CA1077772A - Method of recovering protein of low nucleic acid content from microorganisms - Google Patents
Method of recovering protein of low nucleic acid content from microorganismsInfo
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- CA1077772A CA1077772A CA277,828A CA277828A CA1077772A CA 1077772 A CA1077772 A CA 1077772A CA 277828 A CA277828 A CA 277828A CA 1077772 A CA1077772 A CA 1077772A
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/08—Reducing the nucleic acid content
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- General Health & Medical Sciences (AREA)
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- Preparation Of Compounds By Using Micro-Organisms (AREA)
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Abstract
METHOD OF RECOVERING PROTEIN OF LOW NUCLEIC ACID
CONTENT FROM MICROORGANISMS
Abstract of the Disclosure Proteins of reduced nucleic acid content are obtained from micro-organisms by thermal treatment comprising heating an aqueous suspension of the micro-organism with the optional addi-tion of a material having ribonuclease activity, at a pH value between 7 and 8.5 and at 63 to 67°C, until the 5'-nucleotide content in the medium has reached the desired level, then briefly boiling the resulting mixture and separating the insoluble crude protein from the suspension.
CONTENT FROM MICROORGANISMS
Abstract of the Disclosure Proteins of reduced nucleic acid content are obtained from micro-organisms by thermal treatment comprising heating an aqueous suspension of the micro-organism with the optional addi-tion of a material having ribonuclease activity, at a pH value between 7 and 8.5 and at 63 to 67°C, until the 5'-nucleotide content in the medium has reached the desired level, then briefly boiling the resulting mixture and separating the insoluble crude protein from the suspension.
Description
,1 , :~ 1 The present invention relates to the recovery of proteins from microorganisms, particularly to the obtaining of protein having a reduced nucleic acid content from such materials.
I~ is known that the population of the world is rapidly increasing and therefore an acute shortage of food is to be expected in a few centuries. In many countries this situa-tion already prevails, Protein deficiency constitutes the main problem. ~an's daily protein requirement amo~nts to approximately 70 to lO0 g. In underdeveloped countries this protein requirement is covered mainly by vegetable foods. In contrast to animal protein, however, vegetable protein contains very little of the two essential amino acids, lysine and methionine. I~ only vegetable protein is available, therefore, severe deficiency diseases can occur. For this reason attempts have been made for years to utilize microbial protein in addition to animal and vegetable protein to cover the protein needs of the world~population. Microbial protein has the advantage that it has an amino acid composition similar to that of animal protein. In a diet of microbial protein there is no need to fear the occurrence of the deficiency diseases which can arise in the case of an all-vegetable diet.
.
For decades attempts have been made to use microorgan-isms for the production of foods for human beings and animals.
These minute forms of life have the advantage that they grow, under favorable conditions, much more rapidly than plants or animals. The following table illustrates this.
:
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DOUBLING TIME FOR CERTAIN FORMS OF LIFE
Organisms Biomass Doubling Time 5 Yeasts and bacteria 20 - 120 minutes Microorganisms such as algae 1 - 48 hours Grass 1 - 2 weeks Chicks 2 - 4 weeks Swine 4 - 6 weeks Cattle 1 - 2 months .
The rapid proliferation of microorganisms shows that more protein can be obtained from them in 1 to 2 hours than 5 can be obtained from swine or cattle in 4 to 8 weeks.
However, this kind of protein production entails a number of difficulties:
1. A very great technical investment (gigantic fer menters) is required for the growth of microor-ganisms;
I~ is known that the population of the world is rapidly increasing and therefore an acute shortage of food is to be expected in a few centuries. In many countries this situa-tion already prevails, Protein deficiency constitutes the main problem. ~an's daily protein requirement amo~nts to approximately 70 to lO0 g. In underdeveloped countries this protein requirement is covered mainly by vegetable foods. In contrast to animal protein, however, vegetable protein contains very little of the two essential amino acids, lysine and methionine. I~ only vegetable protein is available, therefore, severe deficiency diseases can occur. For this reason attempts have been made for years to utilize microbial protein in addition to animal and vegetable protein to cover the protein needs of the world~population. Microbial protein has the advantage that it has an amino acid composition similar to that of animal protein. In a diet of microbial protein there is no need to fear the occurrence of the deficiency diseases which can arise in the case of an all-vegetable diet.
