Agro-Industrial Wastes as Feedstock for Enzyme Production: Apply and Exploit the Emerging and Valuable Use Options of Waste Biomass

Agro-Industrial Wastes as Feedstock for Enzyme Production

Apply and Exploit the Emerging and Valuable Use Options of Waste Biomass

Editors

Gurpreet Singh Dhillon

University of Alberta, Edmonton, AB, Canada

Surinder Kaur

University of Lethbridge, Lethbridge, AB, Canada

Table of Contents

Cover image

Title page

Copyright

List of Contributors

Chapter 1. Microbial Enzyme Factories: Current Trends in Production Processes and Commercial Aspects

Introduction

Microorganisms as a Potential Source for Hydrolases

Agro-industrial Residues for Hydrolases Production

Recovery and Purification of Hydrolases

Enzyme Applications and Global Market

Some Commercial Hydrolyses and Their Manufacture

Conclusions

Chapter 2. Fruit and Vegetable Processing Waste: Renewable Feed Stocks for Enzyme Production

Introduction

Characterization and Composition of Fruit and Vegetable Processing Waste

Important Enzymes for Industrial Applications

Enzyme Production Through Fermentation

Potential of Fruits and Vegetable Solid Waste for the Production of Enzymes

Conclusions

List of Abbreviations

Chapter 3. Bioprocesses for Enzyme Production Using Agro-Industrial Wastes: Technical Challenges and Commercialization Potential

Introduction

Major Agro-Industrial Residues/Wastes

Enzymes: The Biological Tools for Industrial Applications

Bioprocesses for Enzyme Production Using Agro-Industrial Wastes

Solid-State Fermentation (SSF)

Commercialization Potential

Effect of Fermentation Parameters

Carbon and Nitrogen Source

Aeration

pH

Temperature

Inoculum Age and Size

Substrate Pretreatment

Technical Challenges and New Trends in Enzyme Production Using Agro-Residues

Heat Dispersal

Scale-Up

Biomass Determination

Conclusions

Chapter 4. Industrial Enzymes: Recovery and Purification Challenges

Major Industrial Enzymes

Enzyme Purification: A Trial and Error Strategy

Purification of Enzymes Largely Depends on Its Application

Challenges in Enzyme Purification

Monitoring the Progress of Enzyme Purification

Important Steps in Enzyme Purification

Determination of Enzyme Activity by Simple Methods

Fractionation of Enzymes With Ammonium Sulfate and Other Solvents

Enzyme Inactivation and Control

Chromatographic Steps

Ultrafiltration

Automation of Protein Purification

Conclusions and Future Perspectives

Abbreviations

Chapter 5. Low-Cost Enzymes and Their Applications in Bioenergy Sector

Lignocellulose Biomass and 2G Ethanol

Enzymes in Biomass Hydrolysis

Orange Bagasse as an Example of Agro-industrial Waste With High Reusable Value

Native Microorganisms

Final Remarks

Chapter 6. Role of Enzymes in Environment Cleanup/Remediation

Introduction

Pollution of the Environment

Methodologies to Cleanup Polluted Environments

Enzymes Important to Pollutant Removal

Enzymes as Indicators of Polluted/Restored Environments

Advantages and Drawbacks of Enzymes as Environmental Actors

Conclusions and Future Perspectives

List of Abbreviations

Chapter 7. Enzymes: Applications in Pulp and Paper Industry

Introduction

Application of Xylanases for the Delignification of Pulp

Mechanism of Delignification of Pulp with Xylanases

Factors Affecting Xylanase Treatment Efficiency

Commercial Availability of Xylanases

Application of Laccases for the Delignification of Pulp

Delignification of Pulp With Fungal Laccases

Delignification of Pulp With Bacterial Laccases

Factors Affecting the Laccase-based Biobleaching of Pulp

Synergistic Effects of Enzymes Involved in Biobleaching of Pulp

Role of Enzymes in Pitch Control

De-inking of Old Newsprint by Enzyme Treatment

Application of Laccase for Grafting of Pulp Fibers

Decolorization and Detoxification Effect of Laccase on Wastewaters From Pulp and Paper Industry

