FR2570715A1 - PROCESS FOR CARRYING OUT AN ENZYMATIC OR MICROBIAL REACTION - Google Patents

Abstract

DEPENDING ON THE INVENTION, THE ENZYMATIC OR MICROBIAL REACTION BETWEEN AT LEAST TWO SUBSTANCES TO BE REACTED BY PLACING AN ENZYME, A MICROBE, AN IMMOBILIZED ENZYME OR A MICROBE IMMOBILIZED IN AN INTERNAL SPACE BETWEEN TWO SHEETS OF NON-POROUS MATERIALS 'ENZYME NOR MICROBE TO PASS, BY PLACING THE SUBSTANCES IN THE TWO EXTERNAL HOUSEHOLD SPACES RESPECTIVELY OUTSIDE THE TWO SHEETS, ALLOWING EACH OF THE SUBSTANCES TO PASS THROUGH A SHEET TO PERFORM THE ENZYMATIC REACTION OR THE MICROBITIC INTERNAL SPACE, AND ALLOWING EACH REACTION PRODUCT TO PASS TO THE EXTERIOR THROUGH A SHEET.

Description

-12- The present invention relates to a new method of implementation

  an enzymatic or microbial reaction, and

  more particularly a process for implementing a reaction

  Enzymatic or microbial action in which at least two substances come into play, which includes the use of a bioreactor which is constructed in such a way that an enzyme or a microbe is placed in an internal space (a compartment for enzyme) defined by two sheets of material

  porous, such as membranes, which do not allow the en-

  zyme or microbe to pass freely through them, and the substances participating in the reaction are placed in external spaces in contact with the two porous materials, so that these substances are free to pass through porous materials in the internal space to be thus subjected to the reaction in the compartment and

  reaction products can pass through materials

  porous membranes in the spaces outside the compartment, whereby the enzymatic or microbial reaction thus

  that the separation of the reaction products can be effected

effectively killed.

  In most enzymatic and micro- reactions

  well, at least two substrates and in addition an activator other than said substrates are usually required to carry out the reaction and it is rare that only a single substance is involved in the reaction. Also, at least two reaction products are usually formed and it is unusual for only one reaction product to be formed. As a result, it is common for at least two

  substrates for the reaction and at least two reaction products

  tion come into play in the enzymatic or micro- reaction

  bienne. However, when two or more reaction products are formed, it is necessary to separate these reaction products after the end of the reaction to obtain the desired products. In addition, in the case of a reaction using at least two substrates, one of them can sometimes have an inhibiting or inactivating effect on the enzyme or on the microbe. It is even sometimes possible that a reaction product itself acts as an inhibitor. Thus, enzymatic and microbial reactions can be carried out more efficiently if the amounts of these inhibitors introduced into the enzymatic or microbial reaction system can be controlled or if such

  inhibitors can be quickly evacuated from the reactive system

  tional. In addition, it is possible to omit a step for separating the reaction products if it is possible to use

  a reactor which also serves to separate the reaction products.

  The enzyme is usually so costly that its reuse

  sation is essential to reduce the cost of production.

  Therefore, it is very important for a bioreactor that an enzyme or a microbe can be easily removed

  of the reaction system for reuse.

  A bioreactor capable of responding to all of these

  gences has not yet been developed. For example, although an enzyme or a microbe can be separated from a reaction system by the use of an immobilized enzyme or microbe, or their reuse is possible, it is impossible to fulfill the conditions required for the implementation simultaneous reaction and separation and for the control of the inhibitor. An attempt was

  made in recent years to give a separation function

  tion of products (adsorbing power) to an immobilized support such as an immobilized enzyme or microbe. However, this method has various drawbacks. For example, when a substance acting as an inhibitor or inactivator is required for the reaction, it is difficult by this method to control the amount of the substance. When the adsorption capacity of the support is to be used, a very large amount of the support is necessary to adsorb the -3-

  products and this represents an obstacle to the industrial application

  trielle. The authors of the present invention have studied a

  reactor to be used in an enzymatic or micro reaction

  bienne and they developed a reactor capable of responding

to the requirements described above.

