FI104457B - Protein composition as well as its preparation and use as well as its containing products and their preparation - Google Patents

Description

104457

The protein composition and its preparation and use, as well as preparations containing it and their preparation

The present invention relates to a protein composition and its preparation and use, as well as to preparations containing it and their preparation. In particular, the invention relates to a substantially non-insulin protein composition and its preparation and use, and preparations containing it, such as substantially non-insulin infant formulas and other nutritional supplements, and their preparation.

Adolescent diabetes mellitus, or insulin-deficient diabetes mellitus (IDDM), is a disease in which the insulin-producing beta cells of the pancreatic islets of Langerhans are destroyed, but the other islets remain intact. This disease usually begins at the latest in childhood.

Despite numerous medical studies, it is not yet known exactly what factors are responsible for the onset of adolescent diabetes. At present, however, it is believed that the risk of children developing adolescent diabetes is influenced by both hereditary and environmental factors. , workers like nutrition.

Several epidemiological studies include: it has been found that exposure to cow's milk proteins during infancy increases the risk of developing adolescent diabetes *: "·· (Gerstein, Diabetes Care 17 (1994) 13-19). Based on epidemiological observations, several hypotheses have been presented regarding the mechanisms by which cow milk proteins could function as diabetogenic agents. It has been found that 30 immune responses to bovine serum albumin (Karjalainen et al., 1992, N. Engl. J. Med. 327: 302-307 (1992)), beta-lactoglobulin (Vaarala et al., Diabetes 45 (1996)). 178-182) or Beta * V 'to T-Casein (Cavallo et al., Lancet 348 (1996) 926-28) could lead to beta-cell reactivity, but up to now, bovine insulin has not even been suspected. - · diabetic risk factor.

• · 2 104457

Cows' strips are normally known to contain small amounts of bovine insulin. The milk insulin content reported in the literature varies depending on the assay method, but for example ELISA has shown concentrations of about 50 ng / ml. The milk has a peak insulin concentration (about 330 ng / ml) immediately after calving, followed by a decline to a constant level (about 46 ng / ml) about 7 days after calving (Aranda et al., J. Dairy Sci. 74 (12) (1991) ) 4320-4325).

10 In its studies, the applicant stated that the administration of a conventional infant formula based on infant formulas produces antibodies, such as insulin antibodies, to bovine insulin. The IgG-grade antibodies to bovine insulin in 6-month-old infants and 6-month-old infants in conventional breast-milk supplement (Enfamil) are shown in Figure 1. These antibodies cross-react with human insulin.

Because the presence of insulin autoantibodies (IAA) precedes and predicts the onset of IDDM, immunization with bovine insulin from cow's milk based products may be detrimental and increase the risk of developing IDDM. market cows' milk, v 25 products and cow's milk products that do not contain immunoreactive bovine insulin.

Liquid chromatography and reversed phase columns have traditionally been used to purify insulin from production and extraction solutions (Kroeff et al., J. Chromatogr. 46: 30 (1989) 45-61; Poli et al., J. Chromatogr. 539 (1991) 37). Cox, J. Chromatogr. 599 (1992) 195-203; Welinder, J. Chromatogr. 542 (1991) 83-99), gel filtration or anion exchange column (WO 90/00176 and WO 90/00177). or a weak cation exchange column (DE 3511270 A1 and GB 35 2173503 A). Reverse phase or gel filtration chromatography «· 1 • · 1 · 104457 3 as such is not suitable for milk treatment as the treated milk should be food grade and reasonably priced.

However, no prior art publication 5 known to the applicant discloses or even suggests the isolation or removal of bovine insulin present in cow's milk.

The Applicant has now discovered how to make cow's milk a reasonably priced food-grade protein 10 composition which is substantially free of bovine insulin and which, as such, is particularly suitable as a protein component of infant formulas, but also of other specialized nutritional products.