Conclusions and Future Perspectives

List of Abbreviations

Chapter 8. Enzymes in Food Processing

Introduction

Enzymatic Applications in Food Processing

Dairy Industry

Conclusions and Future Perspectives

List of Abbreviations

Chapter 9. Seafood Enzymes and Their Application in Food Processing

Introduction

Seafood Enzymes

Applications of Enzymes

Conclusions

List of Abbreviations

Chapter 10. Enzymes for Nutritional Enrichment of Agro-Residues as Livestock Feed

Introduction

Feed Substrates for Animal Nutrition

Forage, Silage, and Related Feed Substrates

Fruit Wastes

Enzymes Used in Animal Feed Supplementation

Use of Exogenous Enzymes in Animal Nutrition

Conclusions

Chapter 11. Potential Applications of Enzymes in Brewery and Winery

Introduction

Enzymes in Brewing

Enzymes in Winery

List of Abbreviations

Chapter 12. Recent Applications of Enzymes in Personal Care Products

Introduction

Application of Nanoparticles for Enzyme Immobilization

Future Perspectives

List of Abbreviations

Chapter 13. Strategies to Enhance Enzyme Activity for Industrial Processes in Managing Agro-Industrial Waste

Introduction

Enzyme Immobilization

Different Strategies for Enzyme Immobilization on Various Supports

Nanomaterials as Dynamic Supports for Enzyme Immobilization

Future Directions

Chapter 14. Biotechnological Production of Enzymes Using Agro-Industrial Wastes: Economic Considerations, Commercialization Potential, and Future Prospects

Introduction

Factors Deciding the Economics of Enzyme Production

Commercialization Potential of Enzymes

Enzyme Technologies With Commercial Potential

Future Perspectives

Index

Copyright

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List of Contributors

R.E. Abraham,     Deakin University, Waurn Ponds, VIC, Australia

A. Alberti,     State University of Ponta Grossa, Ponta Grossa, Brazil

A.T. Awan,     State University of Campinas, Campinas, Brazil

N. Capalash,     Panjab University, Chandigarh, India

F. Carvalho,     Universidade de Lisboa, Lisbon, Portugal

J.C. de Carvalho,     Federal University of Parana, Curitiba, Brazil

M. de la Luz Mora,     University of La Frontera, Temuco, Chile

L.P. de Souza Vandenberghe,     Federal University of Parana, Curitiba, Brazil

S.K. Deshmukh,     The Energy and Resources Institute, New Delhi, India

S. Devatkal

ICAR-Central Institute of Post-Harvest Engineering and Technology, Ludhiana, India

ICAR-National Research Centre for Meat, Hyderabad, India

G.S. Dhillon,     University of Alberta, Edmonton, AB, Canada

P. Fernandes

Universidade Lusófona de Humanidades e Tecnologias, Lisbon, Portugal

Universidade de Lisboa, Lisbon, Portugal

L. Gianfreda,     University of Naples Federico II, Portici, Italy

T.A. Gomes,     Federal University of Paraná, Curitiba, Brazil

N. Gopalan,     CSIR-National Institute for Interdisciplinary Science and Technology (NIIST), Trivandrum, India

G.S. Kaira,     CSIR-Central Food Technological Research Institute, Mysuru, India

M. Kapoor,     CSIR-Central Food Technological Research Institute, Mysuru, India

K. Kaur,     Panjab University, Chandigarh, India

U. Kumar,     The Energy and Resources Institute, New Delhi, India

N. Libardi,     Federal University of Parana, Curitiba, Brazil

A.U. Muzaddadi,     ICAR-Central Institute of Post-Harvest Engineering and Technology, Ludhiana, India

K.M. Nampoothiri,     CSIR-National Institute for Interdisciplinary Science and Technology (NIIST), Trivandrum, India

A. Nogueira,     State University of Ponta Grossa, Ponta Grossa, Brazil

H.S. Oberoi

Guru Nanak Dev University, Amritsar, India

ICAR-Central Institute of Post-Harvest Engineering and Technology, Ludhiana, India

