AMYLASE ECONOMIC AND APPLICATION VALUE

www.wjpr.net 014.Vol 3, Issue 3, 2

Salama et al. World Journal of Pharmaceutical Research

Α AMYLASE ECONOMIC AND APPLICATION VALUE

Nagib A. Elmarzugi1,2,3, Hesham A. El Enshasy1, Mariani AbdulHamid1, Rosnani

Hasham1, Azila Aziz1, Elsayed A. Elsayed4, Nor Zalina Othman1, Mohamed Salama5,*

1Institute of Bioproduct Development (IBD), Universiti Teknologi Malaysia (UTM), 81310

Skudai, Johor, Malaysia. 2Dept. of Industrial Pharmacy, Faculty of Pharmacy, Tripoli University, Tripoli, Libya. 3BioNano Integration Research Group, Biotechnology Research Center, NASR, Libya.

4Bioproducts Research Chair, Zoology Department, Faculty of Science, King Saud

University (KSU), Kingdom of Saudi Arabia. 5Faculty of Pharmacy, Universiti Teknologi Mara, Malaysia

ABSTRACT

The years of the research in biotechnology has boosted the importance

of enzymes in industry, as well as advanced characteristics of using

bioprocess rather than chemical synthesis in their preparation.

Enzymes are important in biological processes because they accelerate

specific biochemical reactions to produce a useful product or effect.

Enzymes are widely used in industrial processes due to their low cost,

large productivity and vast availability. Amylases are one of the most

important enzymes in present-day biotechnology. Amylases contribute

to world enzyme market about 25-30%. The estimated value of world

market of amylase industry is about US$ 2.7 billion at 2012 and

estimated to increase by 4% annually. α-amylase has variable wide

applications. Textile de-sizing based on thermo-stable bacterial

amylase covers about 12% of amylase market. α-amylase uses a very

small quantity of bio-catalysis in bio-washing products instead of very high quantity of

chemical detergents. In food production, enzymes are preferred to alternate the chemical-

based technology. α-amylase is strongly integrated with medical and pharmaceutical

applications. These applications include therapeutic and diagnostic tools for managing a quite

number of diseases ranging from ordinary problems to gene therapy by correcting the enzyme

deficiency.

World Journal of Pharmaceutical ReseaRch

7105 – 2277ISSN Article Review . 4906-4890Volume 3, Issue 3,

Article Received on 12 March 2014,

Revised on 05 April 2014, Accepted on 28 April 2014

*Correspondence for

Author

Prof. Dr.Mohamed Salama

Department of Pharmaceutics,

Faculty of Pharmacy,

Universiti Teknologi Mara,

Level 11, FF1 Building, 42300

UiTM Puncak Alam Campus,

Selangor, Malaysia.

Key Words: Enzyme, α-amylase, textile de-sizing, bio-washing, Starch.

INTRODUCTION

Enzymes play a key role in almost all biological processes, accelerating a variety of

metabolic reactions as well as controlling the energy for genetic information pathways.

Enzymes are biological catalysts responsible for supporting almost all of the chemical

reactions that maintain general homeostasis; because of their role in maintaining life

processes (Kamerlin and Warshel 2010). They are tools of nature and using natures’ own

technology in modern industries, associated with many advantages, compared to chemicals,

enzymes are specific in their action and work at mild conditions (M., S. et al. 1996). The

enzyme is also called Biocatalyst in order to catalyse any reaction. Biocatalysts are vitally

important in the industrial process to be fully exploited. Biocatalysts can be either intact cells

or isolated enzymes (Pisliakov, Cao et al. 2009). The special properties of biocatalysts

include conversion of substrates into the desired products both in single reactions and in

complex cell systems, synthesis of products under mild reaction conditions, which

approximate ambient conditions with regard to pressure, temperature and pH value.

Therefore, using of enzymes in bioprocess industry could offer a number of advantages such

as thirty percent saving in raw materials during production process, and up to thirty percent

energy savings with mechanical process in production (Bonnet and Millet 1996). Enzymes

are proteins, composed of hundreds of amino acids, which are produced by living organisms.

They are responsible for number of reaction and biological activities in plant, animal, human

beings and microorganisms. They are found in the human digestive system to break down

carbohydrates (sugars), fats or proteins present in food. The smaller pieces can be absorbed

into the blood stream; each 10-25% decrease in chlorine chemicals by substituting Xylanase

before the first bleaching step. Moreover, the use of this method in the European paper

industry could cut annual CO2 emissions by 155.000 to 270.000 tons. Therefore, reduction in

the use of chemicals, e.g. chlorine, EDTA and peroxides, by enzymatic deinking of recycled

paper by lipase, cellulases, xylanases or pectinases. The economical paper production by

efficient use of recycling paper and more protection of the environment by reducing the

volume of waste paper and the demand for wood (Namvar-Mahboub and Pakizeh 2012).

Enzymes are catalysts and most of them are protein and few ribonucleoprotein enzymes have

been discovered and, for some of these, the catalytic activity is in the RNA part rather than

the protein part (C., Q. et al. 2011). Enzymes bind temporarily to one or more of the

reactants of the reaction they catalyze. In doing so, they lower the amount of activation

energy needed and thus speed up the reaction, which is represented in figure 1.

