate. Temperature and pH maxima of protease from As#6 strain showed no changes before and after immobilization and remained stable at 450C and pH 9, respectively. However km value was slightly shifted from 4.5mg/ml to 5 mg/ml. Proteases from As#7 strain showed shifting in pH optima to a more alkaline range (10.0) as compared with free enzyme (9.0). Optimum temperature for protease from As#7 strain showed changes after immobilization and shifted from 650C to 850C. However there was no significant effect on Km value but Vmax of immobilized protease from As#7 strain was also shifted from 200U/ml to 370U/ml. Immobilized protease from As#6 strain was reused for 3 cycles with 22% loss in its activity whereas immobilize protease from As#7 strain was reused for 3 cycles with 17% loss in its activity. Protease from As#7 strain has a higher affinity for the substrate and higher proteolysis activity than protease from As#6 strain. The present work concludes that Aspergillus As#7 strain may be a good source of industrial protease
Impact of anthelmintic efficacy of Calotropis procera on tegumental enzymes o...iosrphr_editor
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
Kinetic study of free and immobilized protease from Aspergillus sp.IOSR Journals
In the present investigation partially purified alkaline protease from Aspergillus sp. As#6 and As#7 strains were entrapped in calcium alginate beads and characterized using casein as a substrate. Temperature and pH maxima of protease from As#6 strain showed no changes before and after immobilization and remained stable at 450C and pH 9, respectively. However km value was slightly shifted from 4.5mg/ml to 5 mg/ml. Proteases from As#7 strain showed shifting in pH optima to a more alkaline range (10.0) as compared with free enzyme (9.0). Optimum temperature for protease from As#7 strain showed changes after immobilization and shifted from 650C to 850C. However there was no significant effect on Km value but Vmax of immobilized protease from As#7 strain was also shifted from 200U/ml to 370U/ml. Immobilized protease from As#6 strain was reused for 3 cycles with 22% loss in its activity whereas immobilize protease from As#7 strain was reused for 3 cycles with 17% loss in its activity. Protease from As#7 strain has a higher affinity for the substrate and higher proteolysis activity than protease from As#6 strain. The present work concludes that Aspergillus As#7 strain may be a good source of industrial protease
Impact of anthelmintic efficacy of Calotropis procera on tegumental enzymes o...iosrphr_editor
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
International Journal of Pharmaceutical Science Invention (IJPSI) is an international journal intended for professionals and researchers in all fields of Pahrmaceutical Science. IJPSI publishes research articles and reviews within the whole field Pharmacy and Pharmaceutical Science, new teaching methods, assessment, validation and the impact of new technologies and it will continue to provide information on the latest trends and developments in this ever-expanding subject. The publications of papers are selected through double peer reviewed to ensure originality, relevance, and readability. The articles published in our journal can be accessed online.
Optimum Conditions for Alginaseby Bacilllus Circulans R Isolateiosrphr_editor
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
Bioconversion of Penicillin to CephalosporinIOSR Journals
Cephalosporins are known as 3rd generation broad spectrum Beta lactam antibiotics, which can also be produced synthetically. Commonly, chemical ring expansion followed by an enzymatic removal of the phenylacetyl side chain is commonly employed to convert penicillin G into 7-aminodeacetoxycephalosporanic acid, the precursor for the manufacture of semisynthetic cephalosporins. This process requires several steps, is expensive and highly polluting. Thus there is a need to device a simple biological route to replace the chemical process. A mutant of Streptomyces clavuligerus NP1 was reported to converts Penicillin G to Deacetoxycephalosporin G (DAOG;phenylacetyl-7-aminodeacetoxycephalosporanic acid) enzymatically[5,8] . This enzyme, deacetoxycephalosporin synthase has the potential for the large scale transformation of Penicillin G to deacetoxycephalosporin. The present work studies the conditions required for efficient transformation of Penicillin G to Deacetoxycephalosporin using the wild type strain Streptomyces clavuligerus . Detection of cephalosporin was carried out using various methods. Additionally succinic acid formation was also studied as it could be used as a commercially important by product of the transformation. Deacetoxycephalosporin synthase also extracted and partially purified and characterised.