ICAR-Indian Institute of Horticultural Research, Bengaluru, India

D. Panwar,     CSIR-Central Food Technological Research Institute, Mysuru, India

M. Puri,     Deakin University, Waurn Ponds, VIC, Australia

S. Puri,     Panjab University, Chandigarh, India

V.L. Queiroz,     State University of Campinas, Campinas, Brazil

S.R.F. Raj,     Nesamony Memorial Christian College, Kanyakumari, India

M.A. Rao,     University of Naples Federico II, Portici, Italy

C. Rodrigues,     Federal University of Parana, Curitiba, Brazil

R. Scelza,     University of Naples Federico II, Portici, Italy

P. Sharma,     Panjab University, Chandigarh, India

R. Sharma,     Guru Nanak Dev University, Amritsar, India

G. Singh,     Panjab University, Chandigarh, India

C.R. Soccol,     Federal University of Parana, Curitiba, Brazil

M.R. Spier,     Federal University of Paraná, Curitiba, Brazil

K. Sunar,     The Energy and Resources Institute, New Delhi, India

L. Tasic,     State University of Campinas, Campinas, Brazil

J.O. Ugwuanyi,     University of Nigeria, Nsukka, Nigeria

P. Vijayaraghavan,     Manonmaniam Sundaranar University, Kanyakumari, India

S.G.P. Vincent,     Manonmaniam Sundaranar University, Kanyakumari, India

Chapter 1

Microbial Enzyme Factories

Current Trends in Production Processes and Commercial Aspects

L.P. de Souza Vandenberghe, J.C. de Carvalho, N. Libardi, C. Rodrigues,  and C.R. Soccol     Federal University of Parana, Curitiba, Brazil

Abstract

Microorganisms are the main sources of the most demanded commercial enzymes. They are produced in large scale with the employment of different strains, which can be parental strains or molecularly modified. The capacity of a strain to synthesize higher concentrations of the protein of interest depends not only on the selected microorganism but on the condition of cultivation and media composition. Currently, agro-industrial by-products are viewed as potential economical feedstocks for microroganism cultivation for enzyme production serving dual purposes: to solve environmental problems and to add value to these substrates. Some aspects of enzyme synthesis are described in this chapter, including the main factors that affect the productivity. Some aspects of the enzyme’s process development, mainly hydrolases, such as cellulases, xylanases, phytases, mannanases, amylases, lipases, and others, from batch to industrial scale are reported. General downstream operations for the separation, recovery, and purification of these enzymes are also presented. Finally, the most important hydrolases produced and commercialized are also listed.

Keywords

Agro-industrial subproducts; Amylases; Cellulases; Lipases; Mannanases; Microbial hydrolases; Phytases; Xylanases

Introduction

Enzymes are special proteins, which catalyze chemical reactions with great specificity and rate enhancements. These reactions are the basis of the metabolism of all living organisms, and provide tremendous and economical biocatalyst conversions. The first half of the past century saw rapid development in enzyme chemistry. The Enzyme Commission, set up by the International Union of Biochemists (1965), has published a system of enzyme classification and more than 4000 enzymes have been recognized. However, 25,000 natural enzymes have now been speculated to exist, which means about 90% of the reservoir of biocatalysts still remains to be discovered and characterized (Menon and Rao, 1999).

When designing a new synthetic process, a suitable catalyst for the reaction must be found, and enzymes are ideal candidates for this role. The industrial use of enzymes has developed rapidly because of their specific functions (Ogawa and Shimizu, 1999). Commercial exploitation of microbial enzymes began much before their natures and properties were worked out. For centuries, extracts of plants have been used to bring about hydrolysis of polymeric materials. However, these sources of enzymes were unreliable and expensive too, hence search for alternative sources began. Largely, this was found in the microbial cultures (Menon and Rao, 1999). The first enzyme produced industrially was the fungal amylase takadiastase, employed as a pharmaceutical agent (for digestive disorders) in the United States in 1894. The 1950s saw important growth in the industrial enzyme production and use of microbial enzymes. By 1969, 80% of all laundry detergents contained enzymes, mainly proteases. Additional enzymes, such as lipases, amylases, pectinases, and oxidoreductases were then experimented with in the detergent industry.