Figure1. Effect of activation energy on chemical reaction.

History of enzymes industry

The history of modern enzyme technology could be traced to the beginning of 1874 when

Hansen manufactured Chymosin from the stomach of calves for manufacturing of cheese.

This was the first enzyme preparation of relatively high purity used for industrial purposes.

Jokichi Takamine was the first person to manufacture an enzyme from a microbial source

when he manufactured taka diastase from Aspergillus as a digestive enzyme in 1894. The

method of fermentation suggested by Takamin, the "surface culture" is still actively used in

the production of various enzymes. Bacterial amylase derived from Bacillus Subtilis was used

for desizing, the first time by Boidin and Effront as early as 1917. In 1950, fungal amylase

was used in the manufacture of specific types of syrup, those containing a range of sugars,

which could not be produced by conventional acid hydrolysis. The real turning point was

reached early in 1960, when an enzyme glucoamylase was launched for the first time, which

could completely break down starch into glucose. The process was further improved by the

introduction of a new technique used for the enzymatic pre-treatment of starch by using a

heat stable alpha amylase (Johnson 2013).

Enzyme cofactors

Numerous enzymes require the presence of an additional, non-protein and co-factor. Many of

these are metal ions such as Zn2+ that act as co-factor for carbonic anhydrase, Cu2+, Mn2+, K+,

and Na+. Some cofactors are small organic molecules called coenzymes. The B vitamins

[thiamine (B1), riboflavin (B2) and nicotinamide] are precursors of coenzymes (Sennett,

Kadirvelraj et al. 2012).

In the last few years the time for molecular biological optimization of enzymes has decreased

by a factor of 10-100, while at the same time the stability and productivity of selected

enzymes has unmistakably risen. In the past ten years, the market volume for enzymes rose

by approximately 50 %. A US study made in 2004 anticipates annual growth of the US-

American enzyme market of 6 % and a market volume of US$ 1.9 billion by 2008, the

percentage of total applications of enzymes by market sectors (Flaschel E, Bott M et al.

2006).

Table1. Distribution of the worldwide application of technical enzymes by market

sectors (Flaschel E, Bott M et al. 2006)

Market sector % share of total application Detergent 40 Cheese ripening and aroma 12 Flour and baking products 10 Leather processing 10 Glucose isomerization 7 Fruit utilization and wine 7 Starch degradation 5 Brewery (not in Germany) 5 Silage and feed 2 Pulp and paper, textile 2

Structure

According to computer modular simulation studies, the protein engineering of amylase

enzyme found in human saliva and pancreas. This enzyme can digest starch molecules and

break them down to sugar molecules. Enzyme is made of a sequence of amino acids folded

into a unique three-dimensional structure that determines the function of the enzyme. Even

the slightest change in the sequence of the amino acids can alter the shape and function of the

enzyme.Each enzyme consists of several hundreds of amino acids located in such a delicate

three-dimensional structure as in Figure 2 and 3. This structure determines the properties of

the enzyme such as reactivity, stability and specificity. Enzyme are essential for all

metabolic processes, but are distinguishable from other proteins because they are known as

biological catalysts or substances, which speed up reaction without get used up themselves.

Figure 2. Salivary alpha-amylase Figure 3. Pancreatic alpha-amylase

Production of enzymes

Enzyme molecules are too complex to synthesize by purely chemical means. The problem is

that enzyme produced by microorganisms often expressed in tiny amount mixed up with

many other enzyme and proteins. These microorganisms can also be very difficult to cultivate

under industrial condition, and they may create undesirable by-product. Modern industrial

cultivation of enzymes begins with fermentation of a vial of dried or frozen microorganisms

called a production strain. This production strain is selected to produce large amounts of the

enzymes of interest. The production strain is first cultivated in a small flask containing

nutrients and a gar, the flask placed in an incubator, which provides the optimal temperature

for the previously frozen or dried cells to germinate. Once the flask is ready, the cells are

transferred to a seed fermented, which is a large tank containing previously sterilized raw

materials and water, known as the medium, seed fermentation allows the cell to reproduce

and adapt to the environment and nutrients that they will encounter later on. The cells are

then transferred to a larger tank, the main fermented, where temperature, pH and dissolved

oxygen are carefully controlled to optimize enzyme production, and additional nutrients may

be added to enhance productivity. Therefore, when main fermentation is complete the

mixture of cells, nutrient and enzyme, referred to as the broth, is ready for filtration and

purification.

Amylase Enzyme

Microbial enzymes are widely used in industrial processes due to their low cost, large

productivity, chemical stability, environmental protection, plasticity and vast availability (N.

Sajitha, V. Vasanthabharathi et al. 2011, Yasser Bakri, Hassan Ammouneh et al. 2012).

