FR2639514A1 - Compositions for coating food additives intended for ruminants. - Google Patents

Abstract

Compositions for coating food additives intended for ruminants which are stable to a pH greater than 5 and which liberate the biologically active substance at a pH less than 3.5, containing as substance sensitive to pH variations an organosoluble chitosan derivative consisting of the random linking of the units represented by the formulae: in which: R1 represents an alkanoyl radical(2 to 4 carbon atoms), R2 represents an optionally substituted alkylidene (3 to 14 carbon atoms) or benzylidene radical, R3 represents a hydrogen atom or an acetyl or alkanoyl (3 or 4 carbon atoms) radical, a portion of the radicals R4 represents a hydrogen atom and the other represents an alkanoyl radical identical to R1, whose mean molecular mass is between 10,000 and 80,000 and which contains 60 to 65% of amine functional groups in imine form and 3 to 35-40% of amine functional groups in acetylamino form.

Description

 The present invention relates to compositions for coating food additives or biologically active substances for manogastric or polygastric animals containing, as pH-sensitive substances, novel organosoluble chitosan derivatives.

 Chitosan is a basic polysaccharide, whose average molecular weight is generally greater than 500,000, which consists of 2-amino-2-deoxy-ss-D-glucopyranose 1,4-linked residues.

Chitosan can be obtained by deacetylation of chitin which can itself be extracted for example from the shell of crustaceans where it is found in large quantities.

 Due to its chemical structure, chitosan has the property of being resistant to hydrolysis at the level of the oside bonds in basic medium, of being insoluble at a pH greater than 6.5 and of being soluble and hydrolyzable in acidic conditions.

Given its film-forming properties, chitosan could be an agent of choice to protect various substances whose release must be regulated according to pH. However, chitosan has the disadvantage of being insoluble in common organic solvents, which makes it difficult to produce films or coating films.

 It has now been found, and this is the subject of the present invention, novel chitosan derivatives which are organosoluble and which retain the property of forming films and being sensitive to variations in pH.

dans lesquelles - R1 représente un radical alcanoyle contenant 2 & 4 atomes de carbone, de préférence un radical acétyle, - R2 représente un radical alkylidène contenant 3 à 14 atomes de carbone ou un radical benzylidéne dans lequel le noyau phényle peut être substitué par un ou plusieurs radicaux identiques ou différents choisis, par exemple, parmi les radicaux hydroxy ou néthoxy, - R3 représente un atome d'hydrogéna ou un radical acétyle ou un radical alcanoyle contenant 3 ou 4 atomes de carbone, et - R4 représente un atome d'hydrogène ou un radical alcanoyle contenant 2 & 4 atomes de carbone identique à R1, étant entendu que le chitosan dans le produit final a une masse moléculaire moyenne comprise entre 10.000 et 80.000, qu'il y a 60 à 65 t de fonctions amines sous forme imine (-N=R2) et de 3 à 35-40 t de fonctions amines sous forme acétylamino (-NH-COCH3), le reste des fonctions amines pouvant étre sous la forme amine primaire ou sous la forme acylamino et qu'une partie des radicaux R4 représente un atome d'hydrogène et l'autre représente un radical alcanoyle.The novel chitosan derivatives according to the invention consist of the statistical sequence of motifs which can be represented by the following formulas

in which - R 1 represents an alkanoyl radical containing 2 to 4 carbon atoms, preferably an acetyl radical, - R2 represents an alkylidene radical containing 3 to 14 carbon atoms or a benzylidene radical in which the phenyl nucleus may be substituted with one or a plurality of identical or different radicals chosen, for example, from hydroxyl or methoxy radicals, - R 3 represents a hydrogen atom or an acetyl radical or an alkanoyl radical containing 3 or 4 carbon atoms, and - R 4 represents a hydrogen atom or an alkanoyl radical containing 2 to 4 carbon atoms identical to R 1, it being understood that the chitosan in the final product has an average molecular mass of between 10,000 and 80,000, that there are 60 to 65 t of amine functions in imine form (-N = R2) and from 3 to 35-40 t of amine functional groups in the acetylamino form (-NH-COCH3), the remainder of the amine functions possibly being in the primary amine form or in the acylamino form and a part of the radicals R4 represents a hydrogen atom and the other represents an alkanoyl radical.

