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ENZYMES
To allow enzyme-bearing products to be applied effectively
in an industrial setting, one must have a basic understanding of these
products' characteristics, advantages, and limitations. How Do Enzymes Function? Enzymes
are "biological catalysts." "Biological" means the
substance in question is produced or is derived from some living organism.
"Catalyst" denotes a substance that has the ability to increase the
rate of a chemical reaction, and is not changed or destroyed by the chemical
reaction that it accelerates. Generally speaking, catalysts are
specific in nature as to the type of reaction they can catalyze. Enzymes, as a
subclass of catalysts, are very specific in nature. Each enzyme can act to
catalyze only very select chemical reactions and only with very select
substances. An enzyme has been described as a "key" which can
"unlock" complex compounds. An enzyme, as the key, must have a
certain structure or multi-dimensional shape that matches a specific section of
the "substrate" (a substrate is the compound or substance which
undergoes the change). Once these two components come together, certain
chemical bonds within the substrate molecule change much as a lock is released,
and just like the key in this illustration, the enzyme is free to execute its
duty once again. Many chemical reactions do proceed
but at such a slow rate that their progress would seem to be imperceptible at
normally encountered environmental temperature. Consider for example, the
oxidation of glucose or other sugars to useable energy by animals and plants.
For a living organism to derive heat and other energy from sugar, the sugar
must be oxidized (combined with oxygen) or metabolically "burned".
However, in a living system, the oxidation of sugar must meet an additional
condition; that oxidation of sugar must proceed essentially at normal body
temperature. Obviously, sugar surrounded by sufficient oxygen would not oxidize
very rapidly at this temperature. In conjunction with a series of enzymes
created by the living organism, however, this reaction does proceed quite
rapidly at temperatures up to 100oF (38oC). Therefore, enzymes allow the living
organism to make use of the potential energy contained in sugar and other food
substances. Enzymes or biological catalysts
allow reactions that are necessary to sustain life proceed relatively quickly
at the normal environmental temperatures. Enzymes often increase the rate of a
chemical reaction between 10 and 20 million times what the speed of reaction
would be when left uncatalyzed (at a given temperature). Nutrients locked in certain
organics are complex macromolecules, or in hard-to-digest matrices may be
released or predigested by a high degree of heat or concentrated acid
treatment. In an alternative manner, specific enzymes can promote the
predigesting of certain complex nutrients and facilitate the release of highly
digestible nutrients in organics during processing without the need of
excessive heat or rigorous chemical treatment. What Comprises an Enzyme? The majority of enzymes are purely
protein in nature. Over 750 known enzymes are comprised completely of protein.
Many enzymes must have other components, however, to maintain their activity
and function. Some enzymes require various metals such as sodium, calcium,
zinc, iron, or copper to become active. Many enzymes require other organic,
non-metal components to insure their stability and/or activity. This additional
organic component is known as a "prosthetic group" or "co-enzyme".
Many vitamins are co-enzymes; they are necessary components for the activation
of certain enzyme systems within the body of living organisms. How Are Enzymes Named? One researcher reports treating
grain, sorghum or barley with the enzyme "gumase" while another
reports the same with the enzyme "beta-glucanase". When methodologies
are examined, it is discovered that both of these preparations are the same
product. Unfortunately, this apparent contradiction in terms happens often. Enzymes have been named by several
methods and this fact has been known to cause confusion in their
classification. For example, common or "trivial" names of enzymes,
generally contain a prefix representing the name of the substance or substrate
upon which they act or affect, followed by the suffix "-ase". The
"ase" simply denotes or identifies that the substance is an enzyme.
Examples of this system of nomenclature includes the enzyme that catalyzes the
conversion of proteins into their component amino acids, the name of this
enzyme is "protease" or "proteinase". Another example is the enzyme that
accelerates the breakdown of the two components of starch into sugars. The
components of starch are known as "amylose" and
"amylopectin", thus, the enzyme helping to break them down is called
"amylase". Confusion may exist, however, when
older names of enzymes are used. Included in these older terms are ficin,
pepsin, bromelin and trypsin, which are older trivial names of individual types
of protease preparations, the enzymes that accelerate digestion of proteins.