Many enzymes are commercially available, and numerous industrial applications have been described. An overview was recently published by Li et al. (2012) about the most important applications of enzymes. The role of enzymes in many processes has been known for a long time. People in ancient Greece used enzymes from microorganisms in baking, brewing, alcohol production, cheese making, etc. With better knowledge and purification of enzymes, the number of applications has increased manyfold, and with the availability of thermostable enzymes a number of new possibilities for industrial processes have emerged (Haki and Rakshit, 2003). Thermostable enzymes, which have been isolated mainly from thermophilic organisms, have found a number of commercial applications because of their overall inherent stability (Demirijan et al., 2001).

The food, feed, agriculture, paper, leather, and textile industries are well suited for enzyme technology because products as well as raw materials consist of biomolecules, which can be produced, degraded, or modified by enzymatic processes (van Beilen and Li, 2002). The majority, almost 75%, of currently used industrial enzymes are hydrolytic in action, being used for the degradation of various natural substances. Approximately 200 microbial original types are used commercially. However, only about 20 enzymes are produced on a truly industrial scale. According to a recent research report from the Austrian Federal Environment Agency, about 158 enzymes were used in the food industry, 64 enzymes in technical application, and 57 enzymes in feedstuff, of which 24 enzymes are used in three industrial sectors (Li et al., 2012).

Enzymes significantly contribute to global annual revenue, and therefore, the emphasis has been on engineering them. Although a great deal of data is accumulating on making alterations in microbial enzymes, there is a lack of definite information on redesigning industrial enzymes. Modification of the existing enzymes has become a trend for fine tuning of biocatalysts in the biotech industry. Tolerance to high or low temperatures, exhibiting activity in alkaline or acidic environments, high performance in nonaqueous media, and increased protease resistance are a few of the requisite properties (Joshi and Satyanarayana, 2015). Proteins are then redesigned in such a way that the industrial processes can be carried out in a more economic and greener way. In protein engineering, various methods are employed in modifying the target proteins. These are mainly rational design, directed evolution, and semirational approaches for designing and constructing novel proteins. The rational methods require prior knowledge of the amino acid sequence, three-dimensional (3D) structure and knowledge of the structure–function aspects of the target proteins. When mutations carried out by computational and random mutagenesis are compared, often the best mutants have been developed by either of the methods.

The use of recombinant DNA technology has made possible the manufacture of novel enzymes. Such enzymes may be discovered by screening microorganisms sampled from diverse environments using modern methods of protein engineering or molecular evolution. As a result, several important new enzymes have been developed. Another important achievement is improvement of microbial production strains. For example, several microbial strains recently developed for enzyme production have been engineered to increase enzyme yield by deleting native genes encoding extracellular proteases. Moreover, certain fungal production strains have been modified to reduce or eliminate their potential for production of toxic secondary metabolites (Olempska-Beer et al., 2006).

In many cases, the substrates in industrial processes are artificial compounds or alternative substrates, such as agro-industrial subproducts. In this way, enzymes known to catalyze suitable reactions for such processes are still unknown. Therefore, screening for novel enzymes that are capable of catalyzing new reactions in such mediums is constantly needed.

Microorganisms as a Potential Source for Hydrolases

Over the past few decades, considerable research has been undertaken with the enzymes produced by a wide variety of microorganisms. Enzymes have been derived from several fungi, yeasts, bacteria, and actinomycetes.

One of the most efficient and successful means of finding new enzymes is to screen a large number of microorganisms. The findings of thermophilic organisms, for example, have been possible with the isolation of a large number of beneficial thermophilic microorganisms from different exotic ecological zones of the earth and the subsequent extraction of useful enzymes from them (Demirijan et al., 2001). One extremely valuable advantage of conducting biotechnological processes at elevated temperatures is reducing the risk of contamination by common mesophiles. Elevated process temperatures include higher reaction rates due to a decrease in viscosity and increase in diffusion coefficient of substrates and higher process yield due to increased solubility of substrates and products and favorable equilibrium displacement in endothermic reaction (Kumar and Swati, 2001). Table 1.1 provides some examples of hydrolases produced by different microorganisms.

