DE2059864C3 - Process for the preservation of beer - Google Patents

Description

where η 2, R is an alkyl group with 5.6 or 7 carbon atoms and M is a hydrogen atom, an alkali metal or ¹ I ₂ MoI of an alkaline earth metal and in which two MO groups are attached to the benzene nucleus in 2,4 or 3,4 positions are bound to the group COOR, in an amount of 5 to 60 ppm.

The invention relates to a method for preserving beer. In particular, the invention relates to a Process for preserving beer, which consists in adding a dihydroxybenzoic acid ester to the beer to stabilize it against decomposition and putrefaction during storage.

Preservatives that are generally used with liquid Foods used are benzoic acid, salicylic acid, dehydracetic acid salts, p-hydroxybenzoic acid and salts of sorbic acid.

With these preservatives it sometimes happens that an adequate preservative effect is not achieved if they are added in the amounts permitted by the Food Laws are. Furthermore, their effects depend on the types of foods to which they are added.

In the case of beer, for example, heptyl p-hydroxybenzoate is used in an amount of 10 to 12 ppm and this amount is effective against most putrefactive microorganisms such as yeast and lactobacilli. Since heptyl p-hydroxybenzoate only beschraiikt in beer soluble falls esmanchmal at temperatures of about 0 ⁰ C, and the beer is cloudy.

In the essay by M ο r r i s in »J. At the. Oil Chem. Soc. "November 1949, p. 639, it is stated that Polyhydroxybenzoic acid or its higher alkyl esters, Acyl derivatives or dialkyl-substituted polyhydroxybenzoic acids are effective as antioxidants. However, this reference is by no means indicative on the antimicrobial properties of the compounds to be used according to the invention for preservation from beer.

It has now been found that aromatic carboxylic acid esters which have two hydroxyl groups attached to a benzene nucleus, have an excellent antibacterial spectrum and a very low one Have toxicity as precursors of these compounds occur in nature and in general for the are harmless to the human body.

It has also been found that these compounds are excellently soluble in beer and that they are a have excellent storage stability.

The present invention is a method for preserving beer, thereby is characterized in that it is given a compound of the formula

(MO) _n

where η 2, R is an alkyl group having 5, 6 or 7 carbon atoms and M is a hydrogen atom, an alkali metal or V is ₂ moles of an alkaline earth metal and where two MO groups on the benzene nucleus in 2,4- or 3,4-positions to the Group COOR are bound, adds in an amount of 5 to 6 ppm. In general formula I, the alkyl groups expressed by R may be either straight chain or branched, but in general it is preferable that R is a straight chain alkyl group. Compounds of Formula I which can be used in the present invention are listed in Table I.

Table 1

Compound structural formula No

Solubility (; ■ ml) water

I 5% aqueous solution of ethanol

Melting point (C). Boiling point I C)

OH

HO

COOC ₅ H ₁₁

HO- <r V- COOCH

3.88 ·

1.37

5.82 8.62 10

4.21 · 10

2.50 10

[145 / 0.08 mm Hg]

[147 / 0.1 mm Hg]

[156 / 0.1 mm Hg]

continuation

Compound structural formula

Solubility (; '/ ml) water

15% aqueous solution of ethanol

Melting point (° C), boiling point ( ⁰ C)

OH

4 HO ^ C ^ COOC ₅ H ₁₁ 2.92 ΙΟ ² - 7.80 · ² 93.5-94.5

OH

HO - <^^ - COOQH ₁₀ 1.81-1O ²

OH

6 HO— <f ^X > -COOC ₇ H ₁₅ 7.20 ·

5.43 · 10 ²

1.45- W ²

117

84-86

In the present invention, it is critical that the compound expressed by the formula I has an alkyl group R containing 5.6 or 7 carbon atoms.

The dihydroxybenzoic acid esters according to the invention can be used in the form of a salt if the hydrogen atom of one or more hydroxyl groups through an alkali metal such as sodium or potassium, or replaced by an alkaline earth metal such as magnesium and calcium. When the invention Esters are used as salts, their solubility in water is further improved.

