New infant formulas from cows' milk supplemented ...

ISSN: 0378-2700 NEW INFANT FORMULAS FROM COWS' MILK SUPPLEMENTED WITH MODIFIED WHEY BY Sania M. Abdou; M. B. El-Alfy; M. E. Shenana and Dalia G. Gemiel Food Sci. Dept., Moshtohor Faculty of Agric., Benha Univ., (Accepted, 12/5/2014) SUMMARY

Infant

formulas known as "baby foods" are substitutes for human breast milk usually based on cow's milk and designed for infant consumption. Legislations providing the regulatory frame for the composition of infant formula have been considered in this study. β- Lactoglobulin was removed from rennet whey before its use in the developed formulas. Three proposed formulas were prepared by mixing cows' milk with modified whey at the ratios of 1:1 (Formula 1), 1.5 : 2 (Formula 2) and 2: 3 (Formula 3) respectively. The lactose content of these formulas was adjusted to ~ 7% with lactose powder, fat content to 3.5% with corn oil and 0.5%lecithin was added as an emulsifier. All treatments were heated to 85°C, cooled immediately and vitamins were added. The chemical composition of all proposed formulas was similar to that of human milk except for the low protein content in human milk. Human milk did not clot with rennet without or with adding calcium chloride while the formulas were rennet clotted at different times. The enzymatic digestibility revealed variations between the different formulas. Polyacrylamide gel electrophoresis cleared that the protein patterns of the different formulas were more or less similar to that of human milk. The adequacy of the proposed formulas was ascertained by comparing the growth of rats fed on the prepared products with that of breast–fed rats. The nutritional parameters of the different groups of rats revealed that the body weight gain food efficiency ratio were higher in formulas- fed rats than the control and that the best results were obtained with formulas 3.

INTRODUCTION Human milk is considered to be the best source of nutrition for infants (Picciano, 2001) being almost a complete food and is species-specific. However, milk based formulas and milk substitutes are generally recommended in case of insufficient mothers' milk to satisfy the infant needs..

Major producers of infant formulas developed their brands with specific compositions. The International legislation organizations (such as CODEX), provide the regulatory frame for the composition of infant formula and follow-up formula.

Infant formulas are unique because they may be the only source of nutrition for many infants during the first 4 to 6 months of life.

Infant Formula manufacturers are continuing to research into the modifications necessary to make cows' milk similar to human milk, however, there are so many subtle differences.

Recommendations from the department of Health in the UK and the European Society of Pediatric Gastroerology, Hepatology and Nutrition emphasized that in the absence of a lactating mother, feeding on formula milk should be continued until 12 months of infant age.

When food is designed, not only texture and nutrients must be considered but also, to some extent, protection from diseases. This should be emphasized especially for artificial infant formula. The resistance of breast-fed infants to diseases is generally ascribed to lactalbumin, immunoglobulin,

Egyptian J. Dairy Sci., 42: 105-118 (2014)

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lactoferrin, lysozyeme, lactoperoxidase, the bifidus factor and interferon (Packard et al., 1982). From this point of view, bovine milk whey can be potential source to provide some of these compounds especially immunoglobulin for infant formulas. The direct utilization of whey proteins would increase β-lactoglobulin in infant formula, an allergenic milk protein, which is absent in human milk (Lebenthal, 1975). Therefore, it is desirable to eliminate β-lactoglobulin and to concentrate immunoglobulins and α-lactalbumin in whey before its use in infant formulas. Also, the use of whey would simulate the casein to whey proteins ratio in infant formulas to that in

human milk (Casein N / whey N= 40/60). Thus, most infant formulas are whey predominant. The purpose of technological development of infant feeding is to improve on the efficacy of the diet to maintain the life, health and well-being of infants. Whey proteins are known to be of high nutritional value, rich in essential amino acids and of good digestibility (Hambraeus, 2003). Therefore, this study was carried out to propose infant formulas from cows' milk supplemented with modified whey.

MATERIALS AND METHODS Materials: - Fresh cows' and buffaloe's milk were obtained from the herds of the Faculty of Agriculture, Moshtohor, Benha Univ. - Early morning bulk human milk samples (from 10 individual women) were obtainned from mothers of Child Care Center, Touhk Qalyoubia Governorate. The ten samples were mixed together to form a large bulk sample. - Pure grade lactose was obtained from ElNasr pharmaceutical Co. Adwic, Egypt. Calf rennet was obtained from CHR. Hansen's Laboratories, Copenhagen, Denmark. - Peptic & tryptic Enzymes HOG stomach were obtained from laboratories chemical and reagent, LTD. Netherland. - Mixture of fat and water soluble vitamins and lecithin were obtained from Lucas Meyer GmbH, Hamburg, Germany. Methods: Preparation of modified rennet whey: Mixed cows' and buffaloes' milk (1:1) was coagulated using rennet powder, curd was cut into cubes 1×1cm, the released whey was collected and modified by removing of β-lactoglobulin according to the method of Kaneko and Nakai (1985). Preparation of different infant formulas: Modified rennet whey was added to the standardized cows' milk to