Optimization of process parameters for l asparaginase production by aspergill...eSAT Journals
Abstract L-asparaginase (L-asparagine amido hydrolase, E.C.3.5.1.1) is an extra cellular enzyme that has received considerable attention since it is used as an anticancer agent. L-asparaginase belongs to an amidase group that hydrolyses the amide bond in L-asparagine to aspartic acid and ammonia. The clinical action of this enzyme as an anti-carcinogenic is attributed to the reduction of L-asparagine; tumour cells unable to synthesise this amino acid are selectively killed by L-asparagine deprivation. L-Asparaginase has its application in food industry also. It helps in reducing the content of acrylamide in baked food products by hydrolysing the L-asparagine. L-Asparaginase is majorly produced by microorganisms including bacteria, yeast and fungi. The potential of Aspergillus terreus MTCC 1782 using cauliflower stalk: corn ears (3.75: 1.25) as substrate under SSF is the purpose of the study. Solid state fermentation (SSF) is a very effective technique opposed to submerged fermentation in various aspects. Various fermentation parameters such as types of agro material, their ratios, carbon source, nitrogen source, inoculum level, moisture content, temperature, pH, fermentation time, metal salts, and L-asparagine concentration, which influence the rate of enzyme production under SSF, were optimized. The optimized production of L-asparaginase has been obtained at 35°C for 4 days with a pH of 9.0, along with 50% moisture content, and 20% inoculum volume as the optimized fermentation conditions. The optimization was done using a ‘one-factor-at-a-time’ approach. The highest yield was obtained with, sucrose (1%w/v), ammonium sulphate (1%w/v), NaCl (1%w/v), L-asparagine (1%w/w), added to the fermentation medium, as supplements. Use of cauliflower stalk along with corn ear as potential raw materials for enzyme production could be of great commercial significance. Keywords: L-asparaginase, chemotherapeutic agent, Aspergillus terreus, SSF, mixed substrate, optimization
Effect of estradiol -17 β on arachidonic acid metabolism in sheep uterus: in ...iosrjce
The effect of estradiol-17 β on Arachidonic acid (AA) metabolism in non-pregnant sheep uterus was
studied under in vitro conditions. On incubation of uterine slices with estradiol-17β, the levels of prostaglandins
were altered but not Lipoxygenase (LOX) products. Based on their analysis on conventional TLC technique, the
Cyclooxygenase (COX) products PGF2α, 6-keto PGF1α and PGE2 were shown to be altered over an incubation
period of 0 to 120 minutes. The LOX products, HPETEs and HETEs did not show any change upon incubation
with estradiol-17β. This study gives a preliminary understanding of role of estradiol on AA metabolism.
Isolation and partial characterization of a new strain of Klebsiella pneumoni...GJESM Publication
Glycerol is a promising feedstock for microbial cultivation and production of 1,3 propanediol (1,3 PDO). Here we report a newly isolated bacterial strain BA11 from soil, capable of fermenting glycerol to 1,3 PDO, and
has been identified to be a strain of Klebsiella pneumoniae. Strain BA11 was fast growing showing peak 1,3 PDO production in 6 h of cultivation with productivity of 1.2 g/L-h without the addition of Vitamin B12. Based on the
optimum glycerol utilization (75%) and 1,3 PDO production (8.3 g/L) and yield (0.56 mol/mol glycerol utilized), the
most appropriate glycerol concentration for cultivation was 20 g/L. The strain BA11 could tolerate the pH range of 6
to 8.5 as no inhibitory effects were seen on growth as well as 1,3 PDO production. Strain BA11 was most active and
could produce high 1,3 PDO in the incubation temperature range of 25 to 40 oC. The production of 1,3 PDO was
maximum (9.3 g/L) under aerobic condition with 95.8% glycerol utilization. Addition of glucose to the glycerol fermentation led to increased cell mass but no improvement in the 1,3 PDO production.
International Journal of Pharmaceutical Science Invention (IJPSI) is an international journal intended for professionals and researchers in all fields of Pahrmaceutical Science. IJPSI publishes research articles and reviews within the whole field Pharmacy and Pharmaceutical Science, new teaching methods, assessment, validation and the impact of new technologies and it will continue to provide information on the latest trends and developments in this ever-expanding subject. The publications of papers are selected through double peer reviewed to ensure originality, relevance, and readability. The articles published in our journal can be accessed online.
Optimum Conditions for Alginaseby Bacilllus Circulans R Isolateiosrphr_editor
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
Bioconversion of Penicillin to CephalosporinIOSR Journals
Cephalosporins are known as 3rd generation broad spectrum Beta lactam antibiotics, which can also be produced synthetically. Commonly, chemical ring expansion followed by an enzymatic removal of the phenylacetyl side chain is commonly employed to convert penicillin G into 7-aminodeacetoxycephalosporanic acid, the precursor for the manufacture of semisynthetic cephalosporins. This process requires several steps, is expensive and highly polluting. Thus there is a need to device a simple biological route to replace the chemical process. A mutant of Streptomyces clavuligerus NP1 was reported to converts Penicillin G to Deacetoxycephalosporin G (DAOG;phenylacetyl-7-aminodeacetoxycephalosporanic acid) enzymatically[5,8] . This enzyme, deacetoxycephalosporin synthase has the potential for the large scale transformation of Penicillin G to deacetoxycephalosporin. The present work studies the conditions required for efficient transformation of Penicillin G to Deacetoxycephalosporin using the wild type strain Streptomyces clavuligerus . Detection of cephalosporin was carried out using various methods. Additionally succinic acid formation was also studied as it could be used as a commercially important by product of the transformation. Deacetoxycephalosporin synthase also extracted and partially purified and characterised.