All the compounds of the formula 1 which are used according to the invention are readily soluble in beer. Furthermore, the compounds according to the invention have a structure which is similar to the structure of p-hydroxybenzoic acid ester is. The use of p-hydroxybenzoic acid ester is approved by the World Health Organization (WHO) and del Food and Agriculture Organization (FAO) considered safe and harmless, and it is believed that the internal metabolism of the compounds of the invention is similar the metabolism of p-hydroxybenzoic acid ester. Other compounds which the invention Compounds that are analogous, such as gallic acid, protocatechic acid, and phloroglucinic acid, are in widespread in nature, and their safety is very high.

Since the above compounds generally have high antibacterial activity, sufficient ones can be obtained Effect obtained when added to beer in amounts of 5 to 60 ppm.

In the practice of the present invention, the compounds of Formula 1 can be used at any stage can be added during the production of the beer. They can be in the form of an aqueous solution in alkali hydroxides, such as sodium hydroxide and potassium hydroxide can be added. You can also be inflicted are converted into the corresponding alkali metal salts after one or two of their hydroxyl groups became. Furthermore, they can be in the form of the phosphoric acid esters or glycine ethers or in the form a solution in ethanol, propylene glycol, or other edible harmless solvents such as glycerin and water. For example, they can be in a mixed solvent of propylene glycol and sodium hydroxide can be used.

The antibacterial activity of some compounds of formula 1 used according to the invention is given in Table 11. The numerical Values given in the table are values of the minimum growth inhibition concentration, obtained in antibacterial tests, the tests being carried out at 30 "C for 4 days in a potato dextrose medium or in an agar medium and where one used various bacteria, yeasts and fungi.

Table II

Bacterium Compound (y / ml)
1 4 200 Staphylococcus aureus 209 P. 25th 100 Bacillus subtilis PCI 219 25th 200 Salmonella enteritidis 1891 > 100 400 Escherichia coli 0-55 > 100 100 Proteus vulgaris HX 19 > 100 250 Aspergillus niger IAM 3001 > 250 Rhizopus nigricans IAM 6070 125 Saccharomyces cerevisiae 62.5 125 Hansenula anomala > 250 125 Candida utilis 125

In Table II the numbers correspond to the compounds the numbers in Table I. These numbers are also used in the following tables.

The preservative effects and the antibacterial activities of the compounds of the present invention depend on the position of the hydroxyl groups and the number of carbon atoms in the alkyl group away.

Soft drinks are classified into types that Contain carbonic acid, and those that do not contain carbonic acid. It is allowed. Benzoic acid (600 ml). Dehydracetic acid (50; - ml) and a p-hydroxybenzoic acid ester (100 7 ml) as preservatives

to be used for beer, but even if these substances are added in permissible concentrations, one can often observe the formation of turbidity caused by yeast. The compounds of the invention prevent such decomposition from occurring at the concentration of 20 to 20 γ / ζα \.

For the preservation of beer it is desirable that the alkyl group R in the compounds of the general formula I has a length which preferably corresponds to 6 to 7 carbon atoms. Using these connections, not pasteurized beer can preserve without the occurrence of any turbidity at 0 ⁰ C, which hydroxybenzoate heptyl p in the case of is not possible. By using the compounds described above, the lowering of the foam-retaining properties of beer, as is observed in the case of p-hydroxybenzoate, can be greatly reduced.

Results of the antibacterial tests of the aforementioned species against common micro-organisms, which were carried out by using a potato dextrose (in the case of yeasts and Molds) or an agar medium (in the case of common bacteria) used are shown in Table III specified. The numerical values in the table are values of the minimum growth inhibition ! Concentration (y / ml) if you stay for 4 days cultivated at 30C.

Table III

Bacteria species

prefers the connections just before the biei bottled is to add.

The exploitation of the invention can be made possible by legal provisions, in particular by the food law, be limited.

The following examples illustrate the invention without, however, restricting it.

example 1

Saccharomyces cerevisiae, Saccharomyces ubarurn and Lactobacillus pastorianus, known as putrefactive bacteria for beer, were grown separately in a 3% malt extract broth containing 4% ethanol. Using these broths, the antibacterial activities of Compounds 6 and 5 were examined. A drop of the liquid in which three strains had been cultured for 24 hours was added to a medium containing the preservative compound through an injector, and then incubated at 30 ° C. for 70 days, and the growth of the bacterial samples was examined. The minimum growth inhibition concentration (y / m \) was determined for each compound. For comparison purposes, η - heptyl - ρ - hydroxybenzoate (POBrI) was tested in a similar manner. The results are given in Table V.

