CROSS REFERENCE TO RELATED APPLICATIONS

This application is a National Stage application of PCT/FR2017/052932, filed Oct. 24, 2017, which claims the benefit of France Application No. 16.60521, filed Oct. 28, 2016, both of which are incorporated by reference in their entirety herein.

The invention relates to a composition of collagen peptides from fish skin(s). The invention relates in particular to such a peptide composition obtained by enzymatic hydrolysis of collagen from fish skin(s). The invention also relates to such a composition for use thereof as medicament. The invention therefore relates to the use of such a composition for the curative or preventative treatment of a disease affecting the human or animal body. The invention in particular relates to such a composition for use thereof as medicament in the treatment of a digestive candidiasis in humans or animals and/or in the treatment of an intestinal inflammation in humans or animals. The invention also relates to such a composition for use thereof as medicament in a microbiota-stimulating treatment.

FR 2 720 067 discloses a peptide powder obtained by hydrolysis by papain of a collagen-rich starting material originating from the skin or the skeleton of fish, mollusks or crustaceans. In the peptide powder obtained, 38% of the peptides have a molecular weight of between 10 000 Da and 50 000 Da. The peptide composition of FR 2 720 067 has a broad range of molecular weights, in particular a proportion of greater than 10% of peptides with a molecular weight of greater than 10 000 Da. The peptide composition of FR 2 720 067 is heterogeneous by the size of the peptides and has a fraction of water-soluble peptides of only between 80% and 90%. Such powders of peptides of high molecular weight greater than 10 000 Da are not perfectly water-soluble. They are also not entirely absorbable by the digestive tract and therefore pose problems in terms of their bioavailability.

FR 2 720 067 also describes the use of such a peptide powder obtained from fish living at great depth in the curative treatment of inflammation of the tendons of joints of the lower limbs (knees, fetlocks and hooves) of racehorses. Obtaining such a composition is problematic. This is because it requires the removal of deep-sea fish. In addition, such a powder is limited in its use to the treatment of joint inflammation.

The invention therefore aims to overcome these drawbacks.

The invention aims to propose a novel peptide composition resulting from an enzymatic hydrolysis of collagen from skin(s) of temperate-water fish, said peptides being water-soluble—that is to say 100% water-soluble—and entirely absorbable by the digestive tract.

The invention also aims to propose a novel peptide composition formed by enzymatic hydrolysis of collagen from skin(s) of temperate-water fish.

The invention also aims to propose such a peptide composition that is capable of being used as medicament.

The invention in particular aims to propose such a peptide composition resulting from an enzymatic hydrolysis of collagen from skin(s) of temperate-water fish that is capable of being used as medicament.

The invention aims to propose such a peptide composition having both a distribution of the apparent molecular weights of said peptides that extends over a narrow range of between approximately one hundred and a few thousands of Daltons, that is to say excluding peptides of high apparent molecular weight, and also having a low proportion of peptides of low apparent molecular weight.

The invention aims to propose such a peptide composition that is capable of being used as medicament in the treatment of intestinal candidiasis.

The invention also aims to propose such a peptide composition that is capable of being used as medicament in the treatment of inflammatory digestive diseases, especially inflammatory bowel diseases (IBDs).

The invention also aims to propose such a peptide composition that is capable of being used to promote equilibrium and maintenance of the intestinal flora (or microbiota).

The invention also aims to propose the use of such a peptide composition resulting from an enzymatic hydrolysis of collagen from skin(s) of temperate-water fish as dietary supplement.