Optimization of process parameters for l asparaginase production by aspergill...eSAT Journals
Abstract L-asparaginase (L-asparagine amido hydrolase, E.C.3.5.1.1) is an extra cellular enzyme that has received considerable attention since it is used as an anticancer agent. L-asparaginase belongs to an amidase group that hydrolyses the amide bond in L-asparagine to aspartic acid and ammonia. The clinical action of this enzyme as an anti-carcinogenic is attributed to the reduction of L-asparagine; tumour cells unable to synthesise this amino acid are selectively killed by L-asparagine deprivation. L-Asparaginase has its application in food industry also. It helps in reducing the content of acrylamide in baked food products by hydrolysing the L-asparagine. L-Asparaginase is majorly produced by microorganisms including bacteria, yeast and fungi. The potential of Aspergillus terreus MTCC 1782 using cauliflower stalk: corn ears (3.75: 1.25) as substrate under SSF is the purpose of the study. Solid state fermentation (SSF) is a very effective technique opposed to submerged fermentation in various aspects. Various fermentation parameters such as types of agro material, their ratios, carbon source, nitrogen source, inoculum level, moisture content, temperature, pH, fermentation time, metal salts, and L-asparagine concentration, which influence the rate of enzyme production under SSF, were optimized. The optimized production of L-asparaginase has been obtained at 35°C for 4 days with a pH of 9.0, along with 50% moisture content, and 20% inoculum volume as the optimized fermentation conditions. The optimization was done using a ‘one-factor-at-a-time’ approach. The highest yield was obtained with, sucrose (1%w/v), ammonium sulphate (1%w/v), NaCl (1%w/v), L-asparagine (1%w/w), added to the fermentation medium, as supplements. Use of cauliflower stalk along with corn ear as potential raw materials for enzyme production could be of great commercial significance. Keywords: L-asparaginase, chemotherapeutic agent, Aspergillus terreus, SSF, mixed substrate, optimization
Effect of estradiol -17 β on arachidonic acid metabolism in sheep uterus: in ...iosrjce
The effect of estradiol-17 β on Arachidonic acid (AA) metabolism in non-pregnant sheep uterus was
studied under in vitro conditions. On incubation of uterine slices with estradiol-17β, the levels of prostaglandins
were altered but not Lipoxygenase (LOX) products. Based on their analysis on conventional TLC technique, the
Cyclooxygenase (COX) products PGF2α, 6-keto PGF1α and PGE2 were shown to be altered over an incubation
period of 0 to 120 minutes. The LOX products, HPETEs and HETEs did not show any change upon incubation
with estradiol-17β. This study gives a preliminary understanding of role of estradiol on AA metabolism.
Isolation and partial characterization of a new strain of Klebsiella pneumoni...GJESM Publication
Glycerol is a promising feedstock for microbial cultivation and production of 1,3 propanediol (1,3 PDO). Here we report a newly isolated bacterial strain BA11 from soil, capable of fermenting glycerol to 1,3 PDO, and
has been identified to be a strain of Klebsiella pneumoniae. Strain BA11 was fast growing showing peak 1,3 PDO production in 6 h of cultivation with productivity of 1.2 g/L-h without the addition of Vitamin B12. Based on the
optimum glycerol utilization (75%) and 1,3 PDO production (8.3 g/L) and yield (0.56 mol/mol glycerol utilized), the
most appropriate glycerol concentration for cultivation was 20 g/L. The strain BA11 could tolerate the pH range of 6
to 8.5 as no inhibitory effects were seen on growth as well as 1,3 PDO production. Strain BA11 was most active and
could produce high 1,3 PDO in the incubation temperature range of 25 to 40 oC. The production of 1,3 PDO was
maximum (9.3 g/L) under aerobic condition with 95.8% glycerol utilization. Addition of glucose to the glycerol fermentation led to increased cell mass but no improvement in the 1,3 PDO production.
Slides from my information security management talk at a CalCPA Society Meeting. This story-filled non-technical talk provides real-world guidance executives and their boards need to meet the challenge of cybercrime.
- Why Care: Business Implications of Cyber Crime.
- The Critical Four: Key Questions for Managing Information Risk.
- Why Are We So Vulnerable? Three Inconvenient Truths.
- We Have a Firewall and Antivirus. Isn’t This Enough?
- What Are We Supposed to Do: Information Security Management Objectives.
- How Do We Do It: The Six Key Information Security Management Strategies.
- Leadership and Culture: The Final Frontier.
Abstract— Roots of Panax notoginseng were fermented with 30 fungi respectively. Almost one-third of the products showed increasing antibacterial activity. All products could inhibit GST-CDC25 phosphatase as a potential antitumor agent. HPLC profiles proved that components of unfermented P. notoginseng and fermented P. notoginseng have obviously changes.
IJRET : International Journal of Research in Engineering and Technology is an international peer reviewed, online journal published by eSAT Publishing House for the enhancement of research in various disciplines of Engineering and Technology. The aim and scope of the journal is to provide an academic medium and an important reference for the advancement and dissemination of research results that support high-level learning, teaching and research in the fields of Engineering and Technology. We bring together Scientists, Academician, Field Engineers, Scholars and Students of related fields of Engineering and Technology.