Saccharomyces cerevisiae
Saccharomyces rouxii
Aspergillus niger
Staphylococcus aureus
Bacillus subtilis
Proteus vulgaris

30th Table V Minimal growth inhibition
kon / entralion (; · ml) Saccharo
imccs
»Cash for Lactobaullu
pasloriamis Suixharo
niNccs
cerevisiae 35 Conservation stuff 9 9 Connection 6 9 12th
8th 10
6th 15.6 12th
8th <3.91 40 Connection 6 62.5
50
50 Connection 5
POBH 50

The main antibacterial activities of other compounds are given in Table IV.

45

Table IV

Compound no. Yeast

<7.81-62.5

7.81-125

Lactic Acid Bacilli

250
<5

55

The harmlessness and safety of these compounds is characterized in that they have almost all LD ₅₀ values of 3 to 6 g when administered orally to mice, and the compounds are therefore characterized by very low toxicities.

Sufficient amounts are used to preserve beer Effects regardless of the time when the compounds of formula I during the beer production process be admitted. But it is

Example 2

The preservative effect of compound 6 on beer was investigated. 1 ml of a solution of compound 6 in ethanol was added to 349 ml of unpasteurized beer and then the sample was stored in bottles at room temperature. As a comparison, samples containing POBH incorporated in a similar manner and a beer sample which had been pasteurized by heating were examined at the same time. After storage at room temperature, part of the beer sample was kept at 0 ^° C. for 2 weeks, and then the solubility and stability of the preservation compound at OC was investigated. The results are given in Table VI, with the results obtained on storage at room temperature being shown in the “Storage” column, and the results of the stability test at 0 ° C. are shown in the “Stability” column. In the table, the numeric values indicate the number of months. during which the sample was stored without mold formation. and the symbol "+" means that appearance of white turbidity or precipitation of crystals was observed.

Table VI

concentration
I mil

VLT hi ml u π ü d
l.iiucMini! stability

ΙΌ B H

I-iijici'Uiij! stability

Investigations are given in Table VI, where the "*" sign shows a 5% difference.

5 Table VII

Untreated comparative test (1)

8 4

10> 6

12> 6

14> 6

IS

It was found that the samples containing the compound 6 with concen- trations of 10; ■ m! or more incorporated contained were excellently preserved, in a manner similar to the comparative samples obtained by Heating was pasteurized. No OC was found in the samples containing compound 6 Appearance of white turbidity or precipitated crystals, at a concentration of 8; ■ ml. During white turbidity appeared in the POBH sample at a concentration of 8; · ml. It was even at a concentration of 14; -ml no turbidity observed, and from this it can be seen that the connection 6 is particularly excellent in terms of its solubility wedge and its stability.

Example 3

In this example, the influence of compound 6 examined for the taste of beer. Sample A was made by going to unpasteurized Beer added an ethanol solution of compound 6 (the ethanol content was approximately 0.3%. Calculated on the beer), whereby a bicr solution was formed, which the compound with a concentration of 10 ml. Sample B was made by added the same amount of ethanol to unpasteurized beer as above (the ethanol content was about 0.3%). Sample C was made by ethanol was added in the same amount as above to beer which had been heat pasteurized.

A group of 10 experts was asked for the taste test and pair studies of Samples A and B and Samples A and C were selected repeated twice. The temperature of the beer at the time of drinking was 6 to 9 ° C. The results of the Connection (>

Number of number of Λη / ahl of Examiners, the Λ examiners, the B examiners, the C

essential essential

better than A

judged

than much better
as B or C
judged

rated better than Λ

comparison of
Samples A and B
comparison of
Samples A and C

15 *

From the results shown in Table VII it can be seen that the taste of the samples containing the Compound 6 contained, was not spoiled in comparison with unpasteurized beer and that it Much better rated than beer that went through Heating had been pasteurized.