Bacillus species such as Bacillus subtilis, Bacillus amyloliquefaciens and Bacillus

licheniformis are used as bacterial workhorses in industrial microbial cultivations for the

production of a variety of enzymes as well as fine biochemicals for decades. A large quantity

(20-25g/l) of extracellular enzymes has been produced and secreted by the various Bacillus

strains which have placed them among the most significant industrial enzyme producers. The

estimated value of world market is presently about US$ 2.7 billion and is estimated to

increase by 4% annually through 2012. Detergents (37%), textiles (12%), starch (11%),

baking (8%) and animal feed (6%) are the main industries, which use about 75% of

industrially produced enzymes (Nik C. Barbet, Ulrich Schneider et al. 1996, C. Meintanis,

K.I. Chalkou et al. 2008). Amylases are among the most important enzymes having great

significance for biotechnology, constituting a class of industrial enzymes having

approximately 25-30% of the world enzyme market (Deb, Talukdar et al. 2013). Initially the

term amylase was used originally to designate an extracellular enzymes capable of

hydrolyzing α-1,4-glucosidic linkages in polysaccharides containing three or more 1,4-α-

linked glucose units. The enzyme acts on starches, glycogen and oligosaccharides in a

random manner, liberating reducing groups. These enzymes are found in prokaryote as well

as in eukaryotic organisms. They are widely distributed in microbial, plant and animal

kingdoms (H., S. et al. 2013). Bacterial growth requires a source of energy in order to

assemble the various constituents comprising the cell. The energy supply is derived from the

controlled breakdown of various organic substrates present in the external environment, such

as polysaccharides, lipids, and proteins. Some members of the genus Bacillus are able to

break down starch and utilize it as an energy source. Organisms unable to produce amylase

cannot utilize starch as an energy source (Rothstein, Devlin et al. 1986, Pretorius I, Laing E et

al. 1988). Amylases are classified based on how they break down starch molecules into: I- α-

amylase (alpha-amylase) which reduces the viscosity of starch by breaking down the bonds at

random, therefore producing varied sized chains of glucose. II- ß-amylase (Beta-amylase)

which breaks the glucose-glucose bonds down by removing two glucose units at a time,

thereby producing maltose. III- Amyloglucosidase (AMG), which breaks successive, bonds

from the non reducing end of the straight chain, producing glucose (Carugo, Lu et al. 2001).

The Amylase enzyme has a shape that allows it to wrap around starch (substrate) and cut it up

into individual glucose units (Pandey A, Nigam P et al. 2000). α-amylase is an endo acting

enzyme, catalyzing the random hydrolysis of internal α1 to 4 glycosidic linkages present in

the starch substrate. These enzymes are incapable of hydrolysing α1 to 6 glycosidic linkages

present at branch points of amylopectin chains. One exception to this is the α-amylase

produced by thermoactinomyces vulgaris, which can hydrolyse both α1 to 6 and α1 to 4

glycosidic linkages (Vihinen and Mantsiila 1989). α-amylase has been derived from several

fungi, yeasts, bacteria and actinomycetes, however, enzymes from fungal and bacterial

sources have dominated applications in industrial sectors. Such bulk enzymes are produced

primarily by bacteria by members of the general Bacillus.Fungal α- amylases most commonly

used industrially are produced by species of Aspergillus, most notably A. oryzae and only

one species of Penicillum is Penicillum fellutanum (Flaschel E, Bott M et al. 2006). A few

industrially important enzymes may be obtained from yeast, mainly from species of

Saccharomyces. Amylase growth and production by Bacillus sp. WN11 was studied under

different cultivation conditions. Maximum growth was observed at 65 °C while the highest

amylase production was recorded at 55 °C. Accounted amylase production and growth was

observed at an initial medium pH of 5 and 6. Growth was not detected, and observed at pH 4

and 9. The organism grew better and produced higher levels of amylase activity using

proteose peptone and tryptone as nitrogen sources. Growth was not observed, when NO3+,

NH4+ and urea were used as nitrogen sources. Among defined carbohydrates, tested starch

and maltose supported good growth and amylase production, with the highest productivity

recorded in the presence of starch (24 U/ml) (Mamo and Gessesse 1999).

Amylase substrate

The starch as substrate represents the most abundant storage of polysaccharides in plants, and

next to cellulose, is the most abundant polysaccharide found on earth. The starch polymer

consists exclusively of glucose units. Two forms exist, namely α-amylose and amylopectin.

α-amylose is along, linear polymer in which successive D-glucose molecules are linked by an

α 1 to 4 glycosidic bond. Individual α-amylose chains may vary in length and hence in

molecular masses in the region of 500 kDa. Abundant supplies of starch may obtain from

seeds and tubers, such as corn, wheat, rice, tapioca and potato. The widespread availability of

starch from such inexpensive sources, coupled with large-scale production of amylolytic

enzymes, facilitates production of syrups, containing glucose, fructose or maltose, which is

considerable importance in the food and confectionery industry (Elmarzugi, Enshasy et al.

2010). The initial step in starch hydrolysis entails disruption of the starch granule.

Solubilization of the granules, the process of “gelatinization” facilitates subsequent catalytic

degradation. Gelatinization is normally, achieved by heating the starch slurry to temperatures

in excess of 100°C for several minutes. α-amylase may be added immediately prior to the

heating step, in order to render more efficient the process of granules disruption. Once the

granules have been disrupted, additional α-amylase is added in order to liquefy the starch

This process reduces the viscosity of the starch solution, and this can be shows in figure 4.