Synthesis, Characterization and Biological Evaluation of Oxazolone Derivativesijceronline
A series of six 4-aryl Benzelidene-2-phenyl-5- oxazolone derivatives were synthesized by condensation of aromatic aldehydes with N-benzoyl glycine (Hippuric acid) in the presence of sodium acetate and acetic anhydride at room temperature in ethanol. Six of the compounds are new derivatives. The structures of the compounds were evaluated based on 1H-NMR , IR and FTIR methods and by elemental analysis. .All the derivative compounds prepared were tested for their antimicrobial activity by disk diffusion technique. Test organisms: Bacteria like Staphylococcus aureusMTCC 7443 and Salmonella typhimuriumMTCC 733 Fungi like C.albicans and A.flavus The results were compared with those of the standard 0.5% Ciprofloxacin. The derivatives with Salicylaldehyde and cinnamaldehyde were showed excellent activities against E. coli. and Staphylococcus aureusMTCC 7443 : than Salmonella typhimuriumMTCC 733 bacteria. It also showed reasonable activity withFungi like C.albicans than A.flavus
IJPCBS 2012, 2(1), 110-116 Kavya et al. ISSN: 2249-9504
110
INTERNATIONAL JOURNAL OF PHARMACEUTICAL, CHEMICAL AND BIOLOGICAL SCIENCES
Available online at www.ijpcbs.com
ISOLATION AND SCREENING OF STREPTOMYCES SP. FROM
CORINGA MANGROVE SOILS FOR ENZYME PRODUCTION AND
ANTIMICROBIAL ACTIVITY
M. Kavya Deepthi1*, M. Solomon Sudhakar1 and M. Nagalakshmi Devamma2 1Department of Biotechnology, Rajalakshmi Engineering College, Thandalam, 2Department of Botany, Sri Venkateswara University, Tirupati, Andhra Pr Taadmesihln, aInddui,a I.n dia.
Anticancer Activity of L-asparaginase Produced from Amycolatopsis japonicaAI Publications
The ability of L-asparaginase to inhibit the formation of cancer cells has aroused scientists' curiosity in biological realms. In cancer cells, L-asparaginase suppresses protein synthesis by hydrolyzing L-asparagine to L-aspartic acid and ammonia. As a result, it's a crucial therapeutic enzyme in the treatment of Acute Lymphoblastic Leukemia in combination with other drugs (ALL). This enzyme has recently been discovered to be useful in a number of scientific fields, including clinical research, pharmacology, and the food business. Purification, characterization, and assessment of the cytotoxic effect of Amycolatopsis japonica L-asparaginase were the goals of this study. Amycolatopsis japonica was isolated from the plant rhizosphere and L-asparaginase was recovered. With a molecular weight of 37.5 KDa, partially purified L-asparaginase from A. japonica had a total activity of 1968.98 U with 26.696 mg total protein and a specific activity of 73.75 U/mg, 6.42 purification fold, and 42.86 percent recovery yield. In the presence of EDTA, Mg2+, pH8, 45oC, and 0.13 mM L-asparagine, L-asparaginase from A. japonica demonstrated good activity and stability, with Km and Vmax values of 0.13 mM L-asparagine and 0.43U/ mL, respectively. The cytotoxicity of L- asparaginase from A.japonica against a colon cancer cell line was high; with an IC50 value of 36 L. Amycolatopsis japonica could be a source of L-asparaginase, which could be a new target for cancer cells.
Isolation and purification of peroxidase from soyabeanPooja Walke
Peroxidase (EC. 1.11.1.7), an oxidoreductase, has iron porphyrin ring generally and catalyzes a redox reaction between H202 as an electron acceptor and many kinds of substrates by means of oxygen liberation from HzOz (Brill, 1996).
Optimization of Cultural Parameters for Cellulase Enzyme Production from Fung...IOSR Journals
Cellulalytic fungi synthesize cellulose enzyme for biodegradation of cellulose. This depends on various condition which include the source f isolation. This study was designed to determine the optimum condition necessary for cellulose production by fungi. Cellulose activities at different temperatures, pH and nitrogen sources by Rhizopus oryzae Aspergillus niger; A. flams, P. expansum and A. oryzae in liquid medium was studied and cellulose enzyme assay carried out by dinitrosalicylic acid method. All the fungal isolates have their highest cellulose activity at 400c except Penicillium expansum whose highest value of 1.28mg/ml was obtained at 320c. Cellulase produced 6m was found to be highest in all the isolate at pH 4.0 exception P expansum which occur at pH 5.5 (1.21mg/ml). The highest value e1.45mg/ml was obtained in A niger. Highest cellulose activity for A. niger, A. oryzae & P. expansum occurred in peptone. The study shows the need to determine the best physiological condition that allow for the optimal cellulose activity of fungal isolate. This will enhance their enzyme production.