  The present invention therefore provides a method for carrying out an enzymatic or microbial reaction

  involving at least two substances, which is characterized

  laughed at by the fact that two pairs of porous materials are used, in pairs, which do not allow an enzyme or a microbe to pass through them, an enzyme is placed in an internal space formed between these porous materials , a microbe, an immobilized enzyme or an immobilized microbe, we place in each of the two external spaces

  formed outside of the porous materials each of the

  stanzas involved in the reaction, and these sub-

  stanzas of passing through porous materials in space

  internal to enter a reaction in this space.

  The reaction process according to the present invention is characterized in that two sheets of porous materials, such as membranes, are used in pairs which do not allow an enzyme or a microbe to pass freely through them , an enzyme or a microbe is placed in an internal space defined by the porous materials, the substances involved in the reaction are allowed to pass through the porous materials in the internal space to undergo an enzymatic or microbial reaction, and allows reaction products to pass, after reaction, through porous materials to be removed from the system

  enzymatic or microbial reaction.

  Figure 1 schematically shows an embodiment of a device used in the present invention; in this figure, 1 and 2 represent porous materials, 3 and 5 represent the compartments containing the bodies in reaction, -4-

  4 represents a compartment containing an enzyme and 6 represents

feels an outer envelope.

  FIG. 2 schematically represents an embodiment of a plane membrane type reactor according to the present invention. FIG. 3 schematically represents an embodiment

of a tubular type reactor.

  FIG. 4 schematically represents an embodiment

of a spiral-type reactor.

  The present invention is described in more detail in

  referring to the attached drawings which represent realizations

  preferred locations of the present invention.

  A lipolysis reaction using a lipase is taken as an example of the enzymatic reaction which can be represented by the formula: A + B -C + D (in which A and B are the substances involved in the reaction and

  C and D are the products; for example, in the reaction used

  sant a lipase, A is an oil, B is water, C is a fatty acid and D is glycerol). Referring to FIG. 1, an enzyme is placed in an internal space 4 (enzyme compartment) defined by the porous materials 1 and 2, a substrate A is placed in an external space 3 (body in reaction) and a substrate B is placed in another space

  external 5 (body in reaction). The materials

  porous rials 1 and 2 are made in such a way that enzymes cannot pass through them, substrate A is free to pass through porous material 1 and substrate B is free to pass through porous material 2. In the reactor

  in Figure 1, the substrates A and B placed in the compartments

  respective reacting body elements 3 and 5 can pass

  through the respective porous materials in the compartment

  enzyme 4 where they undergo an enzymatic reaction in the presence of the enzyme to form the products C and D. A preferred porous material 1 which can be used is a material which allows the substrate A to pass through -5-

  this one but which does not allow the substrate B to pass to-

  through it and which allows only one of the reaction products C and D (for example the product C) to pass through it. On the other hand, a preferred porous material 2 is a material which allows the substrate B to pass through it but which does not allow the substrate A to pass through it and which only allows

  the reaction product D to pass through it.

  Porous materials having such properties can be obtained by adequately choosing the quality and size of the pores of the porous material. In the reaction using a lipase, a hydrophobic material such as polypropylene or polyethylene can be used as a porous material which allows only the oil as a substrate and the fatty acid as the reaction product to pass through it, and a hydrophilic material such as a cellulose material (cellulose acetate for example) or an acrylonitrile polymer can be used as

  that porous material that only allows water and gly-

  cerol to get through it. The pore size of porous materials can be such that it does not allow

  not the enzyme or the microbe to pass through them.

  As the size of the enzyme is considered to be of the order of a few tens to a few hundred A (= 10-4 μm) the porous materials must have a pore size of this order. However, in some cases, a pore size

  as small is not necessary, depending on the reaction systems

  tional. For example, in the case of lipolysis with a lipase in the space 4 defined by the porous materials, an oil / water emulsion is formed and the enzyme attaches to the surface of the particles of the emulsion. Consequently, the porous materials can have a pore size such that the emulsion cannot pass through them, and a pore size ranging from 0.1 to a few μm is sufficient to prevent the passage of the enzyme. through porous materials. When using an immobilized enzyme or microbe, it is not necessary

to have small pores.

  Consequently, one can choose porous materials

  depending on the purpose and use, without restrictions

  particular pation regarding the quality or size of the pores, insofar as these can satisfy the

above requirements.

  An enzyme or a microbe which must be placed in the space 4 defined by the porous materials can be found

  in the form of a powder or solution or suspension

  ion in a solvent which does not inactivate either the enzyme or the microbe.