.
For decades attempts have been made to use microorgan-isms for the production of foods for human beings and animals.
These minute forms of life have the advantage that they grow, under favorable conditions, much more rapidly than plants or animals. The following table illustrates this.
:
~ .
7777~
DOUBLING TIME FOR CERTAIN FORMS OF LIFE
Organisms Biomass Doubling Time 5 Yeasts and bacteria 20 - 120 minutes Microorganisms such as algae 1 - 48 hours Grass 1 - 2 weeks Chicks 2 - 4 weeks Swine 4 - 6 weeks Cattle 1 - 2 months .
The rapid proliferation of microorganisms shows that more protein can be obtained from them in 1 to 2 hours than 5 can be obtained from swine or cattle in 4 to 8 weeks.
However, this kind of protein production entails a number of difficulties:
1. A very great technical investment (gigantic fer menters) is required for the growth of microor-ganisms;
2. Thq separation of the organisms from the culture broths on the required scale presents technical difficulties;
3. Undesirable substances (e.g., nucleic acids and toxins) have~to be removed from the microrganisms. -~
:
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In spite of these difficulties, microorganisms are already being cultivated on a large scale for the feeding of domestic animals (fodder yeast). For human nutrition, however, this protein from microorganisms has hitherto been used either not at all or only to a limited extent. The high ribonucleic acid content of this protein, for example, constitutes a serious problem. ~ibonucleic acld (~A) is decomposed in the mammalian organism to uric acid, which in turn can cause gout. The ~NA content of microorganlsm protein must therefore be less than 2%, and preferably less than 0.5%.
One of the most important aims, therefore, is to pro-duce the microbial protein free of ribonucleic acid. This has not been accomplished in a satisfactory manner to date.
Attempts have been made to breed microorganisms of an espe-cially low RNA content. These microorganisms have the dis-!
, advantage, however, that they grow considerably more slowly.
It is known that microorganisms of high RNA content grow more rapidly than those of low RNA content. This results in areal dilemma: the desired fast growth of the microorganisms is to be equated with a high RMA content. If slower produc-tion can be accepted, microorganisms of low RNA content can be cultivated.
The second possibility for eliminating RNA from micro-rganism protein, which is frequently referred to as single-1~t7777Z
cell protein ~SCP)), is based upon the extraction of theRNA followed by separation of the insoluble protein. Sev-eral possibilities for such extraction of RNA have been described as follows:
5 a) Alkaline extraction - German Offenlegungsschrift 16 70 110.3 b) Acid extraction i c) Treatment with high salt concentration (NaCl, KCl, NH4Cl) ~ , d) Treatment with detergents (dodecyl sulfate etc.).
All of these methods have the disadvantage that the microorganisms in some cases have to be processed in a very complicated manner, and the protein is partially hydrolyzed, racemized or otherwise altered by the treatment and by the various additives and thus becomes less suitable if not use-less for nutritive purposes. For example, alkaline extrac-tion modifies the protein in its structure such that it forms colloidal solutions. Separatlon is possible only after neutralization and accordingly necessitates the re-moval of the neutral salts that are formed. Furthermore, the dissolved RNA becomes partially denatured, and can no longer be quantitatively cleaved with enzymes to form soluble mono-nucleotides.
~ 25 - From U.S. Pat, 3t720,585 it is furthermore known, in ~ ~he case of two specific types of yeast, to perform a brief .. . .
1 .
.
}
thermal shock treatment consisting of heating at 60 to 70C
for two to twenty seconds, followed by about 20 minutes of heating at 45 to 50~C at pH values between 4.5 and 7. This process has the disadvantage that it liberates proteases 5 which cleave the useful (digestible) protein and conse-quently the protein yield is considerably reduced. The residual RNA content is reduced but is still greater than the preferred range of 2%.