There are also many subclasses of enzymes. Amylases are a prime example;
subclasses of amylase include: alpha-amylase, beta-amylase, and gluco-amylase,
to name a few. All these enzymes do is accelerate the digestion of starch and
are broadly classified as amylases, but their actions are all slightly
different in nature. To help sort this out, the
International Union of Biochemistry in 1961 proposed a system for enzymes'
classification and naming which is finding acceptance mainly in this
discussion. One example of this system, however, is the term: "alpha 1,
4-glucan glucanohydrolase" which is a name for alpha-amylase. All these systems of nomenclature
may become confusing to someone who has use for only a few types of enzymes or
uses them for industrial or agricultural purposes. Therefore, the use of the
more widely known terms such as "amylase" and "protease"
are more or less universally in these fields. It should be remembered, however,
that there are many types of enzymes that fit into these broad categories that
may be more or less suitable for specific agriculturally related application.
The final selection for a specific application should be made only after
consulting a knowledgeable individual well-versed in the technical aspects of
the particular enzyme requirements and applicable characteristics. Where Do Enzymes Come From? Enzymes have been isolated from
every type of living organism. Many of these biological catalysts are
significant only from an academic or medical standpoint, but some of the
available enzymes from this vast repertoire have been utilized for agricultural
and industrial purposes for years. The table below lists several of the
industrially consequential enzymes and their sources in nature. It is
significant to note that animals, plants, and micro-organisms all yield
industrially important enzymes*. Some enzymes of animal or plant origin have
been used in agricultural applications; however, those enzymes most broadly
used are of microbial origin. Where Do Enzymes Come From? Source Enzyme PLANT Malted grains or tubers Amylase Pineapple Bromelin (Protease) Fig Tree Ficin (Protease) Papaya Papain (Protease) ANIMAL Liver Catalase (Peroxide Breakdown) Calf Stomach Rennet/Chymosin (Milk Clotting) Hog Stomach Pepsin (Protease) Hog Pancreas Pancreatic Enzymes (Several) Digestive Tract Trypain (Protease) MICROBIAL Fungi (Molds and Yeast) Amylase, beta glucanase, hemicellulase,
protease, cellulase, pectinase, lipase, (many types of each), lactase Bacteria Amylase, protease, isomerase, lactase (many types of
each), rennet, oxidase, catalase, beta-glucanase, hemicellulase.
In order to predigest the
potential food sources outside their cell boundaries, many microbes excrete
enzymes out through their enveloping membrane with its supportive cell wall and
into the surrounding environment. Since these "extracellular enzymes"
must function in the environment outside the protection of the cell's wall and
membrane, they must be reasonably stable and have relatively high resistance to
chemicals and must function over a relatively broad temperature range. To
realize the effects of the enzymes they produce, microorganisms also must
produce relatively large quantities of these catalysts. All of these factors
contribute to the industrial significance and durability of extracellular
microbial enzymes. It should be noted that most of
the agriculturally and industrial important enzymes, are those that catalyze
the digestion or "hydrolysis" of certain large organic molecules like
starch, cellulose, and protein. The enzymes actually attack these complex
molecules, accelerating their digestion and yielding simpler substances. Since
this process of digestion is referred to as hydrolysis, the enzymes that catalyze
the process are considered to be "hydrolyzing enzymes" or
"hydrolases". The hydrolyzing enzymes include: (1) Amylases, which catalyze the digestion of starch into small
segments of multiple sugars and into individual soluble sugars. (2) Proteases, (or proteinases), which split up proteins into
their component amino acid building blocks. (3) Lipases, which split up animal and vegetable fats and oils
into their component part: glycerol and fatty acids. (4) Cellulase (of various types) which breaks down the complex
molecule of cellulose into more digestible components of single and multiple
sugars. (5) Beta-glucanase, (or gumase) which digest one type of
vegetable gum into sugars and/or dextrins. (6) Pectinase which digests pectin and similar carbohydrates of
plant origin. How Are Microbial Enzymes Produced and How Are Microbial Enzymes
Labeled? Microbial enzymes are manufactured
by growing the microbial cells under specialized conditions so that these cells
produce their maximal amount of active enzymes. It is important to control
environmental conditions during productions so that a high percentage of these
active catalysts are preserved intact. After the microbial cells have
finished growing and producing their enzymes, they may be inactivated and
harvested along with the enzymes, or the material may be processed in various
ways in order to reach several stages of purificatory of the enzymes. Where
other concerns, like solubility or sprayability of the final enzyme-containing
product enter, the use of a semi-purified solubles or extracted, soluble
products are desirable. Depending upon the products'
degree of processing and selectivity of enzymes contained, the products are
listed as follows in the ingredients statements: Dried _________________ Fermentation Extract Dried _________________ Fermentation Solubles Dried _________________ Fermentation Product Liquid ________________ Fermentation Product The scientific name of the
microorganism used to produce the enzymatic ingredient would be substituted for
the blank provided. The AFFCO Official Publication specifies that enzyme
activity will be guaranteed for those products that represent themselves as
having enzyme activity.