According to the invention the new chitosan derivatives can be obtained by carrying out the succession of the following steps
1) hydrolysis of the optionally purified chitosan to obtain fragments whose average molecular mass is between 10,000 and 80,000, preferably between 10,000 and 20,000
2) condensation of an aliphatic or aromatic aldehyde on the fragments obtained to convert the majority of the amine radicals to imino radicals, and
3) esterification of the primary alcohol functions and partially secondary alcohol functions and optionally total or partial acylation of primary amine functions unreacted with the aldehyde.

 The hydrolysis of the optionally purified chitosan, whose average molecular mass is greater than 500,000 and which contains between 75 and 97 t of free amine radicals and between 3 and 25 t of acetylated amino groups, is generally carried out by heating to a temperature close to 1000C in the presence of an aqueous solution of a strong mineral acid, such as hydrochloric acid.To obtain fragments whose average molecular mass is between 10,000 and 20,000, it is particularly advantageous to heat to a temperature of 1000C a solution of chitosan optionally purified at a concentration of between 30 and 40 g / liter in 0.5N hydrochloric acid for 15 to 20 hours. The chitosan fragments whose average molecular mass is between 10,000 and 20,000 are precipitated after alkalinization to a pH of about 10. The hydrolyzed chitosan is generally separated by filtration after removal of the inorganic salts, for example by washing with water.

 The condensation of an aliphatic or aromatic aldehyde with hydrolysed chitosan is generally carried out at an apparent pH of between 5.5 and 6, preferably close to 5.5 in a hydroalcoholic medium at a temperature of between 10 and 300 ° C.

Generally, a solution of chitosan in an aqueous organic acid, such as aqueous acetic acid, brought to a suitable pH by addition of a mineral base such as sodium hydroxide, is treated with the aldehyde in the presence of an alcohol such as than methanol. In order for the maximum of free amino groups to react with the aldehyde, it is particularly advantageous to use an excess of aldehyde with respect to the amine functions of the hydrolysed chitosan used.

 Depending on the nature of the aldehyde used, it is necessary for the molar ratio between the aldehyde and the hydrolysed chitosan to be greater than 15 and preferably equal to 20, the fixing rate of the aldehyde generally being between 55 and 70. t compared to the amine functions of chitosan.

 The condensation product of the aldehyde on chitosan is generally continuously extracted using a suitable organic solvent such as acetone.

 The esterification and optional acylation of the imine derivative of the hydrolyzed chitosan is generally carried out by means of an activated derivative of an organic acid, such as the anhydride or the acid chloride, operating in a basic organic solvent such as pyridine at a temperature between 10 and 300C. As the organic acid derivative, it is particularly advantageous to use anhydride and more particularly acetic anhydride.

 The chitosan derivatives according to the present invention have the property of being soluble in common organic solvents such as acetone, ethanol or methylene chloride.

Of particular interest are the chitosan derivatives consisting of the statistical arrangement of the units of general formulas (Ia) and (Ib) in which R2 represents an alkylidene radical containing 5 to 12 carbon atoms and more especially 7 to 10 atoms of carbon, the degree of fixation of the aldehyde being between 55 and 65 t relative to the chitosan osidic monomers, R1 represents an acetyl radical, R3 represents a hydrogen atom or an acetyl radical and a part of the radicals R4 represents one hydrogen atom and the other an acetyl radical.

 For example, the organosolubility which corresponds to the fraction of the organosoluble products in a crude product resulting from the reaction sequence can be determined by placing 10 g of ground crude acetylation product in 100 g of solvent and measuring the amount of product that solubilizes.

 In Table 1 are gathered the results giving the organosolubility of acetylated chitosan derivatives according to the invention in acetone.

