This document describes a study to optimize process parameters for maximum production of the enzyme tannase using the Plackett-Burman experimental design method. Key factors that influence tannase production by bacteria will be screened using this method. The document outlines the Plackett-Burman design, discusses parameters that impact tannase production and their typical ranges, and describes the uses of tannase and the work plan for the study. The goal is to identify significant factors that maximize tannase production for applications in food, medical and other industries.

1. By:
Sumit Pannu
Roll No. : 15001511009
M.Tech-3rd
Sem
Optimization of process parameters for maximum
production of Tannase using Plackett-Burman method
Department of Biomedical Engineering
DCRUST-Murthal
Under the Guidance of:
Mr. Saravjeet Singh
Asst. Prof., Deptt. of BME
D.C.R.U.S.T.
Sonipat-Haryana

2. Table of contentsof contents
• Introduction
• Plackett-Burman method
• Tannase production
• Uses of Tannase
• Work plan
• References

3. Introduction
• We get the significant factors for maximum production
of Tannase using Plackett-Burman method .
• Tannase has great commercial significance in the feed,
drink and medical industries.
• Tannase is an enzyme catalyzing the hydrolysis of the
tannic ( C76H52O46 ) , that is present in many different
parts of plants, to produce glucose and gallic acid
(C6H2(OH)3COOH) or Tri-Hydroxy-Benzoic Acid.

4. Plackett –Burman MethodPlackett –Burman Method
Run A B C D E F G H I J K
• 1 + + - + + + - - - + -
• 2 - + + - + + + - - - +
• 3 + - + + - + + + - - -
• 4 - + - + + - + + + - -
• 5 - - + - + + - + + + -
• 6 - - - + - + + - + + +
• 7 + - - - + - + + - + +
• 8 + + - - - + - + + - +
• 9 + + + - - - + - + + -
• 10 - + + + - - - + - + +
• 11 + - + + + - - - + - +
• 12 - - - - - - - - - - -
A 12 run PB design matrix.

5. Plackett –Burman MethodPlackett –Burman Method
• Developed in 1946 by statisticians Robin L. Plackett &
J.P. Burman.
• It is an efficient screening method to identify the
active factors using as few experimental runs as
possible.
• With an 8-Run Plackett-Burman design, we can look at
up to 7 factors; with a 12-Run design, up to 11 factors
and with a 16-Run design up to 15 factors.

6. When to Use Plackett-Burman Design ?
It is particularly helpful to use Plackett-Burman design:
•In screening of process parameters
•In two-level multi-factor experiments
•When there are more than four factors (if there are between
two to four variables, a full factorial can be performed)
•To economically detect large main effects
For N = 12, 20, 24, 28 and 36 (where N = the number of
experiments)

7. Analysis of Experiment Results
• Once the experiment is run and the samples measured, the
data from the experiment are used to calculate the effects
and determine the statistical significance of those effects
by two way ANOVA(analysis of variance) for different
factors and different levels.
• There is no way to know if the effects are due to changes
made in the factor levels or due to common cause
variation in the process unless their statistical significance
is tested.

8. Production of Tannase
• Submerged fermentation: It is the process of producing
enzyme(tannase) with the help of micro-organism.
Micro-organism(bacteria erwinia) breakdown the
nutrients and release desired amount of enzyme in the
solution.
• Micro-organism seeded in liquid(tannic acid ,other
sources of tannic acid,H2O,KCLand NaNO3 ) for some
time(20-24 hours for tannase) to produce tannase enzyme.
• Enzyme activity – moles of substrate converted per unit
time (unit per ml).

9. Process parameters for tannase productionProcess parameters for tannase production
• PH - 2-8
• Temperature - 30-35° c
• Incubation time - 20-24 hours
• Metal ions - k++
and Na++
• Nitrogen and carbon sources

10. Substrate for Tannase productionSubstrate for Tannase production
Substrate Tannase activity
(Unit/ml)
Pomegranate peel 79.5
Grape seed 40
Pine bark 64
Oak bark 46

11. Uses of TannaseUses of Tannase
• Bioconversion of tannic acid by enzyme tannase to
produce gallic acid(Tri-hydroxy benzoic acid).
• Gallic acid is used for preparation of :
- antibacterial drugs
- vitamin supplements
- fruit juice
- coffee flavoured soft drink
• Gallic acid has anti-cancer properties(prevent cellular
mutations) and anti-fungal properties.

12. Work planWork plan
Work Expected time
Experimental work 2-3 months
Statistical work 15-20 days
Other 10-15 days

13. ReferencesReferences
[1].Sahira N. Muslim,Alaa N. Mahammed,Hadeel K. Musafer,Israa
M. S. AL_Kadmy,and Shatha A. Shafiq, Sraa N. Muslim “Detection
of the Optimal Conditions for Tannase Productivity and Activity by
Erwinia Carotovora”, Journal of Medical and Bioengineering Vol. 4,
No. 3, June 2015.
[2]. Swaran Nandini1, Nandini KE2 and Krishna Sundari S2, “Food
and Agriculture Residue (FAR): A Potential Substrate for Tannase
and Gallic Acid Production using Competent Microbes” , Nandini,
et al., J Bioproces Biotech 2014.
[3]. QualityTrainingPortal6,2000-2016 Resource Engineering, Inc.
All rights reserved.
[4]. Manikandan jayakumar,When and How to Use Plackett-Burman
Experimental Design © Copyright iSixSigma 2000-2016.