For this purpose, the invention relates to a peptide composition having an aminogram in which:glycine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 20.0% and 24.5%;hydroxyproline is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 6.0% and 12.0%, especially between 7.0% and 11.0%;proline is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 10.6% and 14.6%, especially between 11.6% and 13.6%, in particular between 12.1% and 13.1%, preferably of the order of 12.6%; the peptide composition having, during analysis by exclusion chromatography during which each peptide of the peptide composition is eluted with a retention time that is representative of the apparent molecular weight of this peptide, an elution curve (that is to say an elution curve of a chromatogram) of the peptides having an area under the curve value (that is to say an area value representative of the amount by weight of peptides) corresponding to the peptides of apparent molecular weight of less than 1400 Da such that the ratio of this area value to the total area under the curve (corresponding to all the peptides of the composition) is less than 40%, especially less than 38%, preferably between 30% and 38%, in particular between 30% and 35%;
said analysis being performed as described below:on a filtration column of dimensions 300×7.8 mm comprising a stationary phase formed of a silica gel with a porosity of 5 μm;the column being kept at a temperature of 40° C.;with, as mobile phase, a solution formed (A) of ultrapure water comprising 0.1% by volume of trifluoroacetic acid and (B) of acetonitrile, wherein the A/B volume ratio is 75/25;introducing, at the top of the gel filtration column, a volume of a solution comprising the peptide composition;the flow rate of the mobile phase in the column being 0.6 ml/min, and;the peptides of the composition being detected by absorbance at a wavelength of 214 nm.

Throughout the text, “aminogram” in intended to mean the list of the free amino acids forming, by peptide chain (or bonding), the sequence of a peptide or the sequence of peptides of a mixture of peptides. Such an aminogram is obtained by analysis—especially by overall analysis or sequence analysis—of the amino acids constituting a peptide or a mixture of peptides.

In particular, the nature and the amount of the amino acids constituting the peptides of the composition according to the invention are determined by any method of overall analysis known per se to those skilled in the art. In particular, this analysis is carried out in accordance with standard ISO 13903:2005 by assay of the free and total amino acids using an amino acid analyzer or using high performance liquid chromatography (HPLC) equipment. Hydroxyproline is assayed by continuous flow analysis and colorimetric detection.

The aminogram obtained by overall analysis of the composition according to the invention is representative of the amino acid composition of collagen from skins of temperate-water fish. The invention therefore relates to such a peptide composition resulting from the enzymatic hydrolysis of collagen from skins of temperate-water fish by a cysteine protease of plant origin—especially a protease from the class EC 3.4.22.2-. The inventors have discovered that the use of such a cysteine protease makes it possible to obtain a composition of peptides which are water-soluble—that is to say 100% water-soluble—and which are entirely absorbable by the digestive tract and which have a distribution of apparent molecular weights which extends over a narrow range between approximately one hundred and a few thousands of Daltons, that is to say excluding peptides of high apparent molecular weights, and also having a low proportion of peptides of low apparent molecular weights.

In addition, a separation of the peptides of the peptide composition according to the invention is carried out as a function of their apparent molecular weight, by subjecting the peptide composition to a step of analytical separation by liquid chromatography on a porous silica gel filtration column (BioSep-SEC-S2000, Phenomenex, Le Peck, France) with a high surface density of silanol groups.

Use is made, as mobile phase, of a solution comprising (A) ultrapure water with trifluoroacetic acid (0.1% by volume) added thereto and (B) acetonitrile (A/B; 75/25; v/v). The gel filtration column is kept at a temperature of 40° C. during the analysis. The flow rate of the mobile phase in the stationary phase is 0.6 ml/min. The volume of peptide composition to be analyzed, introduced at the top of the gel filtration column, is 25 μl and the detection is carried out continuously by absorbance at a wavelength of 214 nm. A chromatogram is obtained, on which each peak is characterized by a duration or retention time value (expressed in minutes following the introduction of the mixture to be analyzed at the top of the column) determined at the maximum absorbance value of the peak. The apparent molecular weight of each peptide corresponding to this retention time value at the maximum absorbance value of each peak is determined by means of a predetermined calibration curve obtained by analysis—under the same chromatographic conditions as described above—of peptides of determined apparent molecular weights. For example, such a calibration curve is produced by analyzing, under these same chromatographic conditions, a mixture of reference peptides/proteins with known apparent molecular weights of between 100 Da and 30 kDa.