The determination compared to the state of the Technology, as described by the journal »Brauwissenschaft« from July 1969. p. 310. and by the OE-PS No. 2 70 543 is shown, technical progress surprisingly achieved is evident from the test reports below I and 11.

Test report 1

1. Comparative experiment

Comparison of the growth inhibition concentration in relation to molds in common culture medium

A culture medium with 1% peplon, 4.0% glucose was used. 1.0% yeast extract. 2.0% agar-agar, with a pH of 5.6. Molds grown at 30 C for 72 hours in the same culture medium were inoculated with platinum loops.

The investigated compounds were added to the culture medium as a dimethyl sulfoxide solution Its solidification was added in such a way that the content of the solution in the culture medium was 1%. It was grown for 48 hours.

Octyl gallate

POBH *)

\ spergillus niger IAM > 40 > 20 50 50 62.5 \ spergillus oryzae var. microsporus > 40 > 20 50 50 > 62.5 Penicillium citrinum IAM 7316 40 50 200 31.25 saccharomyces cerevisiae 40 20th 25th <12.5 15th saccharomyces roudii 20 40 K) 12.5 <12.5 <4 Hansenula anomala > 40 > 20 25th <1Z5 25th Zandida utilis 40 20th <1Z5 25th * l POBH n-HepHl-p-hvdro \> henz <ul 609 632/113

2. Verification attempt

Growth Inhibiting Concentrations

in relation to yeast
in the culture medium containing 4% alcohol. The results of storage for a period of 1 to 14 days are shown in the table below.

Influence of the compounds on the turbidity of the beer, which is low at 0 C

¹⁰ connections

3% alcohol was added to the culture medium containing 25% malt extract, followed by a alcoholic solution of the compound to be investigated was added in such a way that the alcohol concentration in the culture medium was exactly 1% more.

Those grown for the experiment at 30 C for 72 hours in the same culture medium with 4% alcohol Yeasts were inoculated and it was made at a 15 octyl gallate Maintain a temperature of 30 ° C for 72 hours. The growth inhibition concentration was determined by the existing growth.

connection

Microorganisms

S. ubarum S. cerevisiae S. ι

7016 7030 70ί

I; Mil

{; ■ ml)

ml)

Octyl gallate 12th 10 16 POBH 10 8th 10 I. 18th 15th 15th 2 10 8th 8th 3 6th 6th 6th 6th 10 8th 10 Empty attempt I. 4-

As can be seen in the table. are all compounds in the malt culture medium containing 4% alcohol, except connection 1, against the An. . Heaven more effective than octyl gallate. In the beer, which one creates poorer conditions for the growth of microorganisms, lower growth inhibition concentrations can to be expected.

POBH

35

40 concentration

I-ml!

5 10

15 20

10 15 20

10 15 20

5 10 15 20

5 10 15 20

aue iler I..isiciΊΐιιμ

4-

45 3rd comparison experiment

Influence on the cloudiness of beer

Pure Japanese draft beer (Santory Junnama) was used. The beer was before its use stored in a refrigerator at OC for 2 days after which it was subjected to tests.

All compounds were in aqueous 0.1 N NaOH solutions dissolved to form 6330 ppm solutions. These solutions were in 633 ml beer bottles introduced in amounts of 2.1.5, 1 and 0.5 ml, whereupon 0.1 N NaOH solution in amounts of 0.5. 1.0 and 1.5 ml were added; through this became the NaOH content. kept constant in the samples. In a blank experiment, 2.0 ml became 0.1 normal NaOH filled in alone.

The chilled beer was quietly filled into the above bottles under nitrogen pressure until the predetermined one Volume of 633 ml was reached, and after closing it was again at 0 C kept. The cloudiness of the beer was observed with the naked eye

Empty attempt 10 15 20

- means that a cloudiness is not noticeable at all

* means that a slight cloudiness is noticeable.

+ means that the cloudiness is noticeable.

■ Ο- means that the cloudiness is clearly visible.

The table shows that beer preserved with octyl gallate at its effective concentration (5 to 10 r / ml) at 0 ^° C is cloudy due to the octyl gallate, in accordance with the journal »Brauwissenschaft« of July 1969, p. 310

On the other hand, no turbidity occurs in any of the compounds according to the invention that are examined at their effective concentration (1 = 15 to 20 μl / inl, = 8 to 10 μl / ml, 3 = 6 μl ml, cf. experiment 2 above). It follows from this that the compounds to be used in the process according to the invention are superior to octylene alate as a preservative for beer.