The invention thus relates essentially to an insu-15 lineage-free protein composition useful as a protein component of a nutritional preparation or as a raw material for drinking milk and characterized by being a non-fat proteinaceous material derived from cow's milk, such as whey, skimmed milk or curd. has been substantially removed.

When fat and casein are removed from milk, it is obtained. , whey containing whey proteins. About 20% of the total milk protein is whey protein and the remainder is "casein." Whey obtained during cheese making is known as "cheese whey" and whey obtained in casein * is referred to as casein whey.

A: The whey used in the invention may be any whey from cow's milk, such as cheese whey, rennet casein, acid casein whey or sour cheese whey. Preferably, the whey is a cheese whey.

• * ·

In addition to whey, the invention may use skimmed milk or an aqueous solution of skimmed milk powder or an aqueous solution of milk casein, i.e. casein, as the fat-free proteinaceous protein derived from cow's milk.

I · «« «*« • · · 4 104457

The substantially insulin-free protein composition of the invention may conveniently be prepared by the process of the invention, characterized in that 5a) the liquid fat-free proteinaceous material from cow's milk is subjected to an insulin removal treatment, optionally first clarified and subsequently derivatized at pH 5. , 2 to 5.6, 10 through a column packed with a strong cation exchange resin regenerated to the Na + or K + form, b) concentrating the liquid proteinaceous material obtained in step a) by ultrafiltration and diafiltration using 15 membranes of 6,000-20,000 D cut -off membranes, the resulting protein concentrate is evaporated and optionally dried to a protein powder, c) optionally 1) a protein concentration of 1-20% from the protein concentrate 20 or protein powder obtained in step b), 2) hydrolyzed by the protein solution obtained in step cl) ti) with proteases; and 3) ultrafiltration of the protein solution hydrolyzed in step c2), and d) optionally derivatizing the pro- obtained in step c). (e) evaporate the solution obtained in step c) or d) and dry to a powder.

In step (a), the non-fat * * protein * material from cow's milk is subjected to an insulin removal treatment to obtain substantially all or at least a significant portion of the> 35 bovine insulin present in the material. deleted. In the removal of insulin 104457, the liquid material from the cow's milk is treated with a strong cation exchange resin. In addition, it can be clarified, for example, by microfiltration or centrifugation. The whey can also be clarified by ultrafiltration at this stage.

However, the best insulin removal results are obtained when the material to be purified is first clarified as described above and then treated with a strong cation exchange resin.

Whey, such as cheese or casein whey, is most preferably purified from bovine insulin with a strong cation exchange resin. Suitable cation exchange resins are, for example, Amberlite C-20 (Rohm & Haas, France) and Spherosil S (Rhone -Poulenc, France). The whey to be purified, having a pH reduced to 5.2 to 5.6 with a food grade acid such as HCl or ion exchange, is passed at a temperature of 3-65 ° C, suitably at room temperature, into a column filled with a strong cation exchange resin regenerated to Na + or K +. through. In the batch process, the feed rate and volume 20 may vary, but suitably the feed rate is 1 to 10 column volumes (BV) / h and the feed volume is 5 to 60 BV. . preferably 20 BV. In the pH range of 5.2 to 5.6 whey proteins: are generally negatively charged but insulin: "is positively charged because its isoelectric point is 5.6 (Erkama, Biochemistry, p. 185). a substantial part of the bovine insulin in the whey binds to the cation exchange resin by the negatively charged whey proteins through the column.

Preferably, the whey is purified from bovine insulin with a strong cation exchange resin at pH 5.4.

• · ·. ... No bovine insulin was detected on the electro spray mass spectrometer with a strong cation exchange resin.

Also, the bovine lipid concentration of an aqueous solution of skimmed cow's milk and milk casein 35, i.e. a casein solution, can be significantly reduced by a corresponding treatment with a strong cation exchange resin. Thus, the insulin content of the casein solution is reduced by at least about 65% and the insulin content of skimmed milk is reduced by at least about 40%.