Evaluation of Anti-oxidant Activity of Elytraria acaulis Aerial ExtractsIJERA Editor
Elytraria acaulis, a stem less perennial herb of Acantheceae family has many medicinal and therapeutic properties. Anti oxidative activity of the aerial parts of this Elytraria acaulis were assessed in the present study. The aerial parts of the plant (Stem & Leaves) were extracted in different organic solvents such as n-Hexane, Ethanol, Methanol, Ethyl Acetate and Chloroform. Initially, Total Phenolic & Total Flavonoids content in different solvent plant extracts were estimated. The free radical scavenging and antioxidant activity of the Elytraria acaulis aerial extracts in different organic solvents were also assayed by DPPH assay, FRAP assay. The aerial extracts of Elytraria acaulis have shown significant anti oxidant activity. Hence, further studies on this plant will enable elucidation of its therapeutic properties and medicinal applications
Similar to 17.Physicochemical characterization of Indole acetic acid oxidase from Alternaria cepulae (20)
44.Antimicrobial activity in leaf extract of Neem(Azadirachta indica Linn.)
17.Physicochemical characterization of Indole acetic acid oxidase from Alternaria cepulae
1. J. Ecotoxicol. Environ. Moni.l l(2) 147-148 (2001)
O Pilani Paramount Publications-Printed in India
ISSN: 0971 -0965-l I -01- 1 47
PHYSICOCHEMICAL CHARACTERISTIZATION OF
INDOLE ACETIC ACID OXIDASE TRQM- A-LTERNAKIA
CERULAE
B ANNADURAtr*, M SHANMUGAM** AND D B MOTLAG*
*DEPARTMENT OF BIOCIIEMISTRY AND MOLECULARBIOLOGY
UNIVERSITY OF MADRAS, CIIENNAI 600 025 INDIA
**DEPARTMENT OF APPLIED ZOOLOGY, KWEMPU UNIVERSITY
JNANA SAHYADRI, SHIMOGA DISTRICT, KARNATAKA, INDIA
ABSTRACT
The physicochemical properties like change of pH, temperature of the purified IAA oxidase from
Alternaria cepulae were investigated. Molecular weight was estimated by Gel chromatography of Andrews
method. The optimum pH of the enzyme was found to be 5.5. The optimum temperature of IAA-oxidase
activity was at 40oC. The molecular weight was determined as 30,000 daltons.
Key words: Alter:naria cepulae, pll, IAA oxidase, Molecular weight kit.
INTRODUCTION
IAA oxidase (EC1.11.17) was reported to be produced by various pathogens.
Themould Omphaliaflavida w'asreportedlAAoxidasewhileinfectingonCoffeaarabica
and Coleus blumei (Sequeira & Steeves 1954). Similarly IAA oxidase is reported to be
produced in the following cases ofpathogenesis viz Albugo candida while infecting on
Brassica napus (Srivastava et al 1962). Verticillium dahliae on Capsicum annum and
Fusarium oxysporum onMangifera indica (Kumar et al1980). Marasmius perniciosus
whiie infecting on Theobroma cacao (Krupasagar & Sequeira 1969); Protomyces
macrosporus onCoriandyum sativum (Tayal et al l98l),Verticillium dahliae while
infecting on Lycopersicum esculentum Reuveni & Ferrira 1985 and Plasmodiophora
brassica whole infecting on grapes @oemer & Ajarrg 197 4). The auxin content is reduced
in the host tissue due to the aetion of IAA oxidase.
IAA oxidase with acid pH optima has been isolated and purified from several plants
( Psenakova & Kolek 1973;Miyata et al 1981). This enzyme was isolated and purified
from downy mildew fungus Sc/e rospbra graminicola infection on pearl millet (Arora et al
1986), from Erisiphe graminis infection on barley (Fric 1975), from Pseudomonas
solanacearum infection on tobacco (Evident e et al 1986), IAA oxidase is also isolated
from virus infected plants (Gaborj anyi et al 197 3).
2. 148 BANNADURAIETAL
The characterization of IAA oxidase were. reported in many cases (Denckeva &
Klisurska 1986; Talwar et al 1985;Panel & Greppin l97Z;Mazzaet al1968;Mennes
1974; Laurema 1974). The characterization ofIAA oxidase can be altered by naturally
occurring compounds such as Phenols, Coumarins, Manganese salts and plant acids. There
were reportsthatmonophenols act as cofactorswhileparadihydricphmols andpolyphenols
act as inhibitors forIAA oxidase (Pilet 1964; 1966; Runkovaet al1972). Manganese ions
have been found to be a cofactor for IAA oxidase systems (Sequeira & Mineo 1966),
Scoioletin, a naturally occurring coumarin has been shown to inhibit IAA oxidase at higher
concenfrations, here as lower concenffations ofscopoletinwere stimulatory fimbert & Wilson
1970). The physicochemical properties ofthe pudfied IAA oxidase have been invesfigated
and presented.
MATERIALSAND METHODS
Enzymepreparation: CrudeenzymewasobtainedfromthemediumsuggestedbyRay(1956)andthepurification
was carried out according to the procedure of Annadurai ( 1 987; 1 998; 1 999; 2000).