With the chromatogram representing the variation in absorbance at 214 nm over the course of the chromatographic analysis of the peptide composition of the invention, the proportion of peptides of apparent molecular weight of less than 1400 Da is determined by evaluating the ratio of the value of the area under the curve—that is to say the value of the sum of the areas under the peaks of the curve—corresponding to the peptides of apparent molecular weight of less than 1400 Da to the value of the total area under the whole of the curve—that is to say the value of the sum of the areas under each peak of the curve—and corresponding to all the peptides of the peptide composition.

The invention therefore relates to a composition of peptides resulting from an enzymatic hydrolysis of collagen from skins of temperate-water fish by a cysteine protease of plant origin, each peptide of the composition having a number of amino acids of between 2 and a few tens, preferably of between 2 amino acids and 100 amino acids.

In some embodiments, the composition according to the invention has an aminogram in which glycine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 20.0% and 22.4%, especially between 20.0% and 21.9%, in particular between 20.4% and 21.4%.

In some other embodiments, the composition according to the invention has an aminogram in which glycine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 22.4% and 24.9%, especially between 22.9% and 24.4%, in particular between 23.0% and 24.0%.

In some embodiments, the composition according to the invention has an aminogram in which hydroxyproline is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 7.0% and 9.0%, in particular between 7.5% and 8.5%, preferably between 7.7% and 8.5%.

In some other embodiments, the composition according to the invention has an aminogram in which hydroxyproline is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 9.5% and 11.5%, in particular between 10.0% and 11.0%, preferably of the order of 10.5%.

In some embodiments, the composition according to the invention has an aminogram in which glutamic acid is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 8.0% and 13.0%.

In some embodiments, the composition according to the invention has an aminogram in which glutamic acid is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 8.0% and 10.0%, in particular between 8.5% and 9.5%, preferably between 9.0% and 9.5%.

In some other embodiments, the composition according to the invention has an aminogram in which glutamic acid is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 10.5% and 12.5%, in particular between 11.0% and 12.0%, preferably of the order of 11.6%.

In some embodiments, the composition according to the invention has an aminogram in which arginine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 6.9% and 10.9%, especially between 7.9% and 9.9%, in particular between 8.0% and 9.0%.

In some embodiments, the composition according to the invention has an aminogram in which alanine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 7.3% and 11.5%, especially between 8.0% and 10.0%, in particular between 8.1% and 9.6%.

In some embodiments, the composition according to the invention has an aminogram in which aspartic acid is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 3.1% and 7.1%, especially between 4.1% and 6.1%, in particular between 4.6% and 5.6%, preferably between 5.0% and 5.5%.

In some embodiments, the composition according to the invention has an aminogram in which lysine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 1.5% and 5.5%, especially between 2.5% and 4.5%, in particular between 3.0% and 4.0%, preferably between 3.1% and 3.6%.

In some embodiments, the composition according to the invention has an aminogram in which serine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 1.5% and 5.5%, especially between 2.5% and 4.5%, in particular between 3.0% and 4.0%, preferably between 3.2% and 3.6%.

In some embodiments, the composition according to the invention has an aminogram in which threonine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 0.7% and 4.7%, especially between 1.7% and 3.7%, in particular between 2.2% and 3.2%, preferably between 2.4% and 2.8%.

In some embodiments, the composition according to the invention has an aminogram in which leucine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 0.6% and 4.6%, especially between 1.6% and 3.6%, in particular between 2.1% and 3.1%, preferably between 2.4% and 2.9%.

In some embodiments, the composition according to the invention has an aminogram in which phenylalanine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 0.3% and 4.3%, especially between 1.3% and 3.3%, in particular between 1.8% and 2.8%, preferably between 1.8% and 2.4%.

In some embodiments, the composition according to the invention has an aminogram in which valine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 0 and 4.0%, especially between 1.0% and 3.0%, in particular between 1.5% and 2.5%, preferably between 1.8% and 2.5%.

In some embodiments, the composition according to the invention has an aminogram in which isoleucine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 0 and 3.5%, especially between 0.5% and 2.5%, in particular between 0.9% and 2.0%, preferably between 0.9% and 1.6%.