Agro-industrial Residues for Hydrolases Production

The majority of the commercially available hydrolases are produced by submerged fermentation (SmF), overexpressing selected genes in either native or heterologous microbial hosts. However, there are still many research publications testing and comparing SmF with solid-state fermentation (SSF) processes, as an attempt of improvement of the enzymatic productivity, reducing its costs, and improving the viability for commercial application. According to Hussain et al. (2013), the SSF is the method of preference for α-amylase production, regarding the better quality of production, easy follow-through procedures, lesser production costs, energy savings, and no foam formation. However, xylanases (Ho, 2014) and phytases (Lei et al., 2013) are produced in a better way using the SmF process. Fungal lipases are preferably produced in SSF, while bacteria and yeast lipases are produced in SmF (Treichel et al., 2010).

The definition of a medium composition that can significantly affect the product concentration, yield, and volumetric productivity is of central importance when developing an industrial bioprocess. The use of agro-industrial residues is an interesting strategy for the reduction of the costs associated with the culture medium formulation. In recent years, there has been an increasing trend toward efficient utilization of agro-industrial residues in fermentative bioprocess, such as coffee pulp and husk, cassava husk, cassava bagasse, sugarcane bagasse, sugar beet pulp, apple pomace, declassified potatoes, citric pulp, sugarcane molasses and vinasse, soybean molasses and vinasse, etc (Soccol and Vandenberghe, 2003).

Table 1.1

Hydrolases Produced by Different Microorganisms

For α-amylase production, corn steep liquor and chicken feather were already reported as nitrogen sources as well as molasses, sugarcane bagasse, rice husks, corn starch, potato starch, maize starch, wheat starch, and rice starch as carbon sources (Hussain et al., 2013). Several cellulosic residues, such as sugarcane bagasse, wheat bran, paper pulp, corn cob residue, wheat straw, corn stover residue, and many others were reported as carbon sources and inducers of cellulases production in SmF or SSF processes (Singhania et al., 2010). Sahnoun et al. (2015) tested the substitution of yeast extract as the nitrogen source used with agro-industrial residues namely tuna fish powder, wheat gluten waste, and soybean meal, producing α-amylase from Aspergillus oryzae in SSF (1L-Erlenmeyer flask, 30°C). After optimization studies, the authors achieved maximum specific enzyme activity of 22,118.34  U  g−¹. In addition, they found a new high-molecular-weight α-amylase, namely Amy C. Singh and Gupta (2014) used Sal (Shorea robusta) deoiled cake as substrate for α-amylase production using Aspergillus flavus TF-8 in SSF, SmF and mSSF (modified solid-state fermentation), obtaining maximum enzymatic activities of 1.82, 0.36 and 2.51  U  mL−¹, respectively. After optimization of the SmF process, it was reached at 26.38  U  mL−¹.

The most important way to compare different production methods and culture medium compositions, including agro-industrial residues, is the enzyme productivity. Haq et al. (2003) reported the amylase productivity of 102  U  mL−¹  h−¹, for an SmF bioprocess, conducted at 40°C during 48  h, using Bacillus licheniformis as the producer strain with wheat bran and pearl-milled starch as substrate. Esterbauer et al. (1991) reported the cellulase production with a volumetric productivity of 100  FPU  L  h−¹, using wheat straw as substrate. The majority of the published data has been focusing on the maximum enzymatic activity, which makes the comparison between results very difficult.

The great majority of the hydrolases published data are related to bench-scale production processes. Only a few reports deal with the results and problems related to cellulase production in a pilot plant, or to significant parameters for the scaling up of the process from a few to several hundred liters. Oinonen et al. (2004), working in a 700-L fermenter using SmF process, achieved 1160  nkat  mL−¹ of cellulase after 72  h. In 1991, Esterbauer et al. (1991) already reported the cellulase production in a volume of 3000  L, with a yield of 250  mg  cellulase  mg−¹ substrate and maximum cellulase activity of 18  FPU  mL−¹, using a Trichoderma reesei mutant.

In addition, the sustainability of the process should also be taken into the account. Besides the use of agro-industrial residues, strategies such as open culture processes with nonsterile culture conditions could be explored for in situ enzyme production, with mixed cultures of microorganisms (Kleerebezem and Loosdrecht, 2007), taking advantage of ecological selection principles, when high purification degree is not required.