Vcrsuchsbcrichl 11 Experiment 1

Minimal growth inhibition concentration

with respect to yeast in the culture medium containing 4% alcohol

Culture medium

3% alcohol was added to the culture medium containing 2.5% malt extract, followed by a alcoholic solution of the compound to be examined in an amount of 1% (based on the alcohol) Concentration in the culture medium was added, $ o that the alcohol concentration in the resulting Culture medium was 4%.

Below ^r looked for yeasts

The cultured for the experiment at 30 C for 72 hours in the same culture medium with 4% alcohol yeasts were seeded, and it was maintained at a temperature of 30 ⁰ C for 72 hours. The minimum growth inhibition concentration was determined by the existing growth. The results are shown in the table below.

connection

M ik roorganisms

S. ubariim S. cerevi-7016 siac

7030

Heptyl p-hydroxy

benzoate

Oclyl gallate

Ι; · / ηιΙ)

10

S. cerevi

siae

7089

10

16

Connection

Microorganisms S. uharum S. cerevi

7016

10
6th

20th 10 10

siae 7030

(; ·; ΗιΙ)

15th

20 12

1 (amyl ester)

2 (hexyl ester)

3 (heptyl ester)

4 (amyl ester)

5 (hexyl ester)
f> (heptyl ester)

links

(Ester of 2,4-dihydroxybenzoic acid) Compound 1 (amyl ester)

Compound 2 (hexyl ester)

Compound 3 (heptyl ester)

S. cerc \ isiae

7OS9

15 8 6

20 10 10

The numbers of the connections correspond to those in the description.

Attempt 2

Influence of the compounds on the turbidity
of beer stored at 0 ° C

Pure Japanese draft beer (Santory Junnama) was used. Before the test, the beer was ^{stored in a refrigerator at 0 °} C. for 2 days, after which it was subjected to the tests.

All compounds were in aqueous 0.1 N NaOH solutions dissolved to form 6330 ppm solutions. These solutions were in 633 ml beer bottles introduced in amounts of 2, 1.5, 1 and 0.5 ml, whereupon 0.1 N NaOH solution in amounts of 0, 0.5, 1.0 and 1.5 ml was added, reducing the NaOH content was kept constant in the samples. In a blank experiment, 2.0 ml of 0.1 N NaOH solution were used filled in alone.

The cooled beer was calmly decanted into the sample bottles up to the predetermined volume of 633 ml, and after sealing it was again stored at 0 ° C. Cloudiness of the beer became with the detected by the naked eye.

The results when stored for a period of 1 to 14 days are as follows Table.

concentration

Days of storage

25 *) 20 15 10 20 15! 0 5

15 10

Wax! Umsinhibierungskonzenlration

15-18

8-10

- means that a haze is not noticeable ± means that erne slight cloudiness is noticeable.

+ means that the cloudiness is noticeable. + + means that the cloudiness is clearly visible ':'. '.': represents the usable range *) Indicates an astringent taste

13th

continuation
links

Concentrate inn

Days of storage

14th

14th

Grow u mslπhιhicrιιngskon / enlralion

(; Mil

(Ester of 3,4-dihydroxybenzoic acid)
Compound 4 (amyl ester)

Compound 5 (hexyl ester)

Compound 6 (heptyl ester)

Heptyl p-hydroxybenzoate

50 *) 40 20 IO 20 15 K) 5

20 15 K)

20 15 10

4- 4-

20th

10

N K)

X 10

Octvlaallat

comparison

20th

15 10

- means that a cloudiness is not noticeable at all. i means that a little haze is noticeable. + means that the cloudiness is noticeable.
4- means that the cloudiness is clearly visible.
. . : represents the usable area.
* l Indicates an astrinous taste.

IO 16

The technical progress of compounds 3 and 6 can also be found in connection with the test report I.

Claims (1)

Claim: Process for the preservation of beer, characterized in that it is a compound of the formula (MO) ₁ COOR