T~E INVENTION
The invention is therefore addressed to the problem OL
creating a method for the production of single-cell protein, which will not have the above-described disadvantages of 5 the known methods. In particular it is an aim of the in-vention to create a simple process which can be performed on a large technical scale, requires no expensive apparatus, can be performed without the addition of foreign substances, requires no previous treatment for the killing and digestion f the microorganisms, and does not harm the environment.
This problem is solved in accordance with"the invention by a method of obtaining protein of reduced nucleic acid content from microorganisms by thermal treatment, which is 25 characterized in that an aqueous suspension of the micro-organisms, with the addition in some cases of a material having ribonuclease activity, is heated at a pH between 7 ;' :
' ' :
.
" ~
and 805 at a temperature of 63 to 67C, until the content of 5'-nucleotides in the medium has reached a certain value, then the medium is briefly boiled, and the insoluble crude protein is separated from the suspension.
In the performance of the process of the invention, the -~ nucleic acid is cleaved exclusively into 5'-nucleotides which appear in the medium, while no 2'- or 3'-nucleotides are formed. Depending on the nucleic acid content of the microorganism and the desired final content of nucleic acid, it is therefore easy to determine in advance at what 5'-nucleotide content in the medium the nucleic aci~ content will be reduced to the desired leve] and therefore the heat treatment at 63 to 67C can be terminated. It is decisively important that the precise temperature limits between 63 and 67C, and preferably a 65C temperature, be maintained, and that at the same time the pH value remain always in the stated range. Since the p~ decreases in the course of the thermal treatment due to the liberation of the nucleotides, the stated pH range is maintained by the addition of bases, preferably soda lye, potash lye or ammonia.
Endogenous nucleases are activated by the process of the invention and this results in the cleavage of khe nucleic acid with the formation of the 5'~nucleotides which are highly desirable as by-products: adenosine monophosphate (AMP), guanosine monophosphate (GMP), cytosine monophosphate ,'' '.
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.
.
.
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(CMP) and uridine monophosphate (UMP). The heating time required depends on the microorganism used, especially on its content of endogenous nucleases. Since the nuclease content can differ greatly, the heating time will also differ greatly and generally will range between about 20 minutes and about 20 hours.
In the course of the heat treatment of the invention, the protein is largely converted to the insol~uble state.
The precipi-tation of the protein is then completed by the brief boiling, i.e., by heating between about 90 and 100C
at normal pressure, or higher temperatures at elevated pres-sure. Then the insoluble crude protein is separated from the suspension, for example by centrifugation or filtration, 15 and can then be put out for use as a nutrient or can be further processed in any desired manner.
All kinds of microorganisms and protozoa are suitable for the process-of the invention, such as, for example, yeasts, bacteria, molds and the like. Examples of suitable microorganisms are Saccharomyces cerevisiae, Saccharomyces carlsbergensis and Candida boidinii as yeasts, ~ocardia erythropolis as a mycobacterium, Bacillus cereus and Bacil-lus subtilis as bacillaceae, Streptococcus faecalis as an example of streptococci, Pseudomonas fluorescens as an ex-ample of pseudomonads,~ and Aspergillus niger as an example ~f a mold. As previously mentioned, the microorganisms are ~777~Z
used preferably in freshly harvested, viable form, i.e., in the form in which it is harvested from the culture medium.
However, protozoa treated by known methods for the preserva-tion of living microorganisms can also be used, such as 5 especially dried or lyophilized protozoa.
' In a special embodiment of the process of the invention~
; it is possible to reduce the heating time by the addition of a material having a ribonuclease activity. This includes both purified ribonuclease preparations and semirefined or crude biological preparations which contain ribonuclease.
Preferred material having ribonuclease activity are germs of grasses and cereal species/ especially malt germ ~ or extracts thereof, preferably malt germ extract. In this - 15 embodiment, the process of the invention is applicable also . . .