Insulin was analyzed from samples on an electro spray mass spectrometer (VG Quattro II, VG BioTech, Altrincham, England) using fractionation on a C18 reverse phase column (Hewlett Packard HPLC 1090, Hewlett 10 Packard Co. Germany) as eluent (eluent: A: 0.05%). trifluoroacetic acid (TFA) in water, B: acetonitrile (Acn) + 0.05% TFA, gradient: 15% to 40% in 20 min, 40% to 100% in 10 min). The insulin-containing fraction was freeze-dried, redissolved in a concentrated solution and transferred to a mass spectrometer.

The whey can be purified from bovine insulin by clarifying the whey by, for example, microfiltration, ultrafiltration or centrifugation to remove the residual casein and other high molecular weight proteins to which the insulin as a hydrophobic protein is bound.

The whey to be purified by microfiltration is passed, suitably at 10-60 ° C, through microfiltration membranes of I; : the films are 0.05 to 1.4 micrometers, preferably: 0.1 micrometers.

The whey to be purified by ultrafiltration, in turn, is treated with ultrafiltration membranes, which are preferably: films having a cut-off value of 50,000 to 200,000 Dalto:.

For centrifugation, the whey is preferably treated at 30 to 1000 rpm.

• Tue # ·. · # Bleaching treatment reduces whey bovine insulin content by 6 to 10%.

The clarification treatment and the cation exchange resin treatment both reduce the bovine insulin content of the cow's milk material to be treated, but

I · I

The results obtained when the material in question is first clarified and subsequently subjected to a cation exchange resin treatment. The best results are obtained with whey treatment, with microfiltration being the most preferred clarification procedure.

In step a), the proteinaceous material purified from bovine insulin, preferably whey, is concentrated by ultrafiltration and diafiltration to obtain sufficient protein content, then the protein concentrate thus obtained is evaporated and optionally dried to protein protein powder. In this case, material substantially purified from bovine insulin, such as whey, having a pH adjusted to about 6.5 with a food grade base such as NaOH or Ca (OH) 2, or by ion exchange, is subjected to ultrafiltration and diafiltration By means of a semi-permeable membrane, the higher molecular weight proteins are obtained from lactose, salts and smaller molecular weight proteins, such as insulin having a molecular weight of about 5734 D. Suitably the cutoff value of the semipermeable membrane is 6000 to 20,000 D, preferably 20 to 10,000 D for example, a polyether sulfone film having a cut-off value of 10,000 D may be used.

The whey obtained in step a) can be concentrated at a total concentration ratio which is suitably about 120. In this case, the whey treated in step a) is preferably ultrafiltered with 10,000 D cut off membranes first, for example at a concentration ratio of 10, followed by a *; The resulting whey protein concentrate having a protein content of · · * about 90% of the dry matter is evaporated and dried • · · “* • · · *. as a powder, for example by spray drying or freeze drying.

Conventionally, ultra- and diafiltered whey protein-35 powder, having a protein content of 70-80%, contains • ~ 8 104457 43-48 micrograms insulin / gram powder (approximately 60 micrograms insulin / gram protein).

In contrast, the whey protein powder obtained according to steps a) and b) of the process of the invention contains 5 significantly less bovine insulin than the conventionally ultra- and diafiltered whey protein powder mentioned above. With the strong cation exchange resin in step a), no whey protein powder made from bovine insulin-purified whey protein was found on bovine insulin-10 electro spray mass spectrometer. The essentially non-insulin protein composition thus obtained is, as such, suitable as a raw material for, for example, infant formulas and follow-on formulas, because the whey protein it contains is of high nutritional quality and does not require other proteins to supplement the nutritional content.

The cation exchange resin treatment may also be carried out, for example, on skimmed milk or aqueous milk casein, which is a preferred protein formulation. Bovine insulin residues may be removed from the casein solution in a manner similar to milk. However, due to its lower nutritional value than whey protein, casein is not as desirable as the sole source of protein for infant formulas as whey protein.