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make three different formulas coded as follows: Formula 1: 1 part of cows' milk: 1 part of the modified whey. Formula 2: 1.5 parts of cows 'milk: 2 parts of modified whey. Formula 3: 2 parts of cows' milk: 3 parts of modified whey. The composition of the different formulas was adjusted to ~ 3.5% fat with addition of corn oil and lactose content to ~ 7.0% with adding lactose powder. Lecithin (0.5%) was added to the different formulas as an emulsifier, and mixed water and fat soluble vitamins were added to give the following respective contents per one litter of the final mixture (Table 1) as recommended by Codex (1981). The mixtures were then heated to ~85°C and cooled rapidly to room temperature. Three replicates were prepared from each formula and analyzed for chemical composition, some properties and biologically evaluated. Chemical Analysis: The total solids, Ash, Fat, lactose contents and titratable acidity of cows'and human milks, whey and prepared infant formulas were determined according to IDF, (1987), AOAC (1995), IDF, (1996), AOAC, (1990) and Nickerson et al. (1975). The total

New Infant Formulas From Cows' Milk

nitrogen (T.N), non protein nitrogen (NPN) and non casein nitrogen contents (NCN) were determined according to the method of IDF, (2001). The total Albumin nitrogen (AN), βlactoglobulin and α-lactoalbumin were determined as outlined by Fox and Morrissey, (1976). The other nitrogen fractions i.e (proteosepepton, lactoferrin, lysozyme immunoglobulin's…. etc) were calculated by differrence as follows: Other nitrogen fractions = (Whey proteins- Total albumin). pH values were measured using pH meter JENCO model 1671, USA. Table (1): The Added of vitamins to different prepared formulas. Vitamin A 2000 I.U. Riboflavin 0.2mg Vitamin D 400 I.U. Pyridoxine 0.4mg Nicotinamide 2.0mg Thiamine 1.0mg Vitamin B12 1.0mg Vitamin C* 50mg *added after heat treatments.

Determination of minerals content: Zinc, manganese, iron, cupper and magnesium were determined using the atomic absorption spectrophotometer Perkin Elmer model 2380, according to the procedure of AOAC (1990). Potassium & calcium and sodium were determined using a Beckman flame photometer according to the method of Jackson (1967). The phosphorus and chloride were determined by the method of Olsen and Sommers (1982). Digestibility of milk with proteolytic enzymes: Peptic, tryptic and renneting digestion of milk and prepared formulas were determined according to the method of Hawk and Summerson (1953). The soluble digestion products were determined by the method of Lowry et al. (1951). Estimation of the rennet coagulation time (RCT) of cows', human

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milks and prepared infant formulas was carried out according to the method of Joseph and Ashworth, (1970). Gel electrophoresis analysis: Polyacrylamide gel electrophoresis performed using the method of Laemmeli, (1970). Calculation of Caloric value: The caloric values were calculated according to the basis that the caloric values produced by one gram of protein, carbohydrates and fats give 4Kcal, 4Kcal and 9Kcal, respectively (FAO/WHO, 1985). Statistical analysis Statistical analysis of the obtained data was carried out using SAS procedure guide (SAS, 2004) Nutritional adequacy of prepared infant formulas: The nutritional adequacy of the prepared infant formula was carried out biologically using the basal diet recommended by (AOAC 1998) as follows: Thirty female albino rats (90-120g) were fed on basal diet (20g daily per rat) for one week (adaptation period) and then divided randomly into five groups (6 rats / each). The first group was fed on basal diet throughout the experimental period (7 weeks) and considered to be as negative control. The second group was fed on basal diet containing human milk for 7 weeks according to the chemical composition mentioned in table 2 and considered as positive control. The other three groups were fed for 7weeks on basal diet supplemented with different proposed infant formula milk (F1, F2 and F3) as described in Table 2. Diet and water were provided ad libitum. Fresh diet was supplied every day to avoid extra fermentation. The weight of each rat was recorded 3 times weekly and the residues from the fodder were weighed daily. At the end of this experimental, total food intake, body weight gain and food efficiency ratio (Body weight gain/ total food intake) were calculated.

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Table (2): Experimental rat groups. Animal Group diet *

Experimental diets (per 6 rats)

Negative control (Basal diet )

120g basal diet*

Positive control (Basal diet+ Human milk)

67g basal diet + 53g Human milk

Formula1

67g basal diet + 53g Infant milk formula 1

Formula 2


Formula 3


*

Basal diet: As recommended by AOAC 1998

RESULTS AND DISCUSSION Chemical composition of rennet whey and modified whey Table (3) shows that the average total solids (T.S), fat, protein, lactose and ash contents of the rennet whey were 6.80, 0.59, 0.86, 4.87 and 0.50g/100ml, respectively. The minerals content of whey are shown in Table (4). Various elements in milk bind to different compounds. They are present in a colloidal form bound to casein and colloidal calcium phosphate. So, it could be expected that the whey will contain lesser amount of calcium and phosphate than original milk. The acidity and pH of the whey recorded 0.13% and 6.47, respectively. On the other hand, modified whey contains 5.93, 0.05, 0.64, 4.66 and 0.64 g/100ml of TS, fat, protein, lactose and ash, respectively. The pH of modified whey was found 7.84. These results are in agreement with Marshall (1982) and Pescuma et al. (2008). With regard to minerals content, Table 4 revealed that there was a noticeable decrease in modified whey than original whey except Fe and Cu which recorded higher level in the modified whey than the original one due to the use of FeCl3 in removing of βlactoglobulin. No significant differences were found between Cl and Mg in both two types of whey.

