Microbial Processing of Agricultural Wastes to produce Pectinase Enzyme(s) and Evaluation of their Industrial Applications This document summarizes a study on the production of pectinase enzymes from agricultural wastes using microbial processing. Pectinases are important industrial enzymes with applications in juice extraction and clarification. The study aims to optimize pectinase production from bacterial sources using agricultural waste, characterize the enzymes, and evaluate their applications. Key steps include screening bacteria, optimizing fermentation parameters, purifying and characterizing the enzymes, and testing applications in juice clarification and fruit peeling. A hierarchical model is also developed to analyze barriers to industrial pectinase production.

1. Microbial Processing of Agricultural Wastes to
produce Pectinase Enzyme(s)
and Evaluation of their Industrial Applications
By: Meesha Singh

2. Introduction
• Pectic substances (or Pectin) are structural
polysaccharides found in primary cell wall and middle
lamella of fruits and vegetables.
• Pectinases catalyze pectic substance degradation through
depolymerization and de-esterification reactions.
Types of Pectinases Catalysis reactions and products
Pectin esterases (PE) de-esterification of methoxyl group of pectin, forming pectic acid.
Polygalacturonases (PG) and
Polymethylgalacturonases (PMG)
hydrolytic cleavage of a-1,4-glycosidic linkage in pectic acid and
pectin, respectively.
Polygalacturonate Lyase (PGL) and
Polymethylgalacturonate Lyase (PMGL)
cleavage of a-1,4-glycosidic linkage in pectic acid and pectin,
respectively by trans-elimination reaction and forming
unsaturated galacturonates and methyl galacturonates,
respectively.

3. Pectin
Sources
Citrus
fruits
Apple
Papaya
Banana
Guava
Tomatoes
Sugar
beet
Carrots
Pectinolytic
organisms
Bacteria
YeastsMoulds

4. Why Pectinase(s)?
25 % of global food enzymes
produced and their market is
increasing day by day!
• Increasing demand →
Exploring Pectinolytic microbial
strains.
Pectinolytic microbes with
• high extracellular
pectinase activity,
• temperature and pH
stability for a longer
period of time,
• cost-effective production.

5. Juice
clarification
Peeling of
citrus fruits
Extraction of
juice &oil
Wine
making
Maceration
Liquefaction Gelation
Coffee and
tea
fermentation
Functional
foods,
Prebiotics
Waste water
treatment
Retting of
fibre crops
Wood
preservation
Industrial
Applications of
Pectinases

6. FARM
• Agricultural residues
• Soil samples
• Wastes from food
processing units
FERMENTER
• Microbial
processing
• Submerged State
Fermentation
FOOD
• Fruit juices
• Prebiotics
• Coffee and Tea
Waste recycle &
Value addition

7. Objectives
1. Production of pectinase from suitable bacterial sources
using agricultural wastes and market effluents as precursors.
2. Optimization of the physico-chemical parameters for
maximum enzyme production.
3. Purification and characterization of the pectinase(s)
produced.
4. Evaluation of industrial applications of pectinase(s)
produced.
5. Hierarchical framework formulation of various barriers
affecting the industrial production of pectinase(s).

8. Methodologies used
Critical Control Points to be
monitored:
• pH
• Temperature
• Incubation period
• Inoculum size
• Effect of agitation
• Substrate affinity and
concentration
• Moisture content
• Carbon sources
Microbial processing:
• Screening, Isolation and
Identification of suitable
Pectinolytic microbial
strain.
• Characterization of the
strain.
• Assay of enzyme activity.

9. Methodologies used…
Submerged Fermentation
• Fed-batch operation.
• 250ml Erlenmeyer flasks containing cultivation medium.
• Inoculation, Incubation
• Filtration
• Analyse the filtrates daily for pectinase production.

10. Methodologies used…
Kinetic Analysis:
• Determination of the
kinetic parameters
affecting the process.
Statistical Analysis
• Analysis of Variance
(ANOVA) according to
response surface
regression procedure using
Central Composite Design
model.

11. Methodologies used…
Purification of crude
enzyme(s)
1. Ammonium sulphate
fractionation.
2. Chromatographic
techniques.
3. SDS-PAGE
Enzyme immobilization
• Rice straw as support:
activated with glutaraldehyde
using poly(ethylene glycol) as
stabilizing agent.
• Other eco-friendly supports
like coconut fibers, rice straw,
wood cellulignin, chitin,
cotton cloth and olive
pomace.
• Onto UF membranes

12. Laboratory studies of various applications
Laboratory scale studies of the industrial applications of
bacterial pectinase(s) purified.
Mainly:
Juice clarification
Enzymatic peeling of fruits
Removal of mucilage from coffee beans
Waste water treatment

13. Hierarchical model formulation for sustainable
fruit juice supply chain
• Identify the critical barriers affecting industrial production
(enzymatic) of fruit juices by explorative study.
• Ranking of the critical barriers by AHP method.
• Model formulation by prioritizing the barriers.
The model proposed will be beneficial for the efficient
management and HACCP implementation in juice industries.

14. Analytic Hierarchy Process (AHP)
Widely used in business industries for
decision-making process and efficient
supply chain management .
Main steps:
• Divide the problem into various
levels/criteria, structured as a
hierarchy.
• Pair-wise comparison to determine
importance of each criteria.
• Calculate relative weight.
• Rank each criteria.
Hazard Analysis and Critical
Control Points (HACCP) :
a systematic approach to the
identification, evaluation,
and control of food safety
hazards.

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17. THANK YOU!