  In the practice of the present invention, it is preferred that a spacer is placed inside the space 4 and that the enzyme or the microbe is spread or deposited

  on the spacer to allow even distribution

  enzyme or microbe gene. Alternatively, it is possible to circulate in space 4 (the enzyme compartment 4) a solution containing the enzyme or the microbial of a pump as described in more detail below. The spacer which must be placed inside the space 4 can be used without any particular restriction concerning the material, the shape, etc. For example, when a material capable of adsorbing an enzyme, such as an ion exchange fiber, is used as the spacer, the enzyme can be immobilized thereon. In addition, we can give

  the spacer an additional function.

  The substrates present in the reaction body compartments 3 and 5 diffuse into the enzyme compartment 4 by diffusion due to concentration, and the reaction products diffuse from the enzyme compartment 4 into the reaction body compartment 3 or 5 also by concentration diffusion. However, when the diffusion speed due only to the concentration diffusion is too low and that a sufficient reaction speed cannot be obtained, the diffusion can be accelerated by applying a force accelerating the diffusion in each compartment, pressure or temperature for example. Alternatively, it is possible to apply a force accelerating the diffusion by using a difference in osmotic pressure caused by the addition in each compartment of a third substance which does not interfere with either the enzymatic reaction or with the microbial reaction. and which does not cross either

porous material.

  The bioreactor used in the present invention

  can be applied to various enzymatic and micro reactions

  biennes. Examples of such enzymatic reactions in which at least two substances are involved include hydrolysis with a lipase already mentioned, synthesis of triglecrides with a lipase, an ester exchange reaction

  triglycerides in the presence of lipase, the hydrolysis of phospo-

  lipids with a phospholipase, a series of reactions with a transferase as represented by the phosphorolysis of polysaccharide in the presence of phosphorylase, the synthesis of malic acid using fumarase, a series of reactions in the presence of a hydrolase, such as phosphatase or nucleotidase, and a series of reactions in the presence of a

  oxidoreductase such as alcohol dehydrogenase or amino acid-

  oxidoreductase. In addition, the reactor of the present invention can be applied to most microbial reactions, since many substances in addition to the substrates are required for the growth of microbes. For example, glutamic acid can be produced from a substrate consisting of glucose in the fermentation process of glutamic acid using a strain of the microorganism Corynebacterium

  qlutamicum. However, biotin is required for growth

  strain and the glutamic acid yield varies

  depending on the amount of biotin. Micro- reactions

  biennes frequently require vitamins such as biotin, mineral salts, amino acids, etc., in addition to the substrate as in the above microbial reaction, and the reactor of the present invention can be applied

  to most microbial reactions.

  The enzymes and microbes used in the present invention are not always of the highly purified type and may be in the form of an extract, a

  partially purified product or fermented liquor

  tion. In order to effectively carry out the reaction with the bioreactor of the present invention, the bioreactor is constructed, as indicated in FIG. 2, in such a way that several reaction compartments are formed in which each compartment is formed by interposition of 'an enzyme compartment 7 between porous materials 9 and 10, so that porous materials of the same type face each other, a reaction body compartment 5

  is formed between the two porous materials 9 and 9, and a compar-

  timent of reacting bodies 6 is formed between the two other porous materials 10 and 10. Each of the reacting bodies is introduced into each of the reacting body compartments 5 and 6, each by means of pumps 3a and 3b. In the reactor of FIG. 2, it is also possible to introduce an enzyme or a

  microbe in the enzyme compartment 7 by means of a pump 3.

  In FIG. 2, the numbers 1 and 2 each represent a reservoir of reacting bodies, 4 represents a reservoir

  enzyme or microbe, and 8 represents an outer shell.

  Alternatively, the enzyme or microbe can be placed on a

  spacer by placing the spacer inside

  laughing at enzyme compartment 7 and spreading or depositing

  sant the enzyme or the microbe on it, as described

upper.

  The reactor used in the present invention is not limited to a flat membrane type as indicated in FIGS. 1 and 2, but it can have any shape, for example that of a tube as indicated on the figure 3,

  or of a spiral as it is identified on figure 4.