Table 1.2 presents the production of important hydrolases using agro-industrial residues/subproducts.

Table 1.2

Production of Hydrolases Using Different Agro-Industrial Residues

FPU  mL−¹  =  Filter paper unit per milliliter, ECU  mL−¹  =  nkatal per milliliter, U  g−¹  =  units per gram, U  gds−¹  =  units per gram of dry substrate, U  g−¹  DBB  =  units per gram of dry bacterial bran, Qp  =  Volumetric productivity.

Recovery and Purification of Hydrolases

The initial step in all protein purification protocols is the release of the enzyme from the cell or tissue material, or cell extract preparation (Dako et al., 2012). Protein purification varies from a simple one-step purification procedure to large-scale purification processes. The key to obtaining successful and efficient protein-purification strategies is the selection of appropriate techniques that maximize yield and purity and minimize the number of steps required for purification. Each step sequentially enhances the level of purification from a crude extract but also increases purification costs. It is essential to minimize protein losses throughout the process; therefore the use of fewer steps during purification is important since losses can occur in each step. This fact occurs due, for example, to linking to separation matrices, insolubilities, or losses into the fringe fractions during separation procedures (Sá-Pereira et al., 2003).

Preparation of crude samples (preparation, extraction, and clarification, especially for enzymes produced with alternative substrates) to be further purified is a critical step, since it can affect enzyme recovery. Sufficient information can be gathered by complementary techniques that give a profile of the target protein (eg, pI, molecular weight, hydrophobicity) to permit an effective choice of purification strategy.

A pretreatment of the crude enzymatic extract is usually required to remove colored compounds, using dialysis or polyethylenimine (Mishra et al., 1990). Some purification steps involve, for example, ammonium sulfate precipitation, gel filtration, and ion-exchange chromatography and other chromatographic techniques.

Isolation and purification of microbial enzymes has previously been reported. Purification procedure strategies are chosen for each enzyme, including the concentration and purification steps, and the corresponding recovery fields, purification factors, and final specific activities of the different enzymes.

Chromatographic methods are used in a unique or serial steps, when high-level purity enzymes are required, As reported by Sá-Pereira et al. (2003), microbial xylanases are mainly purified by chromatographic methods, using from two to five purification steps (including concentration), and providing recovery yields ranging from 0.2% to 78%. Lower yields are generally obtained when a greater number of purification steps are used. In some cases, higher purification factors correspond to a greater number of purification steps, although no overall correlation was observed. In general, high purification factors correlate with purification schemes for low-molecular-weight xylanases (ranging from 24 to 56  kDa). When two purification steps are used, the purification factors can range from 3 to 36.6, and with three purification-steps can range from 3 to 890.

Ammonium sulfate precipitation generally seems to be an effective method for precipitating high-molecular-weight enzymes. Some reports note that ammonium sulfate promotes flocculation rather than precipitation. In these cases, a decanting funnel can be recommended for visualizing the separation of enzyme phases. Each of the enzyme phases can be analyzed separately in order to balance the enzyme protein concentration and enzyme activity, and to check the purity level of the enzyme by electrophoresis or high-pressure liquid chromatography. Another problem associated with the utilization of ammonium sulfate precipitation is that salt interferes in determinations of activity, for example, leading to overestimations of such activity in crude extracts. Thus, in general a dialysis or desalting step is generally required before the determination of enzyme activity.

Other strategies, such as membrane technology for protein separation, with ultrafiltration, are easily scaled up. Ultrafiltration is based on the principle of separation through a semipermeable membrane filter. Many filter type units are available commercially, either applying a centrifugal or stirred cell type of separation (Bonner, 2007). The principle for all ultracentrifuge methods is the filtration through a membrane that has a selective molecular weight cutoff point, therefore it will separate the extract based on the size of particles, not the charge (Dako et al., 2012).