TABLE 1

<tb><SEP> Aldehyde <SEP> used <SEP>% <SEP> of <SEP> binding <SEP> Mass <SEP> organosolu
<tb><SEP> of <SEP> aldehyde <SEP> molecular <SEP> bility <SEP> (g / l) <SEP>
<tb><SEP> propanal <SEP>. <SEP> 69 <SEP>: <SEP>. <SEP> 20
<tb><SEP> pentanal <SEP> 63 <SEP> 14 <SEQ> 14,500 <SEP>: <SEP> 65
<tb><SEP> heptanal <SEP> 61 <SEP> 17,500 <SEP> 100
<tb><SEP> decanal <SEP> 60-65 <SEP> 15,300 <SEP> 100
<tb><SEP> dodecanal <SEP> 58 <SEP> 80
<tb><SEP> tetradecanal <SEP> 59 <SEP> 50
<tb> hydroxy-4 <SEP> methoxy-3 <SEP> 60-65 <SEP> 15,400 <SEP> 95
<tb> benzaldehyde
<Tb>
The chitosan derivatives obtained by condensation of decanal on hydrolysed chitosan followed by acetylation are soluble, in particular in acetone (30-40% by weight), etbanol (15% by weight) and dichloroethane (10%). % in weight).

 The chitosan derivatives according to the invention are particularly useful for preparing compositions for coating food additives or biologically active substances intended mainly for ruminants which are stable in a medium having a pH equal to or greater than 5.5 and which allow the release of the food additive or the biologically active substance in a medium whose pH is less than or equal to 3.5.

 When ruminants are given certain biologically active substances (drugs, vitamins, amino acids), enzymatic destruction of these substances occurs during the passage through the rumen, favored by the residence time (a few hours to several days). and by the pH (between 5 and 6). As a result, the active substance that is degraded loses most of its effectiveness when it reaches the ruminant's abomasum and gut.

 It is therefore important to be able to protect these biologically active substances by coatings which are stable in the rumen of ruminants, that is to say which are stable to degradation by microorganisms and which allow the release of the biologically active substances in an environment. part of the digestive system, more particularly the abomasum, whose pH is less than or equal to 3.5. While the duration of protection in the rumen must be relatively long (a few hours to several days), the release of the active substance into the abomasum must take place in a relatively short time (a few minutes to a few hours).

 The novel chitosan derivatives according to the present invention can be used as sensitive pH substances in food additive coating compositions or biologically active substances for ruminants. In particular, the novel organosoluble chitosan derivatives may advantageously, totally or partially, replace synthetic sensitive pH substances such as basic copolymers such as copolymers of styrene with vinylpyridines which are usually used in this type of composition.

 The chitosan derivatives according to the invention, which are derived from natural products that are acceptable in the diet, are hydrolysed in the body into substances that are non-toxic to animals, which represents a considerable practical advantage.

More specifically, the new chitosan derivatives can be used as sensitive pH substances in coating compositions which are the subject of French patents
FR 2 514 261, FR 2 582 909, FR 2 575 039, FR 2 575 040, FR 2 603 458 or FR 2 606 597, and they can be implemented in the same way.

 The following examples, given by way of non-limiting example, illustrate the present invention.

EXAMPLE 1
1 - Purification of chitosan
40 g of technical chitosan (from the Company) are dissolved
SIGMA) in 3 liters of an aqueous solution of acetic acid at 4 t (w / v). The solution is filtered through a sieve whose mesh is 125 microns. The chitosan is precipitated by addition of an aqueous sodium hydroxide solution at 25 t to pH 9-11 and then separated by filtration and then washed on a sieve with distilled water and finally with absolute ethanol. The fibrous precipitate is stirred to extract the maximum of solvent and then it is dispersed in 1 liter of absolute ethanol. The mixture is heated at 800C for 2 hours. After filtration, the purified chitosan is dried under reduced pressure at 600 ° C. 35 g of purified chitosan are thus obtained, the characteristics of which are as follows: infra-red spectrum (from tablets mixed with KBr): main characteristic absorption bands at 3400, 2900 and 1650 cm - average molecular weight: greater than 500,000.

 The average molecular weight is determined by high performance liquid chromatography using 5 columns 50 cm long filled with G / CPG glycoprotein (ND Pierce) porosity 3125 (2 columns), 1902, 1038 and 547 A. The phase mobile is a tanpon acetate of 0.2M sodiun and 0.33M acetic acid at pH = 4.2. The flow rate is 1 cm3 / minute and the detection is done by refractometry.

Calibration is done by dextran standards (from the Company
SIGMA) with a molecular weight of 506,000 to 110,000.

percentage of desacetylation: 80% of the saccharide rings of the purified chitosan contain a free primary amine function.