If it is desired to further reduce the bovine insulin content of the protein composition of the invention, enzymatic hydrolysis and, if appropriate, further hydrophobic chromatography may be employed in its preparation.

The protein concentrate or protein powder obtained in step b) then forms a solution in water at a concentration of 1 to 20%, preferably about 5%. The pH of the solution is adjusted to about 8.5 with, for example, Ca (OH) 2 and the temperature to about 50, followed by the addition of animal or microbial enzymes to hydrolyze it. In particular, it efficiently binds bovine insulin protein chain linkages. Such proteases include e.g. chymotrypsin, subtilisin Carlsberg serine protease, subtilisin BPN 'serine protease, Aspergillus oryzean serine and metalloproteases, papain, Bacillus subtilis 5 -neutral protease, thermolysin, Streptomyces griseus serise protease, and pancreatin. The hydrolysis is allowed to continue for 8 hours. In hydrolysis, bovine insulin is cleaved into peptides of up to five amino acids in length, which do not elicit an immune response and do not contain epitopes contained in the former insulin molecule. The hydrolysis mixture thus obtained is subjected to ultrafiltration to remove non-cleaved large molecules through a dense ultrafiltration membrane, suitably a 2000 D 15 cut-off membrane. The resulting permeate is dried, for example, by spray drying to a powder. The product obtained does not contain bovine insulin.

If it is desired to remove bovine insulin hydrolysis products as accurately as possible from the mixture, the whey protein hydrolyzate obtained above may conveniently be subjected to hydrophobic chromatography as a 10% solution which removes hydrophobic peptides and any bovine insulin hydrolysis products from the hydrolyzate.

; 'This hydrophobic chromatography treatment may

I I

:: 25 Suitable hydrophobic adsorption resin, ·: ··; such as Amberlite XAD-16 resin (Rohm St Haas, France), but can also be made with activated carbon which also • is capable of removing hydrophobic compounds. In the production scale, this chromatographic treatment can be carried out ... 30 practically on a column into which the solution to be treated is passed through a packed adsorbent resin. The solution passing through the pyrolysis solution is collected, evaporated and dried to a powder. The powder thus obtained does not contain the present ·: ··; ' methods detectable amounts of bovine insulin. However, the amino acid composition of the product .35 has changed slightly with respect to the hydrophobic amino acids, so that small amounts of phenylalanine and tyrosine have to be added in the preparation of nutritional products.

Most preferably, bovine insulin is removed from the non-fat proteinaceous material, preferably whey, from cow-5 milk with a strong cation exchange resin, and the liquid proteinaceous material thus treated is concentrated by ultra- and diafiltration. Microfiltration as a pretreatment is preferred since insulin is often adhered to macromolecules such as casein dust or denatured whey protein which can be conveniently removed by microfiltration. If the methods have not achieved a sufficiently low bovine insulin content of the product, it may be further reduced by enzymatic hydrolysis and possibly by hydrophobic chromatography.

Infant formulas are traditionally made from milk, cream, vegetable oil, low-salt whey powder, minerals and vitamins, of which milk, cream and low-salt whey powder contain bovine insulin. Whey protein has a high nutritional value; It is suitable as a sole source of protein for infant formulas and other special nutritional preparations.

"** ί The substantially non-insulin protein composition of the invention can be used as a protein component in infant formulas and other special dietary foods and, for example, as a raw material in drinking milk. This provides a non-bovine insulin formulation and thus does not immunize bovine insulin. · *, Thus increasing the risk of developing IDDM.

The invention also relates to a substantially non-insulin-free infant formula, as well as a substantially insul-non-specific nutritional preparation, characterized in that their protein moiety is a non-fat proteinaceous material derived from cow's milk such as whey, skimmed milk or non-fat milk. , preferably whey from which bovine insulin has been substantially removed.