In some embodiments, the composition according to the invention has an aminogram in which hydroxylysine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 0 and 3.5%, especially between 0.5% and 2.5%, in particular between 1.0% and 2.0%, preferably of the order of 1.5%.

In some embodiments, the composition according to the invention has an aminogram in which histidine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 0 and 3.3%, especially between 0 and 2.3%, in particular between 0.5% and 1.5%.

In some embodiments, the composition according to the invention has an aminogram in which methionine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 0 and 2.5%, especially between 0 and 2.0%, in particular between 0.5% and 1.8%, preferably between 0.7% and 1.6%.

In some embodiments, the composition according to the invention has an aminogram in which tyrosine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 0 and 1.5%, especially between 0 and 1.0%, in particular between 0 and 0.9%, preferably between 0.2% and 0.8%.

In some embodiments, the composition according to the invention has an aminogram in which cysteine and cystine are in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 0 and 2%, especially between 0 and 1.0%, in particular between 0 and 0.5%, preferably of the order of 0.03%.

The composition according to the invention has an aminogram in which the essential amino acids (lysine, methionine, phenylalanine, threonine, valine, leucine and isoleucine and histidine) are together between 15% and 20%.

Advantageously, in some embodiments according to the invention, at least 90%—especially at least 95%—of the peptides of the peptide composition have an apparent molecular weight of less than 15 000 Da (Daltons), especially of between 200 Da and 15 000 Da, preferably of between 200 Da and 14 000 Da, in particular of between 200 Da and 13 000 Da. Advantageously, in some embodiments according to the invention, at least 90% of the peptides of the composition have an apparent molecular weight of between 200 Da and 12 000 Da.

The peptides of the composition according to the invention have a tight distribution of apparent molecular weights, extending over a narrow range of between approximately one hundred and a few thousands of Daltons, that is to say excluding peptides of high apparent molecular weights having more than 100 amino acids, and also having a low proportion of peptides of low apparent molecular weights.

Advantageously, in some embodiments according to the invention, the peptides of the composition have a mean apparent molecular weight value of between 2500 Da and 3600 Da, especially of between 2700 Da and 3600 Da. Since each peptide of the peptide composition according to the invention makes a weight contribution to the composition, the mean apparent molecular weight value of peptides of the composition corresponds to the mean of each value, referred to as the weighted value, of the apparent molecular weight of each weighted peptide by a value representative of the weight contribution of each peptide in the composition. The value representative of the weight contribution of each peptide or group of peptides in the composition is expressed as a percentage of the value of the area under the curve corresponding to said peptide or to said group of peptides over the value of the total area under the curve corresponding to all the peptides of the composition.

In practice, the weighted apparent molecular weight value of a group of peptides corresponding to the same peak of a chromatogram corresponds to the apparent molecular weight value read off at the top (maximum) of this peak of the chromatogram, multiplied by the ratio of the value of the area under the curve of this peak to the (total) area under the curve of the chromatogram. “Area under the curve” or “area under the peak” is intended to mean the area of the space between the curve tracing the peak of the chromatogram and the baseline of the chromatogram. In particular, the area under one of the peaks of the chromatogram extends between two minima of the curve of the chromatogram, enclosing a top (or maximum) of the curve of the chromatogram.

According to some embodiments, the peptide composition according to the invention has, during analysis thereof by exclusion chromatography during which each peptide of the peptide composition is eluted with a retention time that is representative of the apparent molecular weight of this peptide, a curve representative of the elution (that is to say a chromatogram) of the peptides having a value of the area under this curve (that is to say an area value representative of the amount by weight of peptides) corresponding to the peptides of apparent molecular weight of greater than 10 000 Da—especially of between 10 000 Da and 50 000 Da such that the ratio of this area value to the total area under the curve (corresponding to all the peptides of the composition) is less than 15%, especially less than 10%. According to some particular embodiments, this ratio is between 2.5% and 8.5%.

According to other particular embodiments, the peptide composition according to the invention may be devoid of any peptide of high molecular weight. According to these other particular embodiments, the peptide composition according to the invention is formed of water-soluble peptides.