 The percentage of deacetylation is determined in the following manner: a powdered sample of chitosan (particles of about 20 microns) is suspended in a water-dimethylsulfoxide mixture (9-1 by volume), the pH of which is brought to 11 by addition of 0.1N sodium hydroxide. The potentiometric assay is carried out with 0.1N hydrochloric acid.

- elementary analysis
C t = 40.90 H t = 6.76 N t = 7.52 O t = 44.82
2 - Acid hydrolysis of purified chitosan
25 g of purified chitosan are dissolved in 650 cm3 of 0.5N hydrochloric acid. The solution is heated in an oil bath at 980C for 18 hours. After cooling, the chitosan is precipitated slowly by addition of 5N sodium hydroxide until the pH of the reaction medium reaches 10. The salts formed are removed by dialysis. The purified chitosan is separated by filtration or centrifugation, washed with absolute ethanol and then dried under reduced pressure at 600C. 21 g of hydrolysed chitosan are thus obtained, the characteristics of which are as follows: average molecular mass: close to 13,000 (determination under the conditions described previously using 4 50 cm long columns filled with G / CPG glycophase (N. ) of porosity 1902, 1038, 547 and 242 A and dextran standards (from the Company
SIGMA) with a molecular weight of 110,000 to 9,000.

- elementary analysis
ct = 40.13 H t = 6.76 N t = 7.72 0 tt 45.39
3 - Condensation of decanal on hydrolysed chitosan
1 g of hydrolysed chitosan is dissolved in 20 cm3 of a 10% aqueous solution of acetic acid. The pH is adjusted to 4.5 by addition of a 5N sodium hydroxide solution and then 40 cm3 of methanol and 23 cm3 of decanal are added. The viscous reaction mixture is stirred for 18 hours at a temperature in the region of 200 ° C. The condensation product is extracted with acetone in a
Soxhlet for 5 hours. After drying under reduced pressure at 350 ° C., 1.48 g of the decanal-chitosan condensation product are obtained, the characteristics of which are as follows: infra-red spectrum (determination from tablets mixed with KBr): main characteristic absorption bands at 3450, 2930 and 2860 cm - percentage of decanal fixation: 60-65 t.

 The determination is carried out after reduction of the imine bond by sodium borohydride in ethanol-water medium (1-1 by volume). The percentage of secondary amine groups is determined by potentiometric determination of the reduced derivative dissolved in acetic acid-ethanol medium (2-1 by volume) by titrating with 0.1N perchloric acid.

4 - Acetylation of the Tuscan decanal condensation product
1 g of the decanal-chitosan condensation product is dispersed in 25 cm 3 of anhydrous pyridine. 5 cm3 of acetic anhydride are added. The reaction mixture is stirred for 24 hours at a temperature in the region of 20 ° C. After evaporation of the pyridine, the residue is taken up twice with toluene and then suspended in pentane. The suspension is filtered and then washed 3 times with 0.1N sodium hydroxide solution and then rinsed with distilled water until neutral. After transfer into a flask, the product is dried by entrainment of the water of acetone.

 The product obtained is purified by centrifugation at 16,000 rpm after dissolution in 50 cm3 of acetone. After evaporation of the supernatant, the residue obtained is washed on n-3 sintered glass with ethyl ether.

dont les caractéristiques sont les suivantes - pourcentage de fixation de l'aldéhyde : 60-65 % - pourcentage d'acétylamino : 20 à 35-40 % - spectre infra-rouge (détermination à partir de comprimés en mélange avec KBr) : principales bandes d'absorption caractéristiques à 3400, 2920-2830, 1750-1230, 1680, 1640 et 1540 cm-1 - spectre de résonance magnétique du 13 (90 MHZ - chloroforme deutéré - déplacement en ppm) : 170 (CO de l'acétyle) - 169 (CH=N) 102 (C-1) - 72,73 et 75,5 (C-2, C-3, C-4 et C-5) - 62,5 (C-6) - 31,8 (CH2 en 2') - 29,4 [(CH2)5 de la chaîne aldéhydique] - 25 (CH3 de
CH3CO) - 22,6 (CH2 en 9') - 20,8 (CH2 en 3') - 14,0 (CH3 en 10') - analyse élémentaire
C 5 2 61,00 H t = 8,17 N t = 3,28 0 t = 27,55 - masse moléculaire Mw = 15300 avec indice de polydispersité de 7,14.There is thus obtained 1.075 g of product of general formula (I) consisting of the statistical arrangement of the units of formula