The process according to the invention for the manufacture of a substantially non-insulin infant formula or any other special nutritional preparation or drinking milk or its raw material is characterized in that the protein component of the product is a substantially non-insulin protein composition according to the invention.

The invention is further described in the following examples.

Example 1 The pH of 6000 ml of fresh cheese whey was lowered with 10% HCl to 5.4. 300 ml of strong cation exchange resin (Amberlite C-20, Rohm & Haas, France) was packed on a 300 ml laboratory-size column and regenerated with 600 ml 17%

NaCl, followed by rinsing the column with 1000 mL of water. 6000 ml of cheese whey (20 column volumes (BV)) was passed through the column at room temperature at a rate of 6 BV / h. Before treatment, whey contained 343 ng / ml insulin, or 68 mg 'insulin / kg of actual protein, after treatment:' no whey was found to have insulin, i.e., the concentration was reduced by 100%. The total protein content of whey was not * · * ' Protein content before treatment was 0.87%, after treatment 0.86%.

The pH was again raised to 6.5 with 10% NaOH, the residue of which was ultrafiltered with 10,000 D cut-off membranes 30 first at a concentration ratio of 10, then diluted to the starting volume and re-filtered. • · · with 12 to give a total concentration ratio of t · 120. The protein concentrate was freeze-dried to a powder containing 90% protein.

35 «· I I« I «<« 12 104457

Example 2 100 liters of cheese whey filtered through microfiltration membranes of 0.1 μιτι are ultrafiltered with 10,000 D cut-off membranes, first at a concentration ratio of 10, then diluted reten-5 to a starting volume and filtered again at a ratio of 12 to 90% retinate. and spray dried to dryness (drying temperatures 180/75 ° C). The untreated cheese whey contained 343 ng / ml or 68 mg / kg of actual protein according to 10 electro spray mass spectrometry analysis. The microfiltered whey contained 324 ng / ml insulin, or 64 mg / kg of actual protein. In the ultra- and diafiltered whey protein concentrate, the insulin content was 21 mg / kg of actual protein 15 by mass spectrometer, i.e., the concentration was reduced by about 69% relative to protein. Conventional ultrafiltered whey protein powder containing 70-80% protein contains about 60 mg / kg of actual protein insulin.

Example 3 20 100 liters of acid whey from the preparation of acid casein were microfiltered through 0.1 μιη: η membranes, respectively. The pH of the whey was 4.5. Microfiltered whey ultra- and whey

t 1 I

Diafiltration was similar to Example 1 to a total concentration ratio of 120. The protein content was then about 90% of the dry matter. Untreated whey contained 320 ng / ml insulin. or 48 mg / kg real: protein, microfiltrated 295 ng / ml or 45 mg / kg real protein and ultra- and diafiltrated whey protein concentrate had 95 ng / ml or 16 mg / kg of 30 real protein as determined by mass spectrometry. · ♦ As a result, the insulin concentration in the treatment was reduced by about 67%.

• · ·

Example 4 • · 60 liters of 50 ° C water were dissolved in 5.040 kg of whey protein powder prepared in Example 1, 11.423 35 kg of vegetable fat blend, 11.232 kg of glucose, 12.260 kg of maltose, DE45, 135 g of vitamin and mineral premix. ta (contains A, D, E, K, B1, B2, B6, B12 vitamins, niacin, folic acid, pantothenic acid, biotin, ascorbic acid, choline, inositol, ferrogluconate, zinc sulfate, manganese sulfate , sodium selenite, copper gluconate) and 70 g of calcium chloride, 300 g of calcium phosphate, 65 g of magnesium sulfate, 125 g of sodium chloride and 620 g of potassium citrate. The dry matter content of the mixture was about 40%. The mixture thus obtained was introduced into a homo-10 generator (150/50 bar) and dried to a powder in a spray dryer with drying temperatures of 180/75 ° C, Lei juped 70/120/30 ° C. The composition, appearance and taste of the product was similar to that of a normal infant formula.