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Nitrogen distribution of rennet whey and modified rennet whey: From the obtained data (Table 5) it is clear that the T.N, N.P.N, C.N, WPN and others N fraction of whey recorded 134, 29, 30, 75 and 19.9 mg/100ml respectively. After modification of whey these values were changed to 101, 46, 3, 52 and 29 mg/100ml, in the same order. The β- lactogloulin was reduced from 35.1 to 2mg/ 100ml. α-lactalbumin and other whey fractions became the predominant protein components as in human milk. The concentration of other whey fractions, which are mainly immunoglobulin and lactoferrin were 29mg/100ml in the modified whey. These changes can be considered essential to make modified whey more suitable to be used in the proposed infant formulas. Thus, the new infant formulas with addition of modified whey may be advantageous immunologically, due to the binding capability of IgG to staphylococci protein A and the antiviral activity of IgG. (Skvaril, 1983). Data illustrated in Fig (1) show the protein bands compared with marker standard. It could be demonstrate that the modified rennet whey contains less amount of βlactoglobulin compared with the rennet whey. From such analysis it could be conclude that we can use the modified whey in the preparation of different formulas without increase in β-lactoglobulin to be easy to digest by infants.


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Table (3): Chemical composition of rennet whey and modified rennet whey (g/100ml). Component Total solids Fat Total protein Lactose Ash Acidity pH

Rennet whey 6.80 0.59 0.86 4.87 0.50 0.13 6.47

Modified rennet whey 5.93 0.05 0.64 4.66 0.64 Traces 7.84

Table (4): Minerals content of rennet whey and modified rennet whey mg/100ml. Sample Ca P Na K CL Fe Mg Zn Cu Mn Rennet whey 66a 95.2a 53a 145.8a 11.4a 0.042b 7.00a 0.434a 0.0128b 0.032a Modified whey

rennet

50b 65.2b 40.2b 87.4b

11.2b 0.0862a 7.00a 0.294b

0.023a

0.013b

*Values with the same latter in each column are not- significant differences at the level of 0.0001.

M

1

2

Fig (1): Poly acrylamide gel electrophoresis of rennet whey lane (1) and modified rennet whey lane (2) compared with marker lane (M).

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Table (5): Nitrogen distribution of rennet whey and modified rennet whey (mg/100ml). Total albumin % Whey Others (N) Type of whey T.N N.P.N P.N C.N N.C.N C.N/ (P.N) (N) WPN α-lacto β-lacto albumin globuli a Rennet whey 134a 29b 105a 30a 104a 75a 40a 20b 35.1 19.9b n Modified rennet whey

101b

46a

55b

3b

98b

52b

5.77b

21a

2b

29a


Chemical composition and calculated energy value. Data illustrated in Table (6) show that the TS contents were 12.64, 12.62, 12.65 and 12.42g/100ml for formula 1, 2, 3 and human milk, respectively. The obtained results of fat content were 3.50, 3.57, 3.50 and 4.03g/100ml for the same previous order. The fat content of the proposed formulas was almost the same due to adjusting of fat to ~ 3.5% with corn oil. The protein contents in the prepared F1-F3 formulas were higher than that of human milk as it recorded 1.86, 1.79, 1.63 and 1.2 g/100ml in order. The protein /energy ratio for the proposed formulas was 2.78, 2.67 and 2.43 while it was 1.79 in human milk. These results are in accordance with Räihä et al. (2002) and with International Recommendations. The calculated energy value of human milk was higher than the prepared formulas which recorded 67.06, 67.61, 66.32 and 69.07 kcal/100ml, of formula 1, 2, 3 and human milk, respectively mainly due to the higher fat content of human milk. The obtained results are within the range recommended by FDA (1985), ESPGHAN (1991), Räihä et al. (1986), Räihä (1994), LSRO (1998), SCF (2001a) and Räihä et al. (2002). The FDA, 1985), ESPGAN, (1977), Codex Alimentrius (1981) and the Committee on Nutrition of the American Academy of Pediatrics (1976) specified the lower limit of protein in infants formula to be 1.8g/100kcal. This level of protein is within the safe level (Joint FAO/WHO/ UNU Expert Consultation, 1985). Fomon (1991) recommended a minimum protein level of 2.2g/100 kcal in

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formulas for infants less than 3 mos and a content of 1.6g/100 kcal for infants over 3 mos. Similar protein level were obtained in this study. The lactose level, was increased in the prepared formulas to simulate human milk. So, the lactose content was adjusted to ~7g/100ml by adding lactose. The lactose recorded 