the characteristics of which are as follows - percentage fixing of the aldehyde: 60-65% - percentage of acetylamino: 20 to 35-40% - infra-red spectrum (determination from tablets mixed with KBr): main bands characteristic absorption at 3400, 2920-2830, 1750-1230, 1680, 1640 and 1540 cm-1 - 13 magnetic resonance spectrum (90 MHZ - deuterated chloroform - displacement in ppm): 170 (CO of acetyl) - 169 (CH = N) 102 (C-1) - 72.73 and 75.5 (C-2, C-3, C-4 and C-5) - 62.5 (C-6) - 31, 8 (CH 2 in 2 ') - 29.4 [(CH 2) 5 of the aldehyde chain] - (CH 3 of
CH3CO) - 22.6 (CH2 at 9 ') - 20.8 (CH2 at 3') - 14.0 (CH3 at 10 ') - elemental analysis
C 5 2 61.00 H t = 8.17 N t = 3.28 0 t = 27.55 - molecular weight Mw = 15300 with a polydispersity index of 7.14.

 The molecular weight Mw is determined by gel filtration using 6 columns in series, a precolumn of 5 cs long (PL gel of 100 A), a 50 cm column (Shodex A 801), a column of 30 cm (PL gel 106 A), a column of 60 cm (PL gel 100) and two mixed columns of 60 cm (PL gel). Chromatography is carried out in dichloromethane with a flow rate of 1.5 cm 3 / minute.

The detection is done by refractometry. Mw 100 polystyrene standards are used at 4.106.

EXAMPLES 2 TO 6
By operating as in Example 1, starting from 1 g of hydrolyzed chitosan, and using different aliphatic aldehydes, the results obtained are collated in Table 2.

TABLE 2

<tb><SEP>:<SEP> Percentage <SEP> Weight <SEP> of <SEP> Product <SEP> Weight <SEP> of <SEP> Product
<tb> Examples <SEP> Aldehyde <SEP> of <SEP><SEP> SEP <SEP> condensation <SEP> condensation <SEP>
<tb> of <SEP> aldehyde <SEP> aldehyde-chitosam <SEP> aldehyde-chitosam <SEP>
<tb><SEP> acetylated
<tb><SEP> 2 <SEP>: <SEP> propanal <SEP> (9 <SEP> cm3) <SEP> 69 <SEP> 1.20 <SEP> g <SEQ> 0.160 <SEP> g
<tb><SEP> 3 <SEP> pentenal <SEP> (13 <SEP> cm3) <SEP> 63 <SEP> 1.36 <SEP> g <SEP> 0.580 <SEP> g
<tb><SEP> 4 <SEP> Heptanal <SEP> (17 <SEP> cm3) <SEP> 61 <SEP> 1.49 <SEP> g <SEP> 0.965 <SEP> g
<tb><SEP> 5 <SEP>: <SEP> dedecanal <SEP> (27 <SEP> cm3) <SEP> 58 <SEP> 1.70 <SEP> g <SEP> 0.900 <SEP> g
<tb><SEP> 6 <SEP>: <SEP> tetradecanal <SEP> (25 <SEP> g) <SEP><SEP> 59 <SEP> 1.82 <SEP> g <SEP> 0.600 <SEP> g
<Tb>
EXAMPLE 7
1 g of hydrolysed chitosan is dissolved in 20 cm 3 of a 10% aqueous acetic acid solution. The pH of the medium is adjusted to 4.5 by addition of a 5N sodium hydroxide solution. 10 cm3 of methanol and then a solution of 18 g of 4-hydroxy-3-methoxybenzaldehyde in 30 cm3 of methanol are added. The dark green reaction mixture is stirred for 18 hours at a temperature of 200C. The precipitate formed is extracted with acetone in a Soxhlet apparatus for 5 hours. After drying at 350 ° C. under reduced pressure, 1.65 g of the condensation product of 4-hydroxy-3-methoxybenzaldehyde are obtained with the hydrolyzed chitosan which is acetylated under the conditions described in Example 1.