- to ribonuclease-free microorganisms, such as for example the E. coli MRE 600 mutant.
.. :
As a rule, the process of the invention permits a re-duction of the ribonucleic acid content in single-cell protein to from 0 to 0.2% by weight. The protein losses are slight, and particularly the cleavage of protein and the loss of value due to racemization are negligible.
., .
:; 25 The protein yield in the method of the in~ention at-tains 90% and more. A special advantage of the method con-sists in the fact that the ribonucleic acid is cleaved vir- ;
:
, .
_g _ ,.
,~ . ' ~6~7777'~
tually quantitatively to the 5'-nucleotides, which them~
selves are valuable substances. It is therefore possible to prepare the nucleotides from the filtrate after the sep-aration of the crude protein. It is especially advantageous, 5 however, to reuse the fi]trate for the cultivation of micro-organisms. The microorganisms use the nucleotides to build the necessary nucleic acid again, so that in this manner a virtual recycling of the nucleotides is achieved. At the same time, the rest of the dissolved cell components, the remainder of the still soluble protein, and the like, are utilized by the microorganisms for the fresh growth. Con-sequently, the addition of biotin or yeast extract, which is otherwise common in the cultivation of microorganisms, can be wholly or largely dispensed witho By the method of the invention, a purer, more digest-ible protein is obtained, which is free of deter~ents, salts or RNA. On the other hand, in the formerly common methods ;of cell digestion with alkali or acids, some of the amino acids in the protein are destroyed or racemized.
A speclal effect of the method of the invention fur-thermore lies in the fact that the peculiar odor of the protein, which is especially pronounced in the case of the - 25 yeasts, is destroyed by it~ This, too, makes the single-cell protein of the invention more suitable for use as a ~ood.
~7777~
The removal of the RNA is performed in an especially simple and economical manner. The 5'-nucleotides can be recovered as by-products. Additional economies can be achieved in the culture media by recycling the filtrate into 5 the culture broths. The process is also environmentally harmless, since the cell waste can be burned without dif-ficulty, which is not the case with a waste material that has been treated with salt, acid or alkali.
;~ 15 ` 20 . I , ', :
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EXAMPLES
The following examples will further explain the in-vention.
Examples 1 7 200 liters of an aqueous suspension of each of the microorganisms listed in Table 1~ with a dry mass content of about 15 to 20%, is heated at 65C and maintained at this temperature for 15 hours with gentle stirring. The pH is maintained constant between 7 and 8.5 by the addition of soda lye. Then the mixture is briefly boiled and the crude protein is separated by centrifugation and dried. The yield 5 is between 25 and 32 kg of crude protein. Table 1 shows the RNA content of the starting material and the RNA cleavage produced in it.
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ut ; J u~ X ~1 ~ ,a o ,~
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As Table 1 shows, the process of the invention pro-duces very good yields of 5'-nucleotides from fresh micro-organisms. When the microorganisms have been standing for a comparatively long time, the yield is lower, and this is 5 attributed to the reduced ribonuclease content.
Examples 8 to 14 -a) 3 kg of malt germs are ground; 30 1 of water is added, and the mixture is held for 40 minutes at 65C. The suspension obtained is used either directly or after removal of the insoluble malt germ fragments in the following process.
' :
5 b) 200 liters of a suspension of each of the same micro-organisms as in Examples 1 to 7, with the same dry mass content as described therein, are heated to about 65C, the malt germ suspension obtained in a) or the extract thereof is added, and then the mixture is heated for between 2 and 15 hours at 65C. The proces-sing is then performed as described in Examples 1 to 7. The yield is between 25 and 35 kg of crude protein, and the average pure protein content is 70O.
~ .
Table 2 shows the RNA cleavage obtained. The yield of the 5'-nucleotides~amounts in each case, for an RNA con-tent in the dry substance o~ about 10o ~ to about 700 to 800 77'772 grams of each of the four different 5'-nucleotides; where ~ the RNA content is lower, the yield is lower according to -~ the starting material.