Figure 4. Controlled hydrolysis of starch, yielding up to a 96% glucose syrup.

Starch may be enzymatically hydrolysed using varying combination of amylolytic enzymes

in order to generate specific end products. The major end products produced commercially

are maltodextrins or corn syrup solids, maltose syrup, glucose syrup or high fructose corn

syrup, and cyclodextrins. Maltodextrins are produced by partial enzymatic hydrolysis of

starch, using α-amylase. The product is often defined as a non sweet nutritive saccharide mix,

largely containing glucose units, which is linked primarily by α1 to 4 glycosidic linkages.

Although the exact product composition can vary, it generally consists of glucose (approx.

1%), maltose (3-5%), maltotriose (5-10%), maltotetraose (6%), with the reminder (75% or

more) being saccharides of higher molecular mass (Rodríguez, Alameda et al. 2006). When

the desired degree of starch hydrolysis attained by α-amylase, the pH value of the slurry

dropped from 6.5 to 3.0 and the slurry, is then heated to boiling for about five minutes in

order to totally inactivate the α-amylase. Filtration and passage through a carbon column

removes any particulates and colour, and the resultant purified maltodextrin mix is

evaporated and subsequently spray dried to yield a powdered product. The reaction rates of

amylase and starch increased as the temperature increased. The absorbance levels of the

starch decreased as the temperature increased. However, after 65 degree trial, the reaction

rates starts to decrease and the absorbance levels increased (Nahas E and Waldemarin MM

2012).

Starch digestion in humans

The action of α-amylase on starch in the human body is digesting starch and producing

shorter chain glucose polymers, which are very similar to the action of α-amylase outside

human body. In humans, the salivary and pancreatic glands produce α-amylase. However, the

action of salivary amylase is limited to the hydrolysis of starch to lower degree of

polymerization polymers. The principal hydrolysis of starch by pancreatic α-amylase occurs

within the lumen of the small intestine; this action produces shorter chain polymers of lower

average polymerization degree than the intact starch molecule. The lower degree of

polymerization polymers are subsequently act on by other enzymes, including glucoamylase,

which cleaves the polymers into individual glucose monomers residues that are absorbed by

the body (Ai, Nelson et al. 2013, Elmarzugi, Enshasy et al. 2013).

The impact of genetic engineering

Genetic engineering continues to have a major impact upon the industrial enzyme sector.

Throughout the 1980s the major enzyme companies initiated research programmed aimed at

producing selected industrial enzymes by recombinant technology. Recombinant strategies

using both homologous and heterologous expression systems. Production of industrial

enzymes by recombinant means displays a number of potential advantages, as compared to

traditional (non-recombinant) approaches. These advantages may include the higher

expression levels attainable; product generally displays a higher degree of relative purity.

Moreover, it is considered economically attractive. Heterologous expression facilities

commercialization of enzymes produced naturally by pathogenic/ non-GRAS-listed species,

also, allows alteration of enzyme’s characteristics via protein engineering. The advent of

recombinant DNA technology has facilitated the cloning and expression of genes coding for

various α-amylases in a variety of recombinant organisms. Human, wheat and bacterial α-

amylases, for example, been expressed in saccharomyces cerevisiae. More recently, the gene

coding for B. licheniformis α-amylase has been expressed in transgenic tobacco plants, this

was among the first examples of the production of bulk industrial enzymes in a recombinant

plant species. Microbial amylases could be potentially useful in the pharmaceutical and fine

chemical industries if enzymes with suitable properties could be prepared. The molecular

mass of the recombinant protein was found to be 64 kDa, compared to 55 kDa in the native

Bacillus species (Deb, Talukdar et al. 2013).

Applications

α-Amylase in textile de-sizing

Initial moves to apply biotechnology have been predominantly in textile finishing.

Biotechnological processes have already proved more advantageous than chemical processes

in the processing of natural fibres, such as cotton and wool. Modern methods of textile

weaving, places considerable mechanical stress on the fabric threads to prevent breakage, the

fabric strands are nearly always coated with a substance known as a ‘size’. The serves to

strengthen the fabric prior to weaving. Essential attributes of sizing materials include good

adhesion to the textile threads, ease of removal after weaving, it is necessary to remove the

size after weaving as it would subsequently prevent proper dyeing or bleaching of the

finished product and inexpensiveness. Subsequent removal of the starch (i.e. ‘desizing’) may

be achieved by steam heating in presence of NaOH, or by oxidants. However, such

treatments can damage the textile and will generate a process effluent which must treat before

disposal. Desizing using α-amylase has thus become popular. Generally, thermo-stable

bacterial α-amylases (e.g. B. licheniformis α-amylase) are mainly used. Depending upon the

enzyme concentration and environmental parameters chosen, the desizing process may last

from several minutes to several hours (Walsh G., 2005; Flashel E. et al., 2006). The

