Impact of physical factors on hydrolytic enzyme production of Alternaria species was studied. Continuous light favored enzyme production in Alternaria species. Maximum enzyme activity occurred in 15 &ndash; 20 days in all the Alternaria species. Alternaria species produced hydrolytic enzymes maximum at 20 &ndash; 30oC temperature.All the species of Alternaria at 5.5 to 6.5 pH value produced maximum enzyme, however pH 3.5 and 8.5 inhibited enzyme activity

Chavan, Ashok M

Rathod, Sulochana R

Current Botany

English

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Current Botany 2012, 3(4): 15-17 ISSN: 2220-4822 Available Online: http://currentbotany.org/    Effect of physical factors on hydrolytic enzyme action of seed bone Alternaria Species  Sulochana R. Rathod1 and Ashok M. Chavan2  1Department of Botany, Government Art’s and Science College, Aurangabad (M.S.), India 2Department of Botany, Dr. Babasaheb Ambedkar Marathwada University, Aurangabad-431004 (M.S.), India.   Abstract  Impact of physical factors on hydrolytic enzyme production of Alternaria species was studied. Continuous light favored enzyme production in Alternaria species. Maximum enzyme activity occurred in 15 – 20 days in all the Alternaria species. Alternaria species produced hydrolytic enzymes maximum at 20 – 30oC temperature.All the species of Alternaria at 5.5 to 6.5 pH value produced maximum enzyme, however pH 3.5 and 8.5 inhibited enzyme activity.  Keywords: Light, pH, enzyme, fungi, seed  INTRODUCTION       The seed borne fungi are known to deteriorate the seeds and its contents [1]. Degradation of these seeds chemical content has been due to production of hydrolytic enzyme like lipase, protease, cellulose etc. and production of hydrolytic enzyme related with physical factors. However the less information is available about the impact of physical factors on hydrolytic enzyme produced by Alternaria. Considering the fact attempt were made to study the impact of physical factors on amylase, protease and lipase produced by Alternaria.   MATERIALS AND METHODS Production of hydrolytic enzyme       Production of hydrolytic enzyme was studied by growing the fungi in liquid medium containing  Starch (in case of amylase)/ gelatin (In case of protease)/ oil (In case of lipase), 1%, KNO3, 0.25% KH2PO4 0.1% and MgSo4. 7H2O 0.05%, pH of the medium was adjusted at 5.5. Twenty five ml of medium was poured in 100 ml conical flask autoclaved and inoculated separately with 01ml spore suspension of the fungi which were grown for 7 days on POA slants. The flasks were incubated for 6 days at 25 ± 10c with diurnal periodicity of light. On 7th day of the flasks were harvested filter no.1 the filtrates were collected in presterilized bottle and teemed as crude enzyme preparation.    Assay of hydrolytic enzyme  Assay of hydrolytic enzyme was done by cup-plate method.  RESULTS AND DISCUSSION        Impact of light, incubation period, temperature and pH on amylase, lipase and protease of Alternaria species were studied and results are summarized in table 1, 2, 3 and 4.       Amylase and lipase production was more in continuous light in all the species of Alternaria. Whereas continuous dark light inhibited amylase and lipase production. Protease production of A.crassa and A. Alternata inhibited by dark light and continuous light (Table 1).      Alternaria alternata, A. dianthicola and A. tenuissima do not show lipase activity on 5th days of incubation period. A. crassa and A. dianthicola do not produce protease at 5th day incubation however as the incubation period increase upto 20th day protease production increased. Effect of temperature on the enzyme activity of Alternaria produced enzyme. At 100c temperature A. citri, A. crassa, A. dianthicola and A. macrospora do not show any amylase production. However 400c temperature for A. citri and A. crassa also inhibited amylase production (Table 2).       Lipase production of A.citri and A. crassa initiated at 100c whereas A. citri, A. crassa, and A. macrospora inhibited lipase production at 400c similarly A. citri initiated its protease production at 600c whereas A. dianthicola and A. macrospora inhibited protease production at higher temperature (Table 3).      At pH 3.5 none of the species of Alternaria produced hydrolytic enzyme Lipase production of A. citri and A. dianthicola was also inhibited at pH 4.0 similarly A. crassa and A. macrospora inhibited protease enzyme production at pH 4.0 however the maximum hydrolytic enzyme production was reported at pH 5.5 to 6.5 in all species of Alternaria (Table 4).       Growth of the microorganisms is directly related to their metabolic activity, therefore, physical factor which are related to enzyme production were also studied in detailed. It is observed from the results that Alternaria species produced maximum amylase, lipase and protease in between 15 to 20 days at 20 to 300c temperature. It is interesting to note that in case of Alternaria species continuous light favored the enzyme production whereas 5.5 to 6.5 pH favored maximum enzyme production, however temperature up to 5 to 100c and pH below 4.5 inhibited the enzyme activity of Alternaria species similar types of work have been reported by   Received: July 18, 2012; Revised: Aug 22, 2012; Accepted: Sept 25, 2012.  *Corresponding Author Sulochana R. Rathod  Department of Botany, Government Art’s and Science College, Aurangabad (M.S.), India.  Email: srrathod@gmail.com    Sulochana R. Rathod   16earlier workers Adam (1981) [2] reported optimum period for amylase production is more 7 days in Humicola lanunginora. Similarly Charya and Reddy (1982) [3] reported optimum period for protease production is more than week. Tarrat et al (1973) [4] reported majority of microorganisms showed their protease activity between 25 to 350c temperature. Sonwane (2002) [5] observed A. alternata favored 20 to 300c for protease production under Alternata light and dark condition similar type of observation in seed mould of Jowar for amylase production have been reported by Panchal (1984) [6].       The range of pH for enzyme production in fungi was found to be variable. Bhosale (1989) [7] reported pH 5.0 to 6.0 is ideal for amylase production in Aspergillus flarus. Jonson (1968) [8] reported that alkaline pH was favored in protease production of A. tenuisima.  Table 1. Effect of light on enzyme production in Alternaria species  Species of Alternaria Illumination of light Continuous light Continuous dark Alternata dark and light Amylase Production A.alternata 29 18 19 A.citri 28 18 20 A.crassa 30 20 20 A.dianthicola 32 08 24 A.macrospora 33 20 20 A.tenuissima 27 20 26 Lipase Production A.alternata 30 25 27 A.citri 27 26 30 A.crassa 20 21 25 A.dianthicola 29 18 26 A.macrospora 25 17 20 A.tenuissima 28 15 20 Protease Production A.alternata 21 26 25 A.citri 23 23 20 A.crassa 16 20 12 A.dianthicola 18 15 20 A.macrospora 15 15 13 A.tenuissima 21 15 20                   Activity zone in mm  Table 2. Effect of incubation period on enzyme species in Alternaria species  Species of Alternaria Incubation period 5 days 10 days 15 days 20 days 25 days Amylase Production A. alternata 08 11 14 18 18 A. citri 11 18 13 14 14 A. crassa 05 14 15 14 10 A. dianthicola 12 15 15 16 15 A. macrospora 07 14 17 15 17 A. tenuissima 10 17 14 14 15 Lipase Production A. alternata -- 12 30 14 18 A. citri 06 06 25 15 20 A. crassa 08 10 26 18 21 A. dianthicola -- 10 18 20 19 A. macrospora 05 11 25 23 20 A .tenuissima -- 08 30 19 19 Protease Production A. alternata 07 18 22 21 20 A. citri 08 16 20 19 20 A. crassa -- 12 17 20 21 A. dianthicola -- 15 16 19 20 A. macrospora 07 12 18 20 17 A. tenuissima 10 17 21 18 18                     Activity zone in mm  Table 3. Effect of temperature on enzyme production in Alternaria species  Species of Alternaria Temperature 0oC 5 10 15 20 30 40 Amylase Production A.alternata -- 13 12 12 18 17 A.citri -- -- 08 10 20 -- A.crassa -- -- 09 10 15 -- A.dianthicola -- -- 10 12 17 21 A.macrospora -- -- 10 11 14 18 A.tenuissima -- 12 13 09 20 14 Current Botany 2012, 3(4): 15-17  17Lipase Production A.alternata -- 12 17 16 20 13 A.citri -- -- 15 19 21 -- A.crassa -- -- 10 15 18 -- A.dianthicola -- 10 20 20 18 12 A.macrospora -- 14 16 18 19 -- A.tenuissima -- 15 15 14 17 15 Protease Production A.alternata -- 14 17 20 20 12 A.citri -- -- 15 18 21 15 A.crassa -- 18 19 20 23 18 A.dianthicola -- 15 17 19 20 -- A.macrospora -- 10 12 20 18 -- A.tenuissima -- 13 18 15 17 15                  Activity zone in mm  Table 4. Effect of pH on enzyme production in Alternaria species  Species of Alternaria pH 3.5 4.5 5.5 6.5 7.5 8.5 Amylase Production A.alternata -- 15 19 20 21 -- A.citri -- 17 18 21 18 -- A.crassa -- 16 18 20 21 -- A.dianthicola -- 19 20 19 19 -- A.macrospora -- 15 16 18 20 -- A.tenuissima -- 13 15 20 19 -- Lipase Production A.alternata -- 20 25 30 30 -- A.citri -- 18 23 28 20 -- A.crassa -- 20 19 18 21 -- A.dianthicola -- 18 20 21 22 -- A.macrospora -- 19 23 27 20 -- A.tenuissima -- 13 18 26 19 -- Protease Production A.alternata -- 17 18 18 17 -- A.citri -- 16 16 20 20 -- A.crassa -- 16 18 17 16 -- A.dianthicola -- 18 16 18 21 -- A.macrospora -- 15 17 18 21 -- A.tenuissima -- 16 18 19 19 --                 Activity zone in mm  REFERENCES  [1] Charya, M.A.S. and S.M. Reddy.1981. Deterioration of Mung (V. radiata) Seeds due to certain seed-borne fungi. Indian J. Bot. 4(2): 187-90. [2] Adam, P.R. 1981. Amylase production by Mucor penicillus and Humicola Lanunginosa as related to myeelial growth. Mycopathologia 76 (2) : 97-102. [3] Charya, M.A.S. and Reddy, S.M. 1982. Toxic effect of Fusarium oxysporum on seed germination and growth of mung. Indian Bot. Rept. 1(2): 169-170. [4] Tarrant, P.J.V. Nadine Jenkins, Pearson, A.M.  and Duston, T.R. 1973. Proteolytic enzyme preparation from pseudomonas Ap. Mic. 25 (6): 996-1005.  [5] Sonwane, V.V. 2002. Studies on seed pathology of pea. Ph.D. Thesis, Dr. B.A.M.U. Aurangabad (M.S.). [6] Panchal V.H. 1984. Studies on seed-borne fungi of sorghum. Ph.D. Thesis, Marathwada University (M.S.). [7] Bhosale, A.M. 1989. Studies on fungal amylase and toxins in grain deterioration of sorghum, Ph.D. Thesis, Marathwada University, Aurangabad (M.S). [8] Jonsson, G.A.1968. Protease production by species of Entomopthora Appl. Microbial. 16:450-451.  

Effect of physical factors on hydrolytic enzyme action of seed bone Alternaria Species

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Effect of physical factors on hydrolytic enzyme action of seed bone Alternaria Species

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