Estimation of IAA oxidase activity: IAA oxidase activity was estimated according to the method suggested by
SequeriaandMineo(1966). TheactivityoflAAoxidasewasdeterminedbytherateofdisappearenceoflAA. The
reaction mixture contained 0.25 ml of 1 mM DCP, 0.75 ml of mixture of I mM IAA and 0.5 ml MnClr. HrO, 3.5 ml
of 0.02 M WV cihate buffer pH 5.5 and 0.5 ml protein. The mixture was shaken and incubation for I hour at 32
+ l"C Aftertheincubationwasover, I mlof Salkowskireagentwasaddedto5ml ofincubationmixture. Thepink
colour developed was read at 530 nm in a shimadzu spectrophotometer. The control treatments were carneci out in
an identical manner except that the enzyme was added after the addition of Salkowski reagent.
IAA oxidase enzyme unit was defined as the one micromoles of IAA destroyed in one Hour{lmber &
Wilson 1972).
Determination of Molecular weight
Gel filtration: The molecular weight of IAA oxidase was determined by gel filtration method according to
Andrews (1964). Sephadex G-100 was allowed to swell in 0.05 M phosphate buffer pH 7.2 for three days. The
solution was decanted and the swollen gel was further washed in 0.05 M Phosphate buffer (pH 7.2). The fine
particles were decanted and packed in a column (2 x 60 cm). The column was equilibrated with 0.05 M TrisI{ Cl
buffer containing 0.,I M NaCl at pH7.2. The column was run at a flow rate of ml,/hr and 5 ml fractions were
gollected. The elution volume (Vo) for each protein was determined by measuring the absorbance at 280 nm. The
void volume (Vo) was determined using Blue Dextran 2000. The standard proteins used were BSA (MW 68,000),
Ovalbumin (MW 45,000), Chymotrypsinogen (MW 325,000) and cytochrome C (MW 12;000). The Kav value
was calculated using the following relationship:
Ve-Vo
Kav = --------------
Vt- Vo
Where Vt is the total bed voiume. The Kav values of standard proteins were plotted in semilogarithmic graph paper
( on the scale) against the corresponding molecular weight (on the logarithmic scale). The straight line obtained was
used for the determination of molecular weight of purified enzymes.
Journal of Ecotoxicology & Environmental Monitoring. Vol.11(2001)
3. Ef{'cct of change of pH on IAA oxidase activit_v: Effect of changc of pH on IAA oxidasc activity was carried out
over the pH of 3.0 to 7.0. IAA oxidase (0.5 nrl) rvas allowed to react with 0.25 ml of I mM DCP. 0.75 ntl of a t.trtxturc
of I nrM IAA and MnCl.. H,O and 3.5 ml of 0.02 M of buffer fi-onr pH 3.0 to 7.0 at 32 + l"C for I hottr -['he
activity of thc cnzynre was estimatcd at the end of the incubation pertod.
Efl'ect of change of temperature on IAA oxidase activitv: 0.25 ml of mM DCP, 0.75 rnl of nrixture olnrM lAA.
'0.5 mM 4nCl.. H.O and 5.5 ml of 0.02 M citrate bu{fler at pH 5.5 was incubatcd with 0.5 ml of IAA oxidase irl
different tempel'arurcs. After the incubatron peilod. IAA oxidase activity was esttnratcd.
RESULTS AND DISCUSSION
Fig. 1 shorvs the determination of nT olecular weight of IAA oxidase in a sephadex
G- 1 00 column using illolecular rveight reference proteins. The molecular weight of I AA
oxidase is found to be 30.000 daltons from the senrilogarithmic graph. Fig.2 shorvs the
effbct of pH on IAA oxidase activity. From the results, it can be seen tlmt the eff-ect enzyme
has an optilnum pH of 5.5 for IAA as a substrate. From tl-re results presertted in Fi-s.3.
enzyme activity- remperature relatronship is seen. It is evident from the activitv that at pH
5.5. the optinllnll temperature fbr enzynte activity is 40"C.
K."
Fig. i Deternrrrration of rrrolccuiar rvc'ighl of IAA oxrclasc b1, gel filtration
IAAO
Activity in
units 100
5.5
Frg.l Elfcci ol'changc oipl-l on i:A orrclasc acr r1'
Jcurnal of Ecotoxicology & Environmental Monitonng Vo!.11(2001)
o.4
0.3
o.2
o.l
150
50
Molecuiar Weight
pH
4. t50 B A}.INADURAIETAL
IAAO
activity
I"AAO
Actlvlty ln
units
Temperature
oC
Fig.3 Effcct ofchange oftentperaturc on tAA oxidase activity of A cepulae'
The rnolecular weight of IAA oxidase fiom G-100 sephadex colufirn (Fig' 1) rvith
the help of low moleculaJweight kit, the MW of IAA oxidase is found to be 30'000
daitons. trAA oxidase was not stable at higher temperature optimlrm temperature for IAA
oxidase was 5 .5. The enzyme is repofiecl to be active between 5 '0 to 7 '0 pH (Krupasagar
& Seciueira 1969)"
ACKNOWLEDGEN1ENT
The author B A is grateful to Dr. s c Dhar and Dr R Puvanakrishnan' Scientists at the Department ol
Biotechnologv. Cl,,RI. cnenn"ai for Laboratory facilities and useful SLlggcstloll and UcC, Neu, Delhi for researcl,t
grani.