European textile industry is exposed to fierce international competition and price wars. In the

last few years this has caused parts of the European textile production to migrate to Asia. As

the European textile industry cannot compete with Asian complying with respect to wages

and the cost of complying with environmental restrictions it is dependent on initiating new

products and new production processes, for example technical textiles (textiles with

customized and functional properties). High growth rates are particularly anticipated for

technical and functional textiles, e.g. textiles for persons with allergies, antimicrobial textiles,

textiles for hospital hygiene and those incorporated into cosmetics and pharmaceuticals. Like

textiles, pulp and paper are also often sized using starch. Sizing of paper protects it from

mechanical damage during manufacture, and enhances the stiffness and feel of the finished

product, in this case desizing is not subsequently carried out. Natural unprocessed starch

slurries are too viscous to be used in paper sizing. α-amylase is used to partially degrade the

starch in order to yield a product of appropriate viscosity for the task at hand. The world

market for enzymes for use in the pulp and paper industry has a volume of around € 10m and

an estimated annual growth 25 % (Longyun Hao, Rui Wang et al. 2013). As bio-

technological processes have proved more advantageous than chemical processes, 80 % of

the European textile industry now uses the biocatalyst amylase to remove starch from natural

fibres (e.g. cotton) during the desizing process. In various areas of textile industry the

increasing application of biotechnological processes as in sourcing cotton has enabled the

branch in the short to medium term to produce more competitively and with less pollution

and to develop more innovative products (Flaschel E, Bott M et al. 2006).

α-Amylase in detergency (bio-washing)

Enzyme is used in cleaning product as cleaning and fabric care agent. Most of the used of

enzyme types breakdown large water insoluble soils and stain which attached to fabrics into

smaller, therefore removed by mechanical action of the washing machine or by interaction

with other detergent agent. Enzyme does not loose its functionality after having worked on

one stain and continues to work on the next one. The most important reason to apply enzyme

in detergency is the use of very small quantity of these bio-catalysts that can replace very

large quantity of chemical detergent (Mitidieri, Souza Martinelli et al. 2006). Amylases are

used to remove residues of starch-based foods like potatoes, spaghetti, custards, gravies and

chocolate. This type of enzyme can be used in laundry detergency as well as in dishwashing

detergency. α-amylases have been used in detergents since the beginning of the 1970s and the

demand is still increasing. The use of a such enzyme could reduce energy consumption, since

washing cycles can be carried out at 30-40°C instead of 80-90°C and less cycles are

necessary, and consequently reduced water consumption (up to 50%). Moreover, a

distinguish saving by enzymatic removal of impurities from cotton fibres, and by using

catalases for hydrogen peroxide removal after bleaching, so a decrease in the amount of

chemicals used and a decrease in the pollution due to reduced wastewater (Roy, Rai et al.

2012). The detergent α-amylases in use today derived mainly from the Bacillus family, have

highly homologous primary amino acid sequences and a tertiary structure comprised of three

domains A, B and C. The active site is typically located in a cleft between domains A and B

and is usually comprised of acidic amino acids such as aspartic and glutamic acids. Like

many proteases, most α-amylases contain essential calcium ions, which are important in

maintaining the tertiary structure of the enzyme molecule. In 2004, a new amylase,

Stainzyme (Trade Mark), was introduced which shows a significantly stronger performance

effect in most detergent segments compared to other traditional commercial amylases. This

enzyme has a broader pH and temperature range as compared to earlier amylases, meaning

that in a typical detergent, for example, nearly equal performance is obtained at both 30◦C as

at 40◦C. Furthermore, the washing time can also be reduced but still nearly, the same wash

performance will be reached (Roy and Mukherjee 2013).

α-Amylase in food industry

α-amylases have been approved and used for many years in food manufacture. Approval of

the use of this enzyme has advantages for food manufacturers by providing a different source

of the α-amylase enzyme. The first, which has greater thermal stability and produces a

different sugar profile. There are no significant disadvantages to food manufacturers,

consumers or government agencies. In 2003 the German food industry employed around

525,000 workers in approximately 5,880 businesses and had a total turnover of around € 128

billion. However, similar to other branches of German industry, it faces increasing

international competition. Innovations could be the key to new product areas. Therefore,

experts take an optimistic view of growth in the area of functional food (Sá FC, Vasconcellos

RS et al. 2013). The world market volume for functional food could rise to about US$ 10 to

22 billion. Future prognoses anticipate annual growth rates in the world market in excess of

20 %, one reason for the increasing demand could be that many food products today, such as

chocolate pudding, ketchup, spaghetti sauce, chilli sauce, fruit puree and baby food, contain

starch in different forms (modified starches) in order to get, among other things, appropriate

viscosity (Flaschel E, Bott M et al. 2006). In food production, enzymes have a number of

advantages:

They are welcomed as alternatives to traditional chemical-based technology, and can

replace synthetic chemicals in many processes. This can allow real advances in the

environmental performance of production processes, through lower energy consumption and

biodegradability

They are more specific in their action than synthetic chemicals. Processes, which use

enzymes, therefore have fewer side reactions and waste by-products, giving higher quality

products and reducing the likelihood of pollution. They allow some processes to be carried

out which would otherwise be impossible. An example is the production of clear apple juice

concentrate, which relies on the use of the enzyme, pectinase (Marrelli M, Loizzo MR et al.

2013).

Therapeutical and Diagnostic application of α-amylase

α-amylase is widely used in pharmaceutical industry in various digestive aid preparations.

