I= STUDIES ON THE DEVELOPMENT OF BIOTECHNOLOGICAL PROCESS FOR BETA-AMYLASE BY MUTANT STRAIN OF BACILLUS POLYMYXA
Pakistan Research Repository Home
 

Title of Thesis
STUDIES ON THE DEVELOPMENT OF BIOTECHNOLOGICAL PROCESS FOR BETA-AMYLASE BY MUTANT STRAIN OF BACILLUS POLYMYXA

Author(s)
Rauf Ahmad
Institute/University/Department Details
University Of The Punjab
Session
1995
Subject
Zoology
Number of Pages
208
Keywords (Extracted from title, table of contents and abstract of thesis)
beta-amylase, bacillus palmyra, starch hydrolysis, mutational studies, fermentation techniques, 14 l stirred fomenter, thermophilie bacterial culture

Abstract
Thermophilie bacterial culture capable of producing extracellular beta-amylase was isolated and identified as Bacillus polymyxa PCSIR-90. The productivity of the enzyme was improved by using UV-irradiation and alkylating at 30oC with 6 percent bacterial inoculums. The production of bacterial enzyme was significantly enganed with liquefied starches of sweet potato and wheat. Among the nitrogen sources examined, ammonium dehydrogenates phosphate, yeast extract, or peptone was found to be suitable for the enzyme formation. The maximum enzymic activity was obtained with corn steep liquor, sunflower meal or corn meal as agricultural by-products. Enzyme synthesis was stimulated the addition of Ca2 + at the level of 5x10-5 M in the culture medium. The ideal cultural conditions in solid state fermentation were wheat bran as best substrate, acetate buffer as a diluents, fermentation time 48 hour at 30oC and finally 1.8cm depth of bran with 68 percent moisture level. Supplementation of wheat bran with soluble starch and dominium hydrogen phosphate slightly enhanced the enzymic yield. However, partial replacement of wheat bran with soluble starch and diammonium hydrogen phosphate slightly enhanced the enzymic yield. However, partial replacement of wheat bran with corn meal in ratio of 1 :1 (w/w), increased the synthesis of bacterial beta-amylase. The culture filtrate was also examined for the detection of maltose by the aid of paper chromatography in order to confirm the presence of beta-amylase. Scale-up studies were carried out in 2 L glass jar fomenters the production of Bacillus polymyxa beta-amylase. It was used along with amyloglucosidase in order to improve the efficiency of starch conversion to maltose. The enzyme was found to be most active and stable in pH range 6.5 to 7.0 and 5.5 to 9.0 respectively. Beta-amylase was stable at 60oC for 60 minutes at pH 6.2 in the presence of 5 percent starch. The activity of the enzyme was enhanced by the addition of 5 mM Ca2+.

Download Full Thesis
2592.8 KB
S. No. Chapter Title of the Chapters Page Size (KB)
1 0 Contents
369.39 KB
2 1 Introduction 1
241.35 KB
  1.1 Structure Of Starch 6
  1.2 Starch Hydrolyzing Enzymes 10
  1.3 Starch Hydrolysis 11
  1.4 Objectives 15
3 2 Review Of Literature 18
464.69 KB
  2.1 Uses And Utilization Of Bacterial Beta-Amylase 18
  2.2 Raw Starch Digestion By Bacterial Beta-Amylase 25
  2.3 Action Pattern Of Bacterial Beta-Amylase 27
  2.4 Isolation Of Bacillus Species 29
  2.5 Mutational Studies 31
  2.6 Fermentation Studies 33
  2.7 Purification And Characterization 40
  2.8 Gene Manipulation 45
  2.9 Immobilization Of The Enzyme 47
  2.10 Kinetic Studies 49
4 3 Materials And Methods 51
368.72 KB
  3.1 Materials 51
  3.2 Methods 53
  3.3 Fermentation Techniques 59
  3.4 Isolation And Purification 64
  3.5 Characteristics 68
  3.6 Kinetic Studies 70
  3.7 Analytical Methods 71
5 4 Results 78
1004.64 KB
  4.1 Isolation Of Microorganisms 78
  4.2 Screening Of Isolates 82
  4.3 Identification Of The Selected Strain 84
  4.4 Culture Improvement 88
  4.5 Submerged Fermentation In Shake Flask 89
  4.6 Production Of Beta-Amylase In Stirred Jar Fomenters 111
  4.7 Studies In 14 L Stirred Fomenter 121
  4.8 Solid State Fermentation 124
  4.9 Enzyme Purification 141
  4.10 Characteristics 143
  4.11 Determination Of Michaelis Menten Constant 153
  4.12 Starch Hydrolysis 153
  4.13 Substrate Specificity 159
6 5 Discussion 163
378.23 KB
  5.1 Summary 182