The cost of the membranes was a serious limitation in the past; however, this barrier is continuously being broken. The effectiveness of membrane separation depends on the enzyme, its molecular weight and conformation, and certainly the adapted porosity and material of the membrane. Ultrafiltration membranes can markedly reduce enzyme recovery yields (MW  >  20  kDa), owing to the ability of the enzyme to pass through ultrafiltration membranes with low-molecular-weight cutoff values (5–10  kDa). Another problem can be the material. Since most commercial ultrafiltration membranes are made of cellulose or its derivatives, for example, the presence of cellulases in a xylanase crude extract should be determined before ultrafiltration. Activity losses of about 50–70% after an ultrafiltration step were reported (Sá-Pereira et al. 2003).

Depending on the properties of the enzyme, certain modifications of purification methods must be considered regarding specific problems that can be encountered throughout the process, such as enzyme insolubility and loss of enzyme activity. Although several classic and more modern methods are available to solve these kinds of challenges, the enzyme purification step remains a major challenge for any method of extraction used (Dako et al., 2012).

Enzyme Applications and Global Market

Enzyme technology was presented as the application of free enzymes and whole cell biocatalysts in the production of goods and services. It is obviously an interdisciplinary field, as an important component of sustainable industrial development. Its applications range from straightforward industrial processes, the degradation of various natural substances in the starch processing, detergent and textile industries, to pharmaceutical discovery and the manipulation of DNA/RNA in biotechnology research.

Table 1.3 presents some examples of the multiplicity of enzyme applications in different industry sectors.

Table 1.3

Industrial Applications of Enzymes

Modified from Soccol, C.R., Vandenberghe, L.P.S., Woiciechowski, A.L., Babitha, S., 2006. Applications Industrial Enzymes. In: Pandey, A., Webb, C., Soccol, C.R., Larroche, C. (Org.), Enzyme Technology. vol. 1. New York, pp. 524–537.

Gross world sales for enzymes are estimated at $8.0  billion in 2015, with a predicted annual growth rate of 7%. Several case studies, recently published by Li et al. (2012), show how industry has implemented biotechnological processes and assessed benefits in terms of costs and sustainability.

The fast growth over the past decade has also been seen in a diversity of other industries from organic synthesis in pharmaceutical industry to diagnostics enzymes. Contrarily, the detergent industry, once the largest sector in the global enzyme market, experienced a decline due in part to the pricing pressures from the main detergent manufactures after the turn of the century. Demand for cleaning enzymes was accelerated by 2005 as the product lines were reformulated with more-effective new enzymes launched continuously. Bioenergy related enzyme demand was limited by the new legislative mandates for grain based ethanol. While the development of the second generation biofuels derived from cellulosic raw materials will be in favor of demand growth over a long time.

Some Commercial Hydrolyses and Their Manufacture

Since the beginning, the production of enzymes has been relatively concentrated in a few developed nations, including Denmark, Switzerland, Germany, Netherlands, and the United States. In Denmark, Novozymes and Danisco together dominate 70% of the total enzyme market. More recently, in 2010, DSM’s sales revenue accounted for 6%. About 100 companies produced enzymes in China, which is estimated less than 1% of the world market share. Some Japanese manufactures are also playing an increasing role in the world enzymes production.

International competition is intense in the enzyme market. Big companies aim to purchase other companies in order to become more efficient and competitive. However, there are still many small- to medium-sized enzyme producers. In China, there are manufacturers that have been gradually washed out from the market. On the other hand, several multinational companies have invested in the enzyme industry in China. Novozymes has three enzyme plants and DSM announced a joint venture with a Chinese company, Yixing Qiancheng Bio-Engineering Company Ltd., to provide α-amylase and xylanase to acquire the food and beverage enzyme markets (Li et al., 2012).

The enzyme market was established in 1832 by Nagase, in Osaka, Japan, producing enzymes for pharmaceuticals, foods, household products, agriculture, and textiles. The leading enzyme manufacturer is Novozymes, with 47% of market share and 902 deposited patents. As the world’s biggest enterprise in the production of enzymes, it produces enzymes for household products, foods and beverages, biopharmaceuticals, bioenergy, feed, and other industrial enzymes.

The diversity of commercial enzymes and their manufacturers can be seen in Table 1.4.

Table 1.4

Commercial Hydrolases and Their Manufacturers