15 Example 5

The whey protein concentrate shown in Example 1 was diluted to 5% with water. The solution was pasteurized at 65 ° C for 20 min and cooled to 50 ° C. The pH was adjusted to 8.5 with 10% Ca (OH) 2. To the mixture was added 6% of 20 protein pancreatin (4 x USP, SPL, USA) - and

Alcalase 0.6 L (Novo Industri A / S; Denmark). During the hydrolysis, the pH of the mixture was maintained at 7.0 by addition of '·' Ca (OH) 2. The mixture was allowed to hydrolyze. "For 8 hours, after which the mixture was heat treated for 5 min at 90 ° C. The mixture was then cooled to 50 ° C and * i ** i ultrafiltered with 2000 D cut-off films and the permeate was collected The permeate obtained was spray-dried to a powder No insulin was detected in the hydrolyzate Example 6 The hydrolyzate obtained in Example 5 was dissolved in a 10% solution · · 30 ml of Amberlite XAD-16 resin (Rohm & Haas). packed in a laboratory scale column regenerated with 60 mL 4% NaOH, rinsed with 1000 mL: ** water and regenerated with 60 mL 4% HCl and rinsed with water until the pH of the permeate was more than 5. Hydrolyzate was passed through a column of 1700 ml of resin, corresponding to 567 g of dry powder hydrolyzate / 100 ml of resin, The solution which had passed through was recovered and freeze-dried as no powder was detected in the hydrolyzate.

Hydrolyzate was used as the sole source of protein for a special dietary product for milk allergies.

Example 7 8.670 kg of whey protein hydro-10 lysate powder prepared according to Example 6, 10.466 kg of vegetable fat blend, 16.058 kg of glucose, 5.233 kg of maltodextrin (DE 21), 135 g of vitamin and mineral blend (containing A-, D-) were dissolved in 60 liters of 50 ° C water. E-, K-,

Vitamins B1, B2, B6, B12, niacin, folic acid, pan-totenic acid, biotin, ascorbic acid, choline, 15 inositol, ferrogluconate, zinc sulphate, manganese sulphate, sodium selenite, potassium gluconate, , 70 g of magnesium sulfate, 165 g of sodium chloride and 620 g of potassium citrate, 1 g of L-tyrosine and 2 g of L-phenylalanine. The dry matter content of the mixture was about 40%. The mixture thus obtained was introduced into a homogenizer (150/50 bar) and dried to a powder in a spray dryer with drying temperatures of 180/75 ° C, in a fluidized bed 70/120/30 ° C. The composition, appearance and taste of the product was as follows: *: * of a special dietary preparation for allergy sufferers, such as milk v 25.

· * · .. Example 8

A strong cation exchange column (30 mL of resin) was regenerated as in Example 1. 600 mL of fat-free milk was adjusted to pH 5.4. The pH-adjusted milk was passed through a column at room temperature as in Example 1, whereby a portion of the calcium was excreted, but also a significant proportion of the insulin. According to RIA analysis (mass spectrometer not available for milk), untreated milk contained bovine insulin 26 μΐυ / ml and treated 17 μΐυ / ml.

I «« tl • «•« <· «104457 15

Insulin concentration was reduced by about 35% during treatment. The treatment was not sufficient to completely remove the insulin from the milk, but the example illustrates that the method is also suitable for treating milk raw materials other than whey.

5 Example 9

The cation exchange resin (30 ml) was regenerated as in Example 1. A 3% solution of sodium caseinate in water was prepared. The pH was lowered to 5.5 with dilute HCl. The caseinate solution was pumped through a cation exchange resin at room temperature like milk in Example 8. Prior to treatment, the caseinate solution contained 26 μΐ / mL of insulin according to RIA analysis and 10 μΐϋ / mL of treated solution. Insulin concentration was reduced by about 62% during treatment.