REFERENCES
Andrcu,s p 1964 Estrnrarron of molccular wcights ofprotcrns by Sephaclex gelfiltratrorr' Biochem' J'91. 221-713 '
;nnaciurar B. palani B. Mahalingam S and Singarav.i, Cl S9s-ifftlt of afla:toxin on RBC' WBC and haemoglobin
of' lialltr'r' ruittts ndr;eglcas Biojournat t 0: 1 65- I 72'
Annadur.ai B, pr.abhakarar.r V, N4d-.. Farrrk S a.a nrr-rttt,imaiun p f CSa Production of anrylase in Aspergillus or;'zae'
or"".r",.l,ll',|,,|il]i]l ll",'Jl;l,t;]-,, o r.o Singaravcruc leee prodLrction orar'raroxi, in conrcnrirrated storccl
I gru,,'t.. J. Ecotoricol. Environ' [tonit'9(l): 13-17'
Anrraciurai B. 1(arr-rnanidhi P and l4alralilrgan-) S t999 6"t'i(}tn',*t' of Alternttria t'epuloe in leatblight clisease of
ontotl..l" Ecobioi' I l(4): 191)-i05 i.- ,^..+r^ri.-L+...-...^,rrnn'
Annaciurar ts and Motlag D ri r99C Extlace llular cnzylrles of Alternaritt celtulue in lcaiblight diseasc of onton
Biojournal 8: 105-lt)9 .r , -r.L^ ^^r..,-r
.nnlicluratB.Coplnatho,nap,ion,RlggSStr-rc.Iicsontherolcofthccelllvalldegr.adrrlgCnzynlcsrnlcafblight
discaseofonron lillltittttccpr(l,tnn.).urr.dtyll'nrnariacepultte'
Biojournall0:l7-l-"1 iE'
.nnadurar B. ,{rul Kurraran P, Kalpana erul t<umai'Li"'JeUJ'f S't'Lan K 1999 ll'lrcrobiologrcal anal-vsrs of
cllorc drrrtlttng rvatt't'sLrppil' Bio' J' I l: l6i- I 7l'
rrnurirrrar B and i4otiag D B 1999 croivth ot ltrci'io''i" ip'lou in leafulight discasc rn onion Bio' J' I I : i6l-
i (i5
Journal of Ecotoxicology & Environmental Monitoring Vol'11(2001)
5. j
!
PI,{YSICOCHi],{IC'AL CI IAI{AC1'ERIS'IIZA'|ION OT INDOt,E A('L]1'IC A(-ID OXIDAST 1.-.I
,rrnariirrrfi L] rnd Motlag D Il 2000 [:lfcit of Various carbon solrrccs on producttot.t ol cndopolVgairctLlrollasc (]l
.iltt'rriortu ( ('pul(tL'. J' Ecotoicoi. Environ. llonit. l0 (l): 37-41
1n1r(.ir Lliti B lnd Nlotleg f) B i999 Elfcct oi tthytohortrones in norntal and int-cctcd onttln 'icr t!s l'r 1/It lit,ti r,i
t't'prrluc'.. llio. ,|. I I: 155- 16().
AnnlclLrrai et.lStlStr,li.roncndopolygalacturonaseinlc-iliblrghrciiscascofoniott(lllitrrtt.t'lr(1 Lilllr.lcaLtsct'l bt
..iltei;ntrria t.cprrluc /Ponnappa) and its irrtcraction rvtth phytohormones. Ph.D. ilresis. litrlverstty ol'
4 ldrtis.