  -9- In the round reactor of FIG. 3, an enzyme compartment 4 is formed between the porous materials 1 and 2, a

  Reaction body compartment 3 is provided between the mem-

  tubular brane 1 consisting of the porous material, a comparison

  timent of reacting body 5 is formed outside the tubular membrane 2 formed by the porous material, and 6 represents an external envelope. In the spiral reactor of FIG. 4, an intermediate sheet 2 on which an enzyme is placed is inserted into the space formed between the membranes 1 and 3 composed of a porous material to form an enzyme compartment. The membranes 1 and 3 are placed between two sheets of outer casing 4 and 4 and they are wound in a spiral shape. A body compartment

  in reaction is formed between an external envelope and the mem-

  brane 1, and another reacting body compartment is

  formed between the other outer shell and the membrane 3.

  One of the features of the present invention

  is that an enzyme or a microbe can be easily removed

  reaction system res, as in the case of an enzyme

  immobilized, and the enzyme or microbe is not immo-

  bilised on porous materials, so that it is possible to exchange only the enzyme or the microbe when they have

  lost their activity, without having to change porous materials.

  Porous materials which are expensive can thus be preserved. In addition, when inhibitors are involved in the enzymatic or microbial reaction, it is possible to control

  the amounts of these inhibitors to be added to the reaction system

  tional by choosing the materials constituting the membranes or by choosing the size of their pores. When a reaction product acts as an inhibitor, the reaction product can be quickly removed from the reaction system. As a result, the inactivation of the enzyme or microbe that can be reused can be prevented. For example, in the case of lipolysis with a lipase, water as a substrate

  acts as a lipase inactivation factor.

  Therefore, when the amount of water increases excessively, the enzyme is considerably inactivated. When it is possible to control the quantity of water passing through the porous material, it is possible to introduce into the reaction system only the quantity of water required and it is possible to avoid

inactivation of lipase.

  As described above, the amounts of substrates

  and reaction products in an enzy- reaction system

  materials of the present invention are frequently higher

  than those present in conventional enzymatic reactions.

  In addition, the concentration of enzyme in the reaction system

  is generally higher than in conventional systems.

  Thus, the environment of the enzyme in the reaction system according to the present invention is different from those existing in conventional systems, which can be a source of

  various advantages. For example, the lipolytic reaction used

  Health of a previously cited lipase has resulted in an increase in the thermal stability of the enzyme. That is to say, the use of the reactor of the present invention makes it possible to carry out the lipolytic reaction with an unstable lipase.

  with heat, at a temperature of 40 to 60 C, with little inactivity

  tivation, although this lipase is considered to be inadequate for the hydrolysis of oil with a high melting point like beef tallow, since it is normally unstable

  heat and should be used at a lower temperature

  less than 40 C. Consequently, the process of the present

  invention sometimes changes the properties of the enzyme itself

  even such as thermal stability or optimal value of

  pH, depending on the conditions of the reaction system.

  Another feature of the present invention is that the reaction and separation of

  reaction products can be performed simultaneously.

  For example, the reaction and separation of fatty acids and glycerol formed as reaction products can be carried out simultaneously in the case of lipolysis with -11 -

lipase.

  The reaction speed in the inventor's bioreactor

  This largely depends on the rate of transfer of a reacting body through the porous membrane of the reactor and the area of the membrane. It is therefore important to choose a porous material allowing substrates

  to pass through it at a high speed in order to increase

  lie about the reaction rate. In addition, since the reaction rate depends largely on the surface area of the membranes, the reaction time can be considerably reduced by

increasing the area.

  As described above, the bioreactor of the

  present invention provides a macro- immobilization method

  scopic of an enzyme or microbe between materials

  porous, in which the enzyme or microbe can be facilitated

  Lately removed from the reaction system. In addition, it makes

  possible reuse of the enzyme or microbe and inhibit

  bition of the inactivation of these, as well as the simultaneous reaction and separation of the reaction products. The bioreactor of the present invention therefore provides an entirely new reaction process with various advantages not offered by conventional processes which use an immobilized enzyme. Consequently, the method of the present

  invention is very useful from the industrial point of view.

  The present invention is illustrated by the examples

  descriptive and non-limiting below.