dont les caractéristiques sont les suivantes - pourcentage de fixation de l'aldéhyde : 60-65 % - pourcentage d'acétylamino : 20 à 35-40 % - spectre infra-rouge (détermination à partir de comprimés en mélange avec KBr) : principales bandes d'absorption caractéristiques à 3400, 2945-2873, 1745-1227, 1691, 1647 et 1602-1507 cm-1 - spectre de résonance magnétique nucléaire du 13C (90 MHz - choroforme deutéré - déplacement en ppm) : 170,1 (CO de l'acétyle) - 152 (C-5') - 151,8 (C-4') - 135,3 (C-2') - 124,6 (C-7') - 123,4 (C-6') 111,0 (C-3') - 102,2 (C-1) - 75,9 à 72,4 (C-2, C-3, c-4 et C-5) 62,5 (C-6) - 56,1 (CH3-O).After purification, 0.990 g of product of general formula (I) consisting of the random sequence of units of formula:

the characteristics of which are as follows - percentage fixing of the aldehyde: 60-65% - percentage of acetylamino: 20 to 35-40% - infra-red spectrum (determination from tablets mixed with KBr): main bands characteristic absorption at 3400, 2945-2873, 1745-1227, 1691, 1647 and 1602-1507 cm-1 - 13C nuclear magnetic resonance spectrum (90 MHz - deuterated choroform - displacement in ppm): 170.1 (CO acetyl) - 152 (C-5 ') - 151.8 (C-4') - 135.3 (C-2 ') - 124.6 (C-7') - 123.4 (C-5 ') 6 ') 111.0 (C-3') - 102.2 (C-1) - 75.9 to 72.4 (C-2, C-3, c-4 and C-5) 62.5 ( C-6) - 56.1 (CH 3 -O).

EXAMPLE 8
According to the technique of the fluidized bed ("spray-coating") with a tank equipped with a WURSTER system, 350 g of previously granulated methionine are coated in the form of spherical particles titrating 98 t, the average diameter of which is between 0.63 and 0.80 mm by a solution having the following composition - stearic acid (mp = 68-690 ° C., acid number 194-198) .. 88 g - chitosan modified according to example 1 ....... ............... 22 g - dichloromethane ............................... .......... 500 cm3 - ethanol .................................... ............. 500 cm3
The solution, maintained at 280C, is sprayed in 60 minutes.

 449 g of granules grading 75% of methionine are thus obtained.

EXAMPLE 9
According to the technique of the fluidized bed with a tank equipped with a WURSTER system, 350 g of lysine hydrochloride granulated in the form of spherical particles, the average diameter of which is close to 0.8 mm, are coated with a solution whose composition is identical to that described in Example 8.

 The solution, maintained at 290C, is sprayed in 1 hour 34 minutes.

 448 g of granules grading 70 ml of lysine hydrochloride are thus obtained.

 In order to demonstrate the sensitivity of the coating compositions to variations in pH, tests are used which make it possible to measure the release of the active ingredient as a function of time at different pH values and, in particular, at pH = 6 and at pH = 2.

 For example, the release of the active substance present in the coated granules is examined by stirring, under specified conditions, a known quantity of granules in the buffered medium maintained at a constant pH at a temperature of 40 ° C. The kinetics of release are compared a sample at different pH values and more particularly at pH = 6 and pH = 2.

 With the granules which are the subject of Examples 8 and 9, the results obtained are collated in Table 3.

TABLE 3

<tb><SEP>%<SEP> of <SEP> substance <SEP> active <SEP> released <SEP>
<tb> Examples <SEP> Title <SEP> in <SEP> to <SEP> pH <SEP> = <SEP> 6 <SEP> after <SEP> to <SEP> pH <SEP> = <SEP> 2 <SEP > after <SEP>
<tb> substance <SEP> active <SEP> 1 <SEP> h <SEP> 6 <SEP> h <SEP> 24 <SEP> h <SEP> 15 <SEP> min <SEP> 1 <SEP> h <SEP > 2 <SEP> h <SEP> 3 <SEP> h <SEP> 5 <SEP> h
<tb><SEP> 8 <SEP> 75 <SEP>% <SEP> (methionine) <SEP> 0 <SEP> 1.2 <SEP> 3.0 <SEP> 0.7 <SEP> 1.1 <SEP> 7.3 <SEP> 44 <SEP> 100 <SEP>
<tb><SEP> 9 <SEP> 70 <SEP>% <SEP> (Hydrochloride <SEP> 2.7 <SEP> 10.0 <SEP> 20.0 <SEP> 1.2 <SEP> 3,4 <SEP> 37 <SEP> 100 <SEP>
<tb><SEP> of <SEP> lysine) <SEP>
<Tb>
The in vivo efficacy of the coating compositions according to the invention can be demonstrated in the following test
Samples of coated granules (about 0.5 g) are introduced into nylon bags having a mesh size of 300 x 300 microns. The pouches are placed in the rumen of fistulated ewes for 6 hours, 15 hours and 24 hours. The bags are recovered and washed. The amount of active substance present in the sachets is determined by an appropriate method.