Due to presence of bacterial α-amylase, starch in the consumed food is better digested and

increase overall digestibility of food. Such digestive aid preparation is used for treatment of

patient whose digestive power is reduced due to certain illness. Many such commercial

formulations of digestive aids are seen in drug stores either as syrup or as tablet (Faulks and

Bailey 1990). The first clinical gene therapy is underway to correct an enzyme deficiency

called Adinoseine deaminase (ADA) in children. There is a correlation between salivary α-

amylase activity and pain scale in patients with chronic pain. The visual analogue scale

(VAS) is commonly used to assess pain intensity. However, the VAS is of limited value if

patients fail to reliably report. Objective assessments are therefore clearly preferable.

Previous reports suggest that elevated salivary α-amylase may reflect increased physical

stress. There is a close association between salivary α-amylase and plasma norepinephrine

under stressful physical conditions. In this study, we have determined the usefulness of a

portable salivary α-amylase analyzer as an objective biomarker of stress (Subramani and

Aalbersberg 2012).

Enzymes are found in all tissues and fluids of the body. Intracellular enzymes catalyze the

reactions of metabolic pathways. Plasma membrane enzymes regulate catalysis within cells in

response to extra cellular signals, and enzymes of the circulatory system are responsible for

regulating the clotting of blood. Almost every significant life process is dependent on enzyme

activity. α-amylase production in aqueous two phase systems by another species of bacteria

Bacillus in polyethylene glycol (PEG 10 000)/dextran (505 000) aqueous two-phase systems

at various concentrations. An increase in the PEG concentration from 7 to 25 % (w/w) in the

aqueous two-phase system resulted in an increase in the phase volume ratio with a

concomitant decrease in the partition coefficient (K) and recovery of α-amylase in the top

phase. However, varying dextran concentrations from 2.5 to 10% (w/w) decreased both the

phase volume ratio and the partition coefficient of α-amylase at dextran concentrations lower

than 2.5% (w/w) the phases could not separate (AKCAN 2011). Bacillus species have

traditionally been utilised in the production of industrially important enzymes for a number of

reasons: with the exception of the Bacillus cereus group, they are all generally recognized as

safe (GRAS) listed and they are widely distributed in nature, they are easily cultured in

relatively inexpensive media, and members of the genera Bacillus are capable of producing

large quantities of many desirable enzymatic activities, most of which are secreted

extacellularly into the fermentation medium. Extra cellular production of enzymes obviously

simplifies subsequent product recovery and purification. Enzyme therapy is an effective safe

alternative way in cancer treatment by reversing the tumour from malignant to benign. This

could be applied, through its function. Firstly, by restore the body internal environment; this

includes blood pH to be slightly alkaline, eliminate body waste, also restore intestinal

bacteria balance, strengthen the immune system, improve digestion and facilitate execration.

Secondly, enzyme has decomposition effect through breaking down the fibrin coating of the

cancer cell to unmask it in order for the immune system to identify as (non-self) and attack

and destroy it. Thirdly, throughout blood purification by eliminate wastes decompose toxin;

maintain right blood pH and fluidity; facilitated blood circulation when blood circulate well

so it can transport oxygen and nutrient to cell and cancer cell can not live in oxygen rich

environment. The enzyme therapy has a valuable advantage on conventional treatment; that

enzyme therapy neither treats the underlying cause and the whole body nor the symptom,

therefore it is considered a natural, non toxic, no side effects and easy to use.

CONCLUSION

Enzymes are tools of nature using natures’ own technology in modern industry associated

with many advantages, compared to chemicals. Amylases have a great commercial value in

biotechnological applications ranging from textile, bio washing, food, therapeutic and

pharmaceutical industries. These uses have placed greater stress on increasing amylase

production and search for more efficient processes.

ACKNOWLEDGEMENT

The authors would like to thank the following students Elrayes H. A., Mohamad M. S., Naji

S. A. for their assistance.

REFERENCES

1. Ai, Y., B. Nelson, D. F. Birt and J.-l. Jane (2013). "In vitro and in vivo digestion of

octenyl succinic starch." Carbohydrate Polymers 98(2): 1266-1271.

2. AKCAN, N. (2011). "High level production of extracellular β-galactosidase from Bacillus

licheniformis ATCC 12759 in submerged fermentation " African Journal of Microbiology

Research 5(26): 4615-4621.

3. Bonnet, P. and J. M. Millet (1996). "Catalytic Properties of Iron Phosphate-Based

Catalysts Containing Fe2(PO3OH)P2O7and α- or β-Fe3(P2O7)2in the Oxidative

Dehydrogenation of Isobutyric Acid." Journal of Catalysis 161(1): 198-205.

4. C., C. H., L. Q., S. C. M., L. J., M. D. C., S. R. R. and C. V. L. (2011). "Membrane

protein complexes catalyze both 4- and 3-hydroxylation of cinnamic acid derivatives in

monolignol biosynthesis." Proc Natl Acad Sci 108(52): 21253-21258.

5. C. Meintanis, K.I. Chalkou, K. Ar. Kormas, D.S. Lymperopoulou, E.A. Katsifas, D.G.

Hatzinikolaou and A.D. Karagouni (2008). "Application of rpoB sequence similarity

analysis, REP-PCR and BOX-PCR for the differentiation of species within the genus

Geobacillus." Letters in Applied Microbiology 46: 395-401.