Atora Y K. N4ehta N. ThakLrr l) P rnd Wagie D S 1986 Enz_vntc changcs associrtcd $ rth ltost PiilusltL- tntci itcttons
bctrvcen Pcarl ntillct an( Dorvny nrilderv furrgus. lSclL'ros1tortt !rtrttirtrt:rtitt- J. Phytopathol. I l6: 9r-
t ()5
Bonircr-ll arrd AianS M R 1974 Dcr-r]onstrattot.r cl indolc acctic acrd or.rdrsc rnhjbitors rn l' usnirtriittltitt)rLt hru.r.rt(1it
rniectcd roots of rapc- Z. Ptlaneuzenlir. Pllanzensclttz' 8l I i2-l i ;-
I)cnuher11 A and I(lrsuriia il iCSt, f'ompatisotr of son-ic i(lnctlc par itlllctcrs olpcroidase and IAA oxtdasc rtl thc
.or,,.s.j l,t g,-o.,",th and diticrcnttatton plaltt cc1ls. Biologi. Plautarutn 28: i()5
EviilenrcA.SurcroL,. lacoball:sNSirnciRandassoClgti6/--N-Acctvl indoic--l-acctvl-r'-L-i.vsrne.3 lllcf;tboiltc
of Iridoic-iacettcacrdil'onr !'sc'ttricttrtouttss.'rut.qu('psttyttstQtlot. Plrltochenlistr-r ?5: Il-'
Frrc F 1975 Activity oi-peroridlsc ancl IAr oidasc il1thc trssucs of r-trild$v inlcctcd bericr il.r.r.ilplte ,!rrttttirtrs i'
sp horder marchacl ). Ph1'tnpath Z 80 6'i -1>
Gabor.ianyi R. Scgi F and Balazs n tgl5 Cro",fh inhibition of virus Infcctcd plilnts. .'ltcrrtrr)ri (rl l)cr'r(lxi.iirsc
enzylt-tcs r1 colrpatrblc- anci rncontpatiblc )rost parasrtc r-elations. ictll I)htol)atltol acatl Sci' Ilrrng li
.[iI-90
lrrbcrr M p and Wrlson I- A 1972 IAA oriclasc preparatiotls li'onr srvct-t potal0 tr:bcrs. [)ltr tocltclttistr-r' I I: ](t-
16
l(rLrpasagarV anci Scclucrra L 19(r9 Auxrn destructionby,,larasnttu. Perfit(t():;Ll' Am. J. Ilot.56. -l.)()-it)^
Kuntar-"1 Bcnrrral SpSanclIlantS I()E0Dcpictronofauxlnsrnntangosccdlingsallccteclw'lilrl)Llllcil)'it)psiaL'rl:
nrango ittali'ortllatrorls. Intl. J. Exptl. Biol' l8: 28(r-2E8
L.aurenra S t97+ inclolc acettc acldc oxrdase rn rcsting ccreal gratns. Phvsiol Plant. -30 .-10l--11)6'
[-orvry O H. Ilosebrough N .i. Fan-A L O and Ranclal] R .] 196 I Protcrn nrcasut-cnlcnt 'ith tlrc l;olin pircnoi rcauctrt.
J. Bio. Chenr. 193: 165-175
,ianncs A N4 l97rt Titc Indolc acctic acid orrclase of l,upitru,s luleus: A quuntitatiYc cot]lllarison of thc actr l1 tll-
thrs enzymc in root nocitilcs and roots Acta. bot' Nerat 22: 694-7(15'
l4rzzaG.C'har.resC, I-loucheslr,l.llrcarcl JadnRynanci U lg6tiEndo..ecnousaurinsinlrcaltlri'rrrcldiscascdl:)ants
Biochem. Biophys. Acta. 167: 89-98.
[,c.el (- and (iraporn ]l 197i Control of glgrvth by', aurins and its spccilic nclrtrill inhibitor'. Plant Cell l'}lrvsiol' l5
i5l-i 5(r
fri]ct F E 1!)(t4 Ar-rxrn conlent and auxrn catabolrsir of thc sterns ol Etrpirctrbtu (1'r(1/.ll7-t-/--rnicctcd b;v i'r0rrttt'r:r
pr.sl. Fh1'tochemistry: 61 ,--{t21 -
pscnilovr -I'and
l(oieli j 1975 Enzynratrc degradation of Indolyl-3-acctlc acid u'ithtn irtatzc (Zcrt arrt.tr.s) Root'
Biochem. Phvsiol. Pllanz. 163 : 5l(r-544.
Rat p i4 i95(t Thc destruction of Indolc acetic acid ll spcctrophotonrctrtc sttrdy olthc crlzr'trrlrttc tcitclton Arch'
Biochem. Biophls. 64. l()5-21(t.
ReL^ clr R and Fcrrtra J R I 9E5
'fhc relationship bctu,ccn pcroxidasc activity and thc tcsislatlcc of totratocs
(1.1,t:1rO"rr,rry esculetttttnt) to Varricilltun dttltliaa. Phltopath' Z' 118 193-l()i '
Runkova l- V, Lis E K. Tontaszanski M and Antonszeu,ski R 1972 Ftrnctroll of Phcnolic substanccs rn thc
,Jcgradation sysrenl ollndolc-.1-acetic acid rn Strarvbcrrics. Biol. Plant. l4: 7l-8 i.
ScqucrraLandMineoLlg66Partial punticatronandmincticsoflndoleaccticrlcrdorrclasclt'otrltobitccorools
Plarrt Pltl'siol 46: 195-,l9(i.
Sri11,astavaBISandSha11,tv{ l9(rlThcbiosynthcsisollndoleaceticacidinMcluntp.:pot-rrllrrl(l)crs).Can.'1.[lot
J0: 509_5 I j
'Trlurtr C. Dcndsay J I, S and CLrpla V K 1985 Kinctic propcrtics of lAA. oxidasc fi'onr nrungbcan cotl'lcdorrs
Phl tochemistrY 21. (:75-676.
1'ayrl i1 S. Sharrnr S Nl and Agaru,al iv,l L l98l Studies on thc polyphcnols, irt'ott-ins. chlorophvlls. IAA ortdasc
ancl aptvlascofn,rrntil ancl falscsnltinfcctcdIcavcsttf'l'haorti.tstlt'es/i't.s ltttl. l'}h1'topathol. l4 l-il-
ile
Journal of Ecotoxicology & Environmental Monitoring' Vol 11(2001)