Example 15 As described in the preceding examples, a suspension of 100 g of dry Saccharomyces cerevisiae with a content of ~' about 45% protein and 1.5% to 3% RNA in the dry substance, was heated at 65C without the addition of any material hav-ing a ribonuclease activity, and was boiled and the crude protein was separated. The composition of the filtrate and of the crude protein was determined and is given in Table ,; 3 below. The values stated show that the protein content ' 15 in the crude protein ,was enriched from 45% to 56%.
I In the case of microorganisms of hlgher initial protein ,', content, the yield of crude protein can increase up to 85%.
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It will be understood that the specification and examples are illustrative but not limitative of the present invention and that other embodiments within the spirit and scope of the invention will suggest themselves to those skilled in the art.
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In spite of these difficulties, microorganisms are already being cultivated on a large scale for the feeding of domestic animals (fodder yeast). For human nutrition, however, this protein from microorganisms has hitherto been used either not at all or only to a limited extent. The high ribonucleic acid content of this protein, for example, constitutes a serious problem. ~ibonucleic acld (~A) is decomposed in the mammalian organism to uric acid, which in turn can cause gout. The ~NA content of microorganlsm protein must therefore be less than 2%, and preferably less than 0.5%.
One of the most important aims, therefore, is to pro-duce the microbial protein free of ribonucleic acid. This has not been accomplished in a satisfactory manner to date.
Attempts have been made to breed microorganisms of an espe-cially low RNA content. These microorganisms have the dis-!
, advantage, however, that they grow considerably more slowly.
It is known that microorganisms of high RNA content grow more rapidly than those of low RNA content. This results in areal dilemma: the desired fast growth of the microorganisms is to be equated with a high RMA content. If slower produc-tion can be accepted, microorganisms of low RNA content can be cultivated.
The second possibility for eliminating RNA from micro-rganism protein, which is frequently referred to as single-1~t7777Z
cell protein ~SCP)), is based upon the extraction of theRNA followed by separation of the insoluble protein. Sev-eral possibilities for such extraction of RNA have been described as follows:
5 a) Alkaline extraction - German Offenlegungsschrift 16 70 110.3 b) Acid extraction i c) Treatment with high salt concentration (NaCl, KCl, NH4Cl) ~ , d) Treatment with detergents (dodecyl sulfate etc.).
All of these methods have the disadvantage that the microorganisms in some cases have to be processed in a very complicated manner, and the protein is partially hydrolyzed, racemized or otherwise altered by the treatment and by the various additives and thus becomes less suitable if not use-less for nutritive purposes. For example, alkaline extrac-tion modifies the protein in its structure such that it forms colloidal solutions. Separatlon is possible only after neutralization and accordingly necessitates the re-moval of the neutral salts that are formed. Furthermore, the dissolved RNA becomes partially denatured, and can no longer be quantitatively cleaved with enzymes to form soluble mono-nucleotides.
~ 25 - From U.S. Pat, 3t720,585 it is furthermore known, in ~ ~he case of two specific types of yeast, to perform a brief .. . .
1 .
.
}
thermal shock treatment consisting of heating at 60 to 70C
for two to twenty seconds, followed by about 20 minutes of heating at 45 to 50~C at pH values between 4.5 and 7. This process has the disadvantage that it liberates proteases 5 which cleave the useful (digestible) protein and conse-quently the protein yield is considerably reduced. The residual RNA content is reduced but is still greater than the preferred range of 2%.
T~E INVENTION
The invention is therefore addressed to the problem OL
creating a method for the production of single-cell protein, which will not have the above-described disadvantages of 5 the known methods. In particular it is an aim of the in-vention to create a simple process which can be performed on a large technical scale, requires no expensive apparatus, can be performed without the addition of foreign substances, requires no previous treatment for the killing and digestion f the microorganisms, and does not harm the environment.