6. Carugo, O., S. Lu, J. Luo, X. Gu, S. Liang, S. Strobl and S. Pongor (2001). "Structural

analysis of free and enzyme-bound amaranth α-amylase inhibitor: classification within

the knottin fold superfamily and analysis of its functional flexibility." Protein Engineering

14(9): 639-646.

7. Deb, P., S. A. Talukdar, K. Mohsina, P. K. Sarker and S. A. Sayem (2013). "Production

and partial characterization of extracellular amylase enzyme from Bacillus

amyloliquefaciens P-001." Springer Plus 2(154): 1-12.

8. Elmarzugi, N. A., H. A. E. Enshasy, E. A. Elsayed, A. Taleb, E. Keleb, S. Naji, O.

Elrmithi and N. Z. Othman (2013). "Production of α-amylase using new strain of Bacillus

polymyxa isolated from sweet potato." IOSR Journal of Pharmacy and Biological

Sciences 5(6): 73-79.

9. Elmarzugi, N. A., H. E. Enshasy, R. A. Malek, Z. Othman, M. R. Sarmidi and R. A. Aziz

(2010). Optimization of cell mass production of the probiotic strain Lactococcus Latis in

batch and fed-batch culture in pilot scale levels. Current research, technology and

education topics in applied microbiology and microbial biotechnology. A. M.-. Vilas.

Medrid - Spain, Formatex. 2: 873-879.

10. Faulks, R. M. and A. L. Bailey (1990). "Digestion of cooked starches from different food

sources by porcine α-amylase." Food Chemistry 36(3): 191-203.

11. Flaschel E, Bott M, Reuss M, Buchholz S, Scriba A, Daubmann T, Sell D, Syldatk C,

Gent R, Takors R, Liese A, Ulber R, Renken E and Zinke H (2006). White

Biotechnology: Opportunities for Germany. Germany, DECHEMA.

12. H., H. C., M. S. M. S., A. M. R., O. Z., E. E. A., S. Ramili, E. N. A., S. M. R., A. R. and

E. E. H. A. (2013). "Bioprocess Development for High Cell Mass Production of the

Probiotic Yeast-Kluyveromyces lactis." IOSR Journal of Pharmacy and Biological

Sciences 8: 49-59.

13. Johnson, K. A. (2013). "A century of enzyme kinetic analysis, 1913 to 2013." FEBS

Letters 587(17): 2753–2766.

14. Kamerlin, S. C. L. and A. Warshel (2010). "At the Dawn of the 21st Century: Is

Dynamics the Missing Link for Understanding Enzyme Catalysis?" Proteins 78(6): 1339-

15. Longyun Hao, Rui Wang, Kuanjun Fang and Jingquan Liu (2013). "Ultrasonic effect on

the desizing efficiency of α-amylase on starch-sized cotton fabrics." Carbohydrate

Polymers 96(2): 474-480.

16. M., K., G. M. S. and N. P. M. (1996). "Nature of the high azocoll lytic activity from

prawn (Metapenaeus brevicornis) muscle." Indian J Biochem Biophys 33(3): 228-230.

17. Mamo, G. and A. Gessesse (1999). "Effect of cultivation conditions on growth and

α‐amylase production by a thermophilic Bacillus sp." Letters in Applied Microbiology

29(1): 61-65.

18. Marrelli M, Loizzo MR, Nicoletti M, Menichini F and Conforti F (2013). "In vitro

investigation of the potential health benefits of wild Mediterranean dietary plants as anti-

obesity agents with α-amylase and pancreatic lipase inhibitory activities." J Sci Food

Agric. .

19. Mitidieri, S., A. H. Souza Martinelli, A. Schrank and M. H. Vainstein (2006). "Enzymatic

detergent formulation containing amylase from Aspergillus niger: A comparative study

with commercial detergent formulations." Bioresource Technology 97(10): 1217-1224.

20. N. Sajitha, V. Vasanthabharathi, R. Lakshminarayanan and S. Jayalakshmi (2011).

"Amylase from an Estuarine Bacillus megaterium." Current Research Journal of

Biological Sciences 3(2): 110-115.

21. Nahas E and Waldemarin MM (2012). "Control of amylase production and growth

characteristics of Aspergillus ochraceus." Rev Latinoam Microbiol. 44(1): 5-10.

22. Namvar-Mahboub, M. and M. Pakizeh (2012). "EXPERIMENTAL STUDY OF

LACTOSE HYDROLYSIS AND SEPARATION IN CSTR-UF MEMBRANE

REACTOR." Brazilian Journal of Chemical Engineering 29 (3): 613-618.

23. Nik C. Barbet, Ulrich Schneider, Stephen B. Helliwell, Ian Stansfield, Michael F. Tuite

and Michael N. Hall (1996). "TOR Controls Translation Initiation and Early GI

Progression in Yeast." Molecular Biology of the Cell 7: 25-42.

24. Pandey A, Nigam P, Soccol CR, Soccol VT, Singh D and M. R. (2000). "Advances in

microbial amylases." Biotechnol Appl Biochem 2: 135-152.