Example 1

  10 sheets of porous membranes (ultra-

  filter having a fractional molecular weight of 20,000) composed of polyacrylonitrile having a surface area of 0.02 m2, and 10 sheets of porous membranes (pore size: 0.1 μm) composed of Teflony (commercial designation of a fluorinated resin ). 2 g of a lipolytic enzyme, a lipase, was suspended in a small amount of soybean oil

  whose activity was 320,000 U / g. On an inter-

  -12- sheet to be placed between the two membranes was deposited about 1/10 of the suspension by interlayer sheet, and the sheet

  interlayer has been placed between the two membranes, provided

  thus being a plane membrane type reactor as shown

  felt in FIG. 2. In the process of this example - the enzyme is placed on the interleaf by depositing the enzyme solution thereon, while the enzyme solution circulates

  by means of a pump in the process shown in Figure 2.

  400 g of soybean oil and 400 g of water were circulated so that the soybean oil was contacted with the Teflon membrane and the water was contacted with the polyacrylonitrile membrane . The temperature was maintained at 30 ° C. in a constant temperature bath

  to hydrolyze soybean oil in the presence of lipase.

  After 24 hours, the rate of hydrolysis of soybean oil was 70%, and the concentration of glycerol in water was 7%. The glycerol content of the oil and the

  fatty acid content of the water was each 0.1% or less.

  From the above results, it is evident that soybean oil and water can pass through the porous membranes in the central compartment containing the enzyme where they react, and after the reaction the fatty acids can pass through the porous membrane and diffuse into the compartment containing the oil, while glycerol can pass through the porous membrane and diffuse into the compartment containing the water. In this way the reaction and the separation of the reaction products can

be performed simultaneously.

Example 2

  The enzyme used in Example 1 was left as it was in the reactor and the fatty acid solution and an aqueous glycerol solution were removed after the reaction of Example 1. 400 g of new soybean oil and 400 g of new water were introduced into the reactor in a manner similar to that described in Example 1. After 24 hours, the hydrolysis rate of the soybean oil was 70%, the same

as in example 1.

  After which, the enzyme was left as it was in the reactor and 400 g of new soybean oil and 400 g of new water were introduced into the reactor. After 24 hours, the third experiment showed that the rate

  soybean oil hydrolysis was 70%.

  Consequently, no inactivation of the enzyme

  manifested in the bioreactor of the present invention.

  The reaction was carried out effectively leaving

  sant the enzyme as it is in the reactor, by evacuating only the solutions after the reaction and by introducing

new oil and water.

  It is clear that a step in the separation of the enzyme

out of the system is useless.

Example 3

  This experiment was carried out under the same conditions as those of Example 1, except that it was used as porous membranes to be brought into contact with the oil 10 sheets of polyvinyl chloride (area: 0, 02 m2, pore size: 2.5 pm). The porous membranes that were to

  come into contact with water were poly membranes

  acrylonitrile as in Example 1. Under conditions similar to those of Example 1, 400 g of soybean oil

  and 400 g of water were introduced into the reactor by circulating

  lation to perform lipolysis. When polyvinyl chloride membranes were used, water diffusion was found in the oil compartment and the side of the oil compartments was therefore submitted

  at a pressure difference of approximately 0.03 kg / cm2 to prevent

  expensive water to diffuse into the oil compartment.

  After 24 hours, the soybean oil hydrolysis rate was 80% and the concentration of glycerol in water was 8.1%. After 48 hours, the rate of hydrolysis of soybean oil was 90% and the concentration of glycerol -14-

in water was 9%.

  It was found that the speed of the reaction varied as a function of the material constituting the membrane and of the size of its pores, by comparing the results with those of Example 1.

Example 4

  The same porous membranes were used as those of Example 1, except that the number of membranes was equal to 5. 400 g of soybean oil and 400 g of water were introduced into the reactor. After 24 hours, the rate of hydrolysis

soybean oil was 44%.

  It was therefore found that the reaction speed depended

  to a large extent of the area of the membrane.

Example 5

  5 sheets of acetate membranes were prepared.

  cellulose (reverse osmosis membranes with an elimination rate

  10% ordinary salt) with an area of 0.02 m2,

  and 5 sheets of porous membranes (pore size: 0.1 µm).

  These membranes were placed as shown in Figure 2 to thereby form a porous structure. Using a pump, it was circulated between the acetate membranes

  cellulose and polyethylene a suspension of compressed enzyme

  2.4 g of lipase and 300 g of soybean oil, while circulating using a pump 300 g of soybean oil and 300 g of water in such a way as oil and water were brought into contact with the polyethylene membrane respectively

  and with the cellulose acetate membrane. The reaction system

  was kept at 500C in a temperature container

  constant to hydrolyze soybean oil with lipase.