According to some embodiments, the peptide composition according to the invention has, during analysis thereof by exclusion chromatography during which each peptide of the peptide composition is eluted with a retention time that is representative of the apparent molecular weight of this peptide, a curve representative of the elution (that is to say a chromatogram) of the peptides having a value of the area under this curve (that is to say an area value representative of the amount by weight of peptides) corresponding to the peptides of apparent molecular weight of between 1800 Da and 10 000 Da such that the ratio of this area value to the total area under the curve (corresponding to all the peptides of the composition) is greater than 35%—especially between 35% and 70%, in particular between 45% and 65%. According to some particular embodiments, this ratio is between 49% and 55%.

According to some embodiments, the peptide composition according to the invention has, during analysis thereof by exclusion chromatography during which each peptide of the peptide composition is eluted with a retention time that is representative of the apparent molecular weight of this peptide, a curve representative of the elution (that is to say a chromatogram) of the peptides having a value of the area under this curve (that is to say an area value representative of the amount by weight of peptides) corresponding to the peptides of apparent molecular weight of between 600 Da and 1800 Da such that the ratio of this area value to the total area under the curve (corresponding to all the peptides of the composition) is between 15% and 45%—especially between 20% and 40%, in particular between 25% and 35%. According to some particular embodiments, this ratio is between 27% and 32%.

According to some embodiments, the peptide composition according to the invention has, during analysis thereof by exclusion chromatography during which each peptide of the peptide composition is eluted with a retention time that is representative of the apparent molecular weight of this peptide, a curve representative of the elution (that is to say a chromatogram) of the peptides having a value of the area under this curve (that is to say an area value representative of the amount by weight of peptides) corresponding to the peptides of apparent molecular weight of less than 600 Da such that the ratio of this area value to the total area under the curve (corresponding to all the peptides of the composition) is less than 10%. According to some particular embodiments, this ratio is between 8.5% and 14.5%.

Advantageously, the composition according to the invention has, by chromatographic analysis on an anion exchange column during which each peptide of the peptide composition is eluted from the column with a retention time that is representative of its charge:an area value under a peak corresponding to the anionic peptides;an area value under a peak corresponding to the neutral peptides, and;an area value under a peak corresponding to the cationic peptides;
such that the ratio of this area value under the peak corresponding to the anionic peptides to the sum of the area values under the peaks corresponding to the anionic peptides, to the neutral peptides and to the cationic peptides of the composition is between 27.0% and 45%, especially between 30% and 45%, in particular between 35% and 43%, preferably between 35% and 40%;
the value of the area under the peak corresponding to the anionic peptides, the value of the area under the peak corresponding to the cationic peptides and the value of the area under the peak corresponding to the neutral peptides being determined by chromatographic analysis under the conditions described below:using a chromatographic column of dimensions 100×7.8 mm comprising, as stationary phase, a hydrophilic anion exchange resin functionalized with quaternary ammonium groups with a particle size of 10 μm;using, as first mobile phase for elution of the cationic peptides and neutral peptides, a 5 mM aqueous Tris buffer (C) at pH 8.35 for a duration of 7 minutes starting from the introduction of the composition to be analyzed at the top of the column, then a second mobile phase for elution of the anionic peptides, in which the ratio of the volume of a buffer (D) formed of 5 mM Tris, 5 M NaCl at pH 8.35 to the volume of buffer (C) increases linearly from 0 to 100% in 30 minutes;with a flow rate of the mobile phase of 1 ml/min in the column;the analysis being performed at a temperature of 25° C., and;with detection by absorbance at a wavelength of 214 nm at the column outlet.