25. Pisliakov, A. V., J. Cao, S. C. L. Kamerlin and A. Warshel (2009). "Enzyme millisecond

conformational dynamics do not catalyze the chemical step." Proceedings of the National

Academy of Sciences 106(41): 17359-17364.

26. Pretorius I, Laing E, Pretorius GH and M. J (1988). "Expression of a Bacillus alpha-

amylase gene in yeast." Curr Genet. 14(1): 1-8.

27. Rodríguez, V. B., E. J. Alameda, J. F. M. Gallegos, A. R. Requena and A. I. G. López

(2006). "Enzymatic Hydrolysis of Soluble Starch with an α-Amylase from Bacillus

licheniformis." Biotechnology Progress 22(3): 718-722.

28. Rothstein, D. M., P. E. Devlin and R. L. Cate (1986). "Expression of alpha-amylase in

Bacillus licheniformis." Journal of Bacteriology 168(2): 839-842.

29. Roy, J. K. and A. K. Mukherjee (2013). "Applications of a high maltose forming, thermo-

stable α-amylase from an extremely alkalophilic Bacillus licheniformis strain AS08E in

food and laundry detergent industries." Biochemical Engineering Journal 77(0): 220-230.

30. Roy, J. K., S. K. Rai and A. K. Mukherjee (2012). "Characterization and application of a

detergent-stable alkaline α-amylase from Bacillus subtilis strain AS-S01a." International

Journal of Biological Macromolecules 50(1): 219-229.

31. Sá FC, Vasconcellos RS, Brunetto MA, Filho FO, Gomes MO and Carciofi AC (2013).

"Enzyme use in kibble diets formulated with wheat bran for dogs: effects on processing

and digestibility." J Anim Physiol Anim Nutr (Berl). 97(1): 51-59.

32. Sennett, N. C., R. Kadirvelraj and Z. A. Wood (2012). "Cofactor Binding Triggers a

Molecular Switch To Allosterically Activate Human UDP-α D glucose

6 Dehydrogenase." Biochemistry 51 (46): 9364–9374.

33. Subramani, R. and W. Aalbersberg (2012). "Marine actinomycetes: An ongoing source of

novel bioactive metabolites." Microbiological Research 167(10): 571-580.

34. Vihinen, M. and P. Mantsiila (1989). "Microbial Amylolytic Enzyme." Critical Reviews

in Biochemistry and Molecular Biology 24(4): 329-418.

35. Yasser Bakri, Hassan Ammouneh, Samir El-Khouri, Muhanad Harba and Philippe

Thonart (2012). "Isolation and identification of a new Bacillus strain for amylase

production." Research in Biotechnology 3(6): 51-58.

AMYLASE ECONOMIC AND APPLICATION VALUE

Documents

Transcript of AMYLASE ECONOMIC AND APPLICATION VALUE

A COMPARISON OF ADDED SALIVA, α-AMYLASE

EE 340 Economic Dispatch - UNLV

a-amylase ( boucharef & Abbad)

business & the greek economy · 2016-09-09 · 2 business & the greek economy Economic indicators Economic sentiment 2015/2014 Dec / Dec 2016/2015 June / June Economic climate ↓

Optimization of alpha-amylase production, immobilization ...jipbs.com/VolumeArticles/FullTextPDF/246_JIPBSV3I405.pdf · ... pulp and paper industries, ... Amylase isolated from bacteria

Amylolytic Yeasts: Producers of α-amylase and Pullulanase

Production of α-Amylase by Aspergillus oryzae, Penicillium ...

EE 340 Economic Dispatch - UNLVeebag/Economic Generator Dispatch.pdf · EE 340 Economic Dispatch Spring 2013. G1 G2 G3 System Incremental cost curves. 975 $9.163/MWh 9.16 9.16 9.16

Sparinvest Global Value

Presentation on Amylase enzyme

In Vitro Antioxidant and α-Amylase Inhibition Activities .... Engida Dessalegn.pdf · Alpha-amylase inhibition activity • 1 mL of α-amylase (from Aspergillus oryzae) was added

Production and optimization of α-amylase from thermo ...

ASSAY PROCEDURE (AMYLOGLUCOSIDASE/ α-AMYLASE METHOD)

AMYLOGLUCOSIDASE/ α-AMYLASE METHOD

Value Investing During an Economic Crisis: The Case of Greece

Simultaneous Increase of Glucoamylase and -Amylase ...

ARCHIVES OF ECONOMIC HISTORY€¦ · ARCHIVES OF ECONOMIC HISTORY ... times at Constantinople. ... EL. RIGA: Banks in ancient Greece ...

Immobilization of B. amyloliquefaciens α-amylase and ...

ALPHA-AMYLASE - Megazyme · ALPHA-AMYLASE ASSAY PROCEDURE (AMYLASE SD METHOD) K-AMYLSD 04/19 HIGH SENSITIVITY METHOD FOR THE MEASUREMENT OF α-AMYLASE IN CEREAL GRAINS AND FOOD PRODUCTS

Εγκεκριμένος Οικονομικός Φορέας Authorized Economic Operator (AEO)