This problem is solved in accordance with"the invention by a method of obtaining protein of reduced nucleic acid content from microorganisms by thermal treatment, which is 25 characterized in that an aqueous suspension of the micro-organisms, with the addition in some cases of a material having ribonuclease activity, is heated at a pH between 7 ;' :
' ' :
.
" ~
and 805 at a temperature of 63 to 67C, until the content of 5'-nucleotides in the medium has reached a certain value, then the medium is briefly boiled, and the insoluble crude protein is separated from the suspension.
In the performance of the process of the invention, the -~ nucleic acid is cleaved exclusively into 5'-nucleotides which appear in the medium, while no 2'- or 3'-nucleotides are formed. Depending on the nucleic acid content of the microorganism and the desired final content of nucleic acid, it is therefore easy to determine in advance at what 5'-nucleotide content in the medium the nucleic aci~ content will be reduced to the desired leve] and therefore the heat treatment at 63 to 67C can be terminated. It is decisively important that the precise temperature limits between 63 and 67C, and preferably a 65C temperature, be maintained, and that at the same time the pH value remain always in the stated range. Since the p~ decreases in the course of the thermal treatment due to the liberation of the nucleotides, the stated pH range is maintained by the addition of bases, preferably soda lye, potash lye or ammonia.
Endogenous nucleases are activated by the process of the invention and this results in the cleavage of khe nucleic acid with the formation of the 5'~nucleotides which are highly desirable as by-products: adenosine monophosphate (AMP), guanosine monophosphate (GMP), cytosine monophosphate ,'' '.
;~ -7-., ~ .
.
.
.
~77q~
(CMP) and uridine monophosphate (UMP). The heating time required depends on the microorganism used, especially on its content of endogenous nucleases. Since the nuclease content can differ greatly, the heating time will also differ greatly and generally will range between about 20 minutes and about 20 hours.
In the course of the heat treatment of the invention, the protein is largely converted to the insol~uble state.
The precipi-tation of the protein is then completed by the brief boiling, i.e., by heating between about 90 and 100C
at normal pressure, or higher temperatures at elevated pres-sure. Then the insoluble crude protein is separated from the suspension, for example by centrifugation or filtration, 15 and can then be put out for use as a nutrient or can be further processed in any desired manner.
All kinds of microorganisms and protozoa are suitable for the process-of the invention, such as, for example, yeasts, bacteria, molds and the like. Examples of suitable microorganisms are Saccharomyces cerevisiae, Saccharomyces carlsbergensis and Candida boidinii as yeasts, ~ocardia erythropolis as a mycobacterium, Bacillus cereus and Bacil-lus subtilis as bacillaceae, Streptococcus faecalis as an example of streptococci, Pseudomonas fluorescens as an ex-ample of pseudomonads,~ and Aspergillus niger as an example ~f a mold. As previously mentioned, the microorganisms are ~777~Z
used preferably in freshly harvested, viable form, i.e., in the form in which it is harvested from the culture medium.
However, protozoa treated by known methods for the preserva-tion of living microorganisms can also be used, such as 5 especially dried or lyophilized protozoa.
' In a special embodiment of the process of the invention~
; it is possible to reduce the heating time by the addition of a material having a ribonuclease activity. This includes both purified ribonuclease preparations and semirefined or crude biological preparations which contain ribonuclease.
Preferred material having ribonuclease activity are germs of grasses and cereal species/ especially malt germ ~ or extracts thereof, preferably malt germ extract. In this - 15 embodiment, the process of the invention is applicable also . . .
- to ribonuclease-free microorganisms, such as for example the E. coli MRE 600 mutant.
.. :
As a rule, the process of the invention permits a re-duction of the ribonucleic acid content in single-cell protein to from 0 to 0.2% by weight. The protein losses are slight, and particularly the cleavage of protein and the loss of value due to racemization are negligible.
., .
:; 25 The protein yield in the method of the in~ention at-tains 90% and more. A special advantage of the method con-sists in the fact that the ribonucleic acid is cleaved vir- ;
:
, .
_g _ ,.