Advantageously according to the invention, the peptides of the composition according to the invention have, during a reversed-phase liquid chromatography hydrophobicity analysis, a retention time of between 16 min and 36 min; said hydrophobicity analysis being performed under the conditions below:using a chromatography column of dimensions 250×4.6 mm having a stationary phase formed of silica grafted with butyl groups, of a particle size of 5 μm and of a porosity value of 300 Å;using, as first mobile phase for elution of the hydrophilic peptides, a solution (E) of trifluoroacetic acid at 0.1% in ultrapure water for a duration of 7 minutes starting from the introduction of the composition to be analyzed at the top of the column, then a second mobile phase for elution of the hydrophobic peptides in which the ratio of the volume of a solution (F) of trifluoroacetic acid at 0.1% (by volume) in water comprising 40% acetonitrile to the volume of the solution (E) increases linearly from 0 to 40% in 30 minutes;with a flow rate of the mobile phase of 0.6 ml/min in the column;the analysis being performed at a temperature of 40° C., and;with detection by absorbance at a wavelength of 214 nm at the column outlet.

In some embodiments, a composition according to the invention comprises peptides which are hydrophobic by nature which are eluted from the column mentioned above and under the conditions specified above with a retention time corresponding to a percentage of acetonitrile of between 12% and 38%. The median retention time of the peptides of the composition according to the invention is 26 min, corresponding to a percentage of acetonitrile of 25% in the eluent.

Advantageously according to the invention, the peptide composition is in the liquid state. It may be a solution of the peptide composition according to the invention in a liquid solvent, especially in an aqueous solvent.

Advantageously according to the invention, the peptide composition is in the solid state. The peptide composition may be in the form of a solid in the divided state. It may in particular be a solid in the at least partially dehydrated state. The peptide composition according to the invention may be in powder form.

Advantageously according to the invention, the peptide composition is devoid of carbohydrates.

Advantageously according to the invention, the peptide composition is devoid of fats.

Advantageously according to the invention, the solids of the peptide composition comprises a proportion by weight of collagen peptides of greater than 95%, especially greater than 99%. Thus, the peptide composition has an amount of collagen peptides such that the weight ratio of the collagen peptides of the solids of the peptide composition to the solids of the peptide composition is greater than 95%, especially greater than 99%.

Advantageously according to the invention, the peptides of the composition are water-soluble. Advantageously, the peptides of the peptide composition are 100% water-soluble. Advantageously, the peptide composition is hydrocompatible.

Advantageously according to the invention, the peptides of the peptide composition result from a controlled enzymatic hydrolysis of collagen from skin of at least one fish selected from the group formed of fish from the family Pangasiidae—especiallyPangasius hypophtalmus(orPangasianodon hypophtalmus),Pangasius pangasius, Pangasius bocourti—and from the family Cichlidae—especially from the genusOreochromis, in particularOreochromis niloticusor from the genusTilapia. Advantageously, the peptides of the peptide composition result from a controlled enzymatic hydrolysis of collagen from skin of at least one fish found in a temperate water of a temperate region.

The invention also extends to the use of such a peptide composition in a therapeutic treatment for the human or animal body. The invention therefore also extends to such a peptide composition for use thereof as medicament. The invention therefore extends to such a peptide composition for use thereof as medicament in the preventative or curative treatment of at least one disease of the human or animal body.

The invention also extends in particular to a peptide composition for use thereof as medicament in at least one of the following treatments:treatment of a digestive disease;treatment of an intestinal candidiasis;treatment of a digestive inflammation, and;maintenance of the intestinal microbiota.

The invention also extends to any use of the peptide composition according to the invention in human food. According to some embodiments, the invention also extends to any use of a peptide composition according to the invention in human food, excluding any use as medicament. In particular, the composition according to the invention is advantageously used as dietary supplement.

The invention also extends to a peptide composition obtained by a process in which:skins of temperate-water fish are selected—especially from the family of the Pangasiidae and/or from the family of the Cichlidae, then;the following are carried out in succession:at least one step of washing the skins, then;at least one step of acid or alkaline treatment of the skins, suitable for enabling extraction of at least a portion of the collagen from the skins, then;at least one step of hydrolysis of the collagen by at least one cysteine protease of plant origin—in particular at least one protease ofCarica papaïa—at a temperature of less than 75° C., then;interruption of the enzymatic hydrolysis by heating the collagen hydrolyzate to a temperature greater than the denaturation temperature of each cysteine protease, so as to form the peptide composition.