Waste processing levels usually
dictate some variation in physical conditions under which the enzyme products
must function. In order to utilize enzymes to their optimal potential in
catalytic ability, we must be familiar with the basic principles that can
affect the activity and stability of these enzymes. Enzymes, being biological
compounds and being comprised of a high percentage of protein, are subject to
many environmental effects. Although the following principles hold true for
most biological enzymes produced for commercial agricultural use: The pH of the environment has a
profound affect on enzyme activity and stability. Activity optimal for pHs of
various enzymes vary; however, the optimal pH's for the biological catalysts
produced by most commercial strains of microorganisms lies between pH 4.0 and
7.5. This range is from moderately acidic to mildly alkaline in nature. These
are the pH levels normally encountered. Figure 1, indicates a difference in activity
levels that various enzymes exhibit at varying pH levels. Another major affect of enzyme
activity and stability is temperature. Since enzymes are biochemical catalysts,
made up at least partially of protein, they are sensitive in varying degrees to
heat. Raising temperatures of the environment generally multiplies the degree
of activity by the enzyme. Once an optimum temperature has been reached,
however, even higher temperatures cause rapid degradation of the enzyme with
concurrent and irreversible loss in activity (See Figure 2). Optimal
temperatures generally range from 98oF to 140oF (37oC to 60oC) for most
hydrolytic enzymes. High temperatures (over 150oF, 66oC) generally have
detrimental effects on the enzymes. However, there is broad variation in
resistance and sensitivity to heat among the enzymes' types. Bacterial enzymes
such as those from Bacillus subtilin are less sensitive to heat than are the
fungal enzymes of A. oryzae. Some amylase preparations prepared by the
fermentation of Bacillus species can withstand even boiling for short periods
and have optimal activities in the 158o-176oF (70o-80oC) range. Our laboratory
has determined that approximately 85% of the activity from B.
subtilin/licheniformis alpha-amylase survives high heat. A. oryzae amylases,
however, showed a greater than 90% loss activity in high heat. When the
enzyme-bearing, dried fermentation products of these two microorganisms are
kept dry, they are much more resistant to environmental temperature stress than
if they are moistened. In fact, very few stability problems are encountered
with most enzymes in typical situations. Basic Knowledge of Enzymes Applied Making use of general knowledge
about enzymes including how they act, under what conditions they perform, and how
to preserve their activity is important in applying the technology of enzymes
to organic waste digestion. HELPFUL REFERENCES ON ENZYMES: Aunstrup, K. 1979. In Enzyme Technology, L.B. Wingard, Jr., E.
Katchalek-Katzir, L. Goldstein (ed). Academic Press, Inc., New York. Boyer, P.D. (ed). 1971. The enzymes, 3rd, ed. Vol. 5. Academic
Press, Inc., New York. Fogarty, W. M. 1974. Enzyme technology - projects and
developments. In Projects and prospects in industrial fermentation: proceedings
of meeting held in Holly Royde. A. J. Powell and J. D. Bu'Lock (ed). U. of
Manchester. Manchester, England. Godfrey, T. and J. Reichelt (eds). 1983. Industrial enzymology:
the application of enzymes in industry. Nature Press. New York. Kulp, K. 1975. Carbohydrases and other enzymes
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