• · • · • · * ·

»T I

• • • • • • • • • • • • • • • («

I I I

«I

• «• · · t ·«

Claims (13)

104457 Claims 1. A substantially non-insulin protein composition useful as a protein component of a nutritional preparation or as a raw material for drinking milk, characterized in that it is a non-fat proteinaceous material derived from cow's milk, such as whey, skimmed milk or bovine insulin-free. A process for the preparation of a substantially insulin-free protein composition, characterized in that: a) the liquid non-fat proteinaceous material derived from cow's milk is subjected to an insulin deprotection treatment, which may be first clarified and subsequently conducted at a pH of 5.2 to 5.6; Through a column packed with a strong cation exchange resin regenerated to the Na + or K + form, b) the liquid proteinaceous material obtained in step a) is concentrated by ultra- and diafiltration using membranes from 6,000 to 20,000 D cut-off membranes, and the protein concentrate obtained is evaporated and optionally drying to a protein powder, c) optionally. 1) forming a solution of the protein concentrate or protein powder obtained in step b) in a water having a protein content of 1-20%, 2) hydrolyzing the protein solution obtained in step cl). - • · ·; enzymatically with proteases and: 3) ultrafiltration of the protein solution hydrolyzed in step c2), and d) optionally subjecting the pro-, protein hydrolyzate obtained in step c) to hydrophobic chromatography 17 104457 and evaporating the solution obtained in step c) or d) and powder. Process according to claim 2, characterized in that whey, skimmed milk or casein solution, preferably whey, is used as the liquid non-fat proteinaceous material in step a). Process according to Claim 2 or 3, characterized in that in step a) the liquid non-fat proteinaceous material is clarified prior to treatment with the cation exchange resin by filtration through microfiltration membranes, which are 0.05 to 1.4 micrometer membranes, preferably 0, 1 micrometer membranes. A process according to claim 2 or 3, characterized in that in step a) the liquid non-fat proteinaceous material is clarified before treatment with the cation exchange resin by treatment with ultrafiltration membranes, preferably 50,000 to 20,200,000 D cut-off films. Process according to claim 2 or 3, characterized in that in step a), the liquid non-fat proteinaceous material is clarified prior to the cation exchange resin treatment, preferably by centrifugation at 1000 to 10,000 rpm. The process according to any one of claims 2 to 6, characterized in that in step b), membranes of 10 · · · · 000 D-cut-off are used for edge and diafiltration. films. The me- according to any one of claims 2 to 7: * · *; A process characterized in that, in step b), the ultra- and diafiltered protein concentrate is formed into an aqueous solution of 1-20%, preferably about 5%, which is enzymatically hydrolyzed with proteases, such as pancreatin and alkalase. 104457 Process according to claim 8, characterized in that the protein hydrolyzate obtained in step c) is subjected to hydrophobic chromatography, whereby it is treated with activated carbon or, preferably, a hydrophobic polystyrene-based adsorption resin. Use of a protein composition according to claim 1 or a method according to any one of claims 2 to 9 as a protein component of infant formula 10 or other special nutritional preparation or as a raw material for drinking milk. 11. A substantially non-insulin infant formula, characterized in that its protein moiety is a non-fat proteinaceous material derived from cow's milk, such as whey, skimmed milk or casein solution, preferably whey from which bovine insulin has been substantially removed. 12. A substantially non-insulin specific nutritional preparation, characterized in that its protein moiety is a non-fat proteinaceous material derived from cow's milk, such as whey, skimmed milk or casein solution, preferably whey from which bovine insulin has been substantially removed. ; ·. 13. A process for the manufacture of a substantially non-insulin-containing infant or other special nutritional preparation, or a raw material for drinking milk, characterized in that the protein component of claim 1 or * is used in the manufacture of the product. · A substantially insulin-free ptotein composition produced by the method of any one of claims 2 to 9. • · · • «· • · 19 104457