Advantageously, in a process according to the invention, after the step of acid or alkaline treatment of the skins, at least one liquid/solid extraction of the collagen is carried out in water brought to a temperature of between 60° C. and 98° C. Advantageously, in a process according to the invention, a step of separation is next carried out—especially a step of separation by settling out—of a fraction comprising solids (and fats) and of a solution comprising the extracted collagen, then the solution comprising the extracted collagen is subjected to a purification step, for example by filtration over earth and/or demineralization on an ion exchange resin, suitable for forming a purified solution comprising collagen, the solids of the purified solution comprising a proportion by weight of collagen of at least 99%, especially at least 99.5%, in particular at least 99.8%. A purified solution of collagen is formed, comprising substantially pure collagen. In particular, such a purified solution of collagen is formed which is substantially colorless. In particular, such a purified solution of collagen is formed which is substantially—especially totally—devoid of elastin. The purified solution is concentrated so as to form a purified collagen gel, and the purified collagen gel is then subjected to the step of hydrolysis of the collagen.

Such a process makes it possible to obtain a peptide composition according to the invention which is substantially colorless. Such a process makes it possible to obtain a peptide composition according to the invention which is substantially devoid of elastin. In particular, it makes it possible to obtain such a peptide composition according to the invention without any chromatographic step of purification of the peptide composition.

In such a process, advantageously and according to the invention, a subsequent step of filtration of the peptide composition is carried out. Advantageously, a step of pasteurization of the peptide composition is also carried out for a duration of at least 2 minutes at a pasteurization temperature of between 85° C. and 90° C. at least.

In such a process, advantageously and according to the invention, a step of drying the peptide composition is carried out. This drying step is carried out by spray drying so as to form a composition according to the invention that is substantially dehydrated and in powder form.

The invention therefore extends to a peptide composition obtained by a process in which:skins of temperate-water fish are selected—especially from the family of the Pangasiidae and/or from the family of the Cichlidae, then;the following are carried out in succession:at least one step of washing the skins, then;at least one step of acid treatment of the skins, suitable for enabling extraction of at least a portion of the collagen from the skins, then;at least one step of hydrolysis of the collagen by at least one cysteine protease of plant origin—in particular at least one protease ofCarica papaïa—at a temperature of less than 75° C., then;interruption of the enzymatic hydrolysis by heating the collagen hydrolyzate to a temperature greater than the denaturation temperature of each cysteine protease, so as to form the peptide composition;
said peptide composition having an overall analysis of the amino acids in which:glycine is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 20.0% and 24.5%;hydroxyproline is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 6.0% and 12.0%;proline is in a molar amount such that the ratio of this amount to the sum of the molar amounts of the amino acids in the composition is between 10.6% and 14.6%; the peptide composition having, during analysis by exclusion chromatography during which each peptide of the peptide composition is eluted with a retention time that is representative of the apparent molecular weight of this peptide, an elution curve (that is to say an elution curve of a chromatogram) of the peptides having an area under the curve value (that is to say an area value representative of the amount by weight of peptides) corresponding to the peptides of apparent molecular weight of less than 1400 Da such that the ratio of this area value to the total area under the curve (corresponding to all the peptides of the composition) is less than 40%;
said analysis being performed as described below:on a filtration column of dimensions 300×7.8 mm comprising a stationary phase formed of a silica gel with a porosity of 5 μm;the column being kept at a temperature of 40° C.;with, as mobile phase, a solution formed (A) of ultrapure water comprising 0.1% by volume of trifluoroacetic acid and (B) of acetonitrile, wherein the A/B volume ratio is 75/25;introducing, at the top of the gel filtration column, a volume of a solution comprising the peptide composition;the flow rate of the mobile phase in the column being 0.6 ml/min, and;the peptides of the composition being detected by absorbance at a wavelength of 214 nm.

The invention also relates to a peptide composition, such a peptide composition for use thereof as medicament, a process for obtaining such a peptide composition and such a peptide composition obtained by such a process for obtaining same, characterized in combination by all or a portion of the characteristics mentioned above or below.