,~ . ' ~6~7777'~
tually quantitatively to the 5'-nucleotides, which them~
selves are valuable substances. It is therefore possible to prepare the nucleotides from the filtrate after the sep-aration of the crude protein. It is especially advantageous, 5 however, to reuse the fi]trate for the cultivation of micro-organisms. The microorganisms use the nucleotides to build the necessary nucleic acid again, so that in this manner a virtual recycling of the nucleotides is achieved. At the same time, the rest of the dissolved cell components, the remainder of the still soluble protein, and the like, are utilized by the microorganisms for the fresh growth. Con-sequently, the addition of biotin or yeast extract, which is otherwise common in the cultivation of microorganisms, can be wholly or largely dispensed witho By the method of the invention, a purer, more digest-ible protein is obtained, which is free of deter~ents, salts or RNA. On the other hand, in the formerly common methods ;of cell digestion with alkali or acids, some of the amino acids in the protein are destroyed or racemized.
A speclal effect of the method of the invention fur-thermore lies in the fact that the peculiar odor of the protein, which is especially pronounced in the case of the - 25 yeasts, is destroyed by it~ This, too, makes the single-cell protein of the invention more suitable for use as a ~ood.
~7777~
The removal of the RNA is performed in an especially simple and economical manner. The 5'-nucleotides can be recovered as by-products. Additional economies can be achieved in the culture media by recycling the filtrate into 5 the culture broths. The process is also environmentally harmless, since the cell waste can be burned without dif-ficulty, which is not the case with a waste material that has been treated with salt, acid or alkali.
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EXAMPLES
The following examples will further explain the in-vention.
Examples 1 7 200 liters of an aqueous suspension of each of the microorganisms listed in Table 1~ with a dry mass content of about 15 to 20%, is heated at 65C and maintained at this temperature for 15 hours with gentle stirring. The pH is maintained constant between 7 and 8.5 by the addition of soda lye. Then the mixture is briefly boiled and the crude protein is separated by centrifugation and dried. The yield 5 is between 25 and 32 kg of crude protein. Table 1 shows the RNA content of the starting material and the RNA cleavage produced in it.
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As Table 1 shows, the process of the invention pro-duces very good yields of 5'-nucleotides from fresh micro-organisms. When the microorganisms have been standing for a comparatively long time, the yield is lower, and this is 5 attributed to the reduced ribonuclease content.
Examples 8 to 14 -a) 3 kg of malt germs are ground; 30 1 of water is added, and the mixture is held for 40 minutes at 65C. The suspension obtained is used either directly or after removal of the insoluble malt germ fragments in the following process.
' :
5 b) 200 liters of a suspension of each of the same micro-organisms as in Examples 1 to 7, with the same dry mass content as described therein, are heated to about 65C, the malt germ suspension obtained in a) or the extract thereof is added, and then the mixture is heated for between 2 and 15 hours at 65C. The proces-sing is then performed as described in Examples 1 to 7. The yield is between 25 and 35 kg of crude protein, and the average pure protein content is 70O.
~ .
Table 2 shows the RNA cleavage obtained. The yield of the 5'-nucleotides~amounts in each case, for an RNA con-tent in the dry substance o~ about 10o ~ to about 700 to 800 77'772 grams of each of the four different 5'-nucleotides; where ~ the RNA content is lower, the yield is lower according to -~ the starting material.
Example 15 As described in the preceding examples, a suspension of 100 g of dry Saccharomyces cerevisiae with a content of ~' about 45% protein and 1.5% to 3% RNA in the dry substance, was heated at 65C without the addition of any material hav-ing a ribonuclease activity, and was boiled and the crude protein was separated. The composition of the filtrate and of the crude protein was determined and is given in Table ,; 3 below. The values stated show that the protein content ' 15 in the crude protein ,was enriched from 45% to 56%.
I In the case of microorganisms of hlgher initial protein ,', content, the yield of crude protein can increase up to 85%.