 The results obtained are collated in Table 4.

TABLE 4

<tb> Time <SEP> of <SEP> stay <SEP>% <SEP> of <SEP> residual methionine <SEP><SEP>%<SEP> of <SEP> residual lysine <SEP>
<tb> in <SEP> the <SEP> rumen <SEP> Product <SEP> of <SEP> Example <SEP> 8 <SEP> Product <SEP> of <SEP> Example <SEP> 9
<tb><SEP> 6 <SEP> 99 <SEP> 1 <SEP> - <SEP>
<tb><SEP> 15 <SEP> 98.7 <SEP> 1 <SEP><SEP> 71 <SEP><SEP> 7
<tb><SEP> 24 <SEP> 96.8 <SEP><SEP> 0.6 <SEP> 67.5 <SEP><SEP> 2.5
<Tb>

Claims (8)

 1 - Composition for coating food additives or biologically active substances intended for feeding ruminants which are stable at a pH greater than 5 and which allow the release of the active substance at a pH below 3.5 characterized in that it contains, as pH-sensitive substance, an organosoluble derivative of chitosan which is constituted by the statistical sequence of motifs represented by the formulas

 in which - R 1 represents an alkanoyl radical containing 2 to 4 carbon atoms, - R2 represents an alkylidene radical containing 3 to 14 carbon atoms or a benzylidene radical in which the phenyl nucleus may be optionally substituted with one or more identical or different radicals. chosen from hydroxyl or methoxy radicals, - R3 represents a hydrogen atom or an acetyl or alkanoyl radical containing 3 or 4 carbon atoms, - R4 represents a hydrogen atom or an alkanoyl radical containing 2 to 4 identical carbon atoms at R1, it being understood that the chitosan in the final product has an average molecular mass of between 10,000 and 80,000, that there are 60 to 65 t of amine functional groups in the form of ixine -N = R2 and 3 to 35-40 t of amino functions in acetylamino -NH-CCCH3 form the rest of the amine functional groups may be in the primary amine form or in the acylamino form and that a part of the R 4 radicals represents a hydrogen atom. hydrogen and the other an alkanoyl radical.  2 - Composition according to claim i characterized in that in the sequence of units constituting the organosoluble derivative of chitosan R1 represents an acetyl radical - R2 represents an alkyl radical containing 5 to 12 carbon atoms - R3 represents a hydrogen atom or an acetyl radical, and a part of the radicals R4 represents a hydrogen atom and the other represents an acetyl radical.  3 - Composition according to claim 1 characterized in that in the sequence of units constituting the organosoluble derivative of chitosan - R1 represents an acetyl radical - R2 represents a radical decylidene - R3 represents a hydrogen atom or an acetyl radical, and - a part of the symbols R4 represents a hydrogen atom and the other an acetyl radical.  4 - Composition for coating food additives or biologically active substances intended mainly for feeding ruminants characterized in that it consists of an organosoluble chitosan derivative according to one of claims 1, 2 or 3 in combination with a hydrophobic substance whose melting point is greater than 600C.  5 - Coated granules characterized in that they consist of a core of active substance surrounded by a continuous film of a coating composition containing as pH-sensitive substance an organosoluble derivative of chitosan as defined in one of claims i, 2 or 3.  6 - Coated granules characterized in that they consist of a core of active substance surrounded by a continuous film of a coating composition according to claim 4.  7 - coated granules according to one of claims 5 or 6 characterized in that the active substance is chosen from drugs, vitamins and essential amino acids.  8 - coated granules according to one of claims 5 or 6 characterized in that the active substance is selected from methionine or lysine.