  After 48 hours, the rate of hydrolysis of soybean oil was 92%. The fatty acid solution and the aqueous glycerol solution were then removed while leaving the enzyme as it was and 300 g of new soybean oil and as much of the following were introduced into the reactor, as described above. water. After 48 hours, the rate of hydrolysis of soybean oil was 92%. 300 g of new soybean oil and as much water were then introduced into the reactor.

  while leaving the enzyme as is.

  As a result of the third reaction, the same rate of soybean oil hydrolysis was obtained after 48 hours (i.e.

  92%). The reactor of the present invention has therefore made it possible to

  the recovery of the product and the reuse of the

  zyme while maintaining the enzymatic activity at high temperature (namely 50 C) without alteration: - of the properties of the enzyme itself or prior treatment thereof, such as immobilization. Comparative Example The procedure of Example 5 was followed, except that the substrate (namely the soybean oil), the water and the lipase were mixed with stirring without using the membrane reactor to reuse thus. the enzyme. 300 g of soybean oil, 300 g of water and 2.4 g of the enzyme were mixed with stirring at 50 ° C. as in Example 5. The rate of hydrolysis of the soybean oil was 92% after 48 hours. After 48 hours, the mixture was centrifuged

  reaction at 8,000 rpm for 5 minutes at 50 C.

  The reaction mixture was thus separated into three layers, i.e. an oil layer (upper), a layer

  emulsion (intermediate) and an aqueous layer (lower).

  About 90% of the enzyme was distributed in the emulsion layer.

  sion while the remaining 10% of it was

  in the aqueous phase. We then introduced the emulsion layer.

  sion in the subsequent enzymatic reaction while subjecting the aqueous phase to ultrafiltration to recover by this means the enzyme contained therein. Namely, the aqueous phase was subjected to ultrafiltration through a

  polyacrylonitrile membrane with fractional power-

  20,000 molecular weight, 9.81 x 104 Pa (1 kg / cm2).

  The enzyme remaining in the membrane was then collected

  and reused in the subsequent enzymatic reaction.

  -1 6- The concentrated enzyme solution thus recovered from the emulsion layer and from the membrane was then added to 300 g of soybean oil and as much water and stirred

  for 48 hours at 50 C as for the first reaction.

  In the second reaction, the rate of hydrolysis of soybean oil was 45%. The enzyme was recovered by the same separation technique and subjected to the third reaction. In the third reaction, the rate of hydrolysis

soybean oil was 6%.

  The activity of the enzyme therefore considerably decreased in the process in which the lipase was mixed with oil and water with stirring without any membrane reactor, at a high temperature (50 ° C.), as the number of reactions increased, so it was

cannot reuse the enzyme.

Example 6

  The procedure of Example 5 was followed, except that the soybean oil was replaced by beef tallow which was solid at room temperature. After carrying out the reaction at 50 ° C. for 48 hours, the rate of hydrolysis of beef tallow was 90%. The second and the third reaction. also gave, after 48 hours,

a hydrolysis rate of 90% each.

  Solid fats such as beef tallow can therefore be hydrolyzed with little inactivation using

  the reactor of the present invention.

-1 7-

Claims (4)

  1. Method for carrying out an enzymatic reaction   or microbial between at least two substances to react, charac-   terized in that it comprises the steps consisting in poaching an enzyme, a microbe, an immobilized enzyme or a immobilized microbe in an internal space formed between two sheets of porous materials which do not allow the enzyme or the microbe to pass, placing the substances in the two external spaces provided respectively outside the two sheets, allowing each of the substances to pass through a sheet to carry out the enzymatic or microbial reaction in the internal space, and allowing each reaction product to pass outside through leaf.   2. Method according to claim 1, characterized in that one of the sheets easily allows only one of the substances and only one of the reaction products to pass through it, and the other easily allows the other substance and to the other   reaction product to pass through it.   3. Method according to claim 1 or 2, characterized in that several pairs of two sheets are provided and aligned so that two sheets of the   same porous material face each other.   4. Method according to one of claims 1 to 3, ca   characterized in that the hydrolysis of oils and fats is carried out using water and oil as   substances to react and a lipase as an enzyme.