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It will be understood that the specification and examples are illustrative but not limitative of the present invention and that other embodiments within the spirit and scope of the invention will suggest themselves to those skilled in the art.
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Claims (7)
1. Method of obtaining protein having a reduced nucleic acid content from microorganisms which process comprises forming an aqueous suspension of the microorganisms, heating same at a temperature of from 63 to 67°C and at a pH of from 7 to 8.5 until the 5'-nucleotide content in the suspension has reached the desired level, briefly boiling the resulting mixture, and separating the insoluble crude protein from the suspension.
2. Method as claimed in claim 1, wherein a material having ribonuclease activity is added prior to heating the suspension.
3. Method as claimed in claim 1, wherein the pH is maintained in the range of 7 to 8.5 by the addition of an aqueous base.
4. Method as claimed in claim 2, wherein said aqueous base is soda lye, potash lye or ammonia.
5. Method as claimed in claim 2, wherein said material having ribonuclease activity is cereal germs or an extract thereof.
6. Method as claimed in claim 1, wherein the solution remaining after said insoluble crude protein has been separated from the suspension is returned to the starting aqueous suspension of microorganisms.
7. Method as claimed in claim 1, wherein the solution remaining after said insoluble crude protein has been separated from the suspension is utilized for obtaining 5'-nucleotides.
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| DE2622982A DE2622982B1 (en) | 1976-05-21 | 1976-05-21 | Process for the production of protein with a low nucleic acid content from microorganisms |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CA1077772A true CA1077772A (en) | 1980-05-20 |
Family
ID=5978741
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA277,828A Expired CA1077772A (en) | 1976-05-21 | 1977-05-06 | Method of recovering protein of low nucleic acid content from microorganisms |
Country Status (6)
| Country | Link |
|---|---|
| JP (1) | JPS52143290A (en) |
| CA (1) | CA1077772A (en) |
| FI (1) | FI58156C (en) |
| SE (1) | SE433501B (en) |
| SU (1) | SU884574A3 (en) |
| YU (1) | YU125877A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4999193A (en) * | 1987-06-16 | 1991-03-12 | Etablissements Guyomarc'h S.A. | Feed additive for animals, feeds containing such an additive and method for improving the growth of animals |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| MX352973B (en) * | 2011-09-22 | 2017-12-15 | Danisco Us Inc | Endogenous dnase activity to reduce dna content. |
Family Cites Families (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| GB1440642A (en) * | 1973-09-24 | 1976-06-23 | Ranks Hovis Mcdougall Ltd | Production of edible protein containing substances |
| GB1466078A (en) * | 1973-09-26 | 1977-03-02 | British Petroleum Co | Treatment of proteinaceous material |
| SE7401668L (en) * | 1974-02-07 | 1975-08-08 | Scp Exploatering Ab |
-
1977
- 1977-05-06 CA CA277,828A patent/CA1077772A/en not_active Expired
- 1977-05-16 SE SE7705701A patent/SE433501B/en not_active IP Right Cessation
- 1977-05-19 YU YU125877A patent/YU125877A/en unknown
- 1977-05-20 FI FI771621A patent/FI58156C/en not_active IP Right Cessation
- 1977-05-20 JP JP5862677A patent/JPS52143290A/en active Granted
- 1977-05-20 SU SU772485597A patent/SU884574A3/en active
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4999193A (en) * | 1987-06-16 | 1991-03-12 | Etablissements Guyomarc'h S.A. | Feed additive for animals, feeds containing such an additive and method for improving the growth of animals |
Also Published As
| Publication number | Publication date |
|---|---|
| SU884574A3 (en) | 1981-11-23 |
| FI771621A (en) | 1977-11-22 |
| FI58156B (en) | 1980-08-29 |
| JPS5637799B2 (en) | 1981-09-02 |
| FI58156C (en) | 1980-12-10 |
| SE433501B (en) | 1984-05-28 |
| SE7705701L (en) | 1977-11-22 |
| JPS52143290A (en) | 1977-11-29 |
| YU125877A (en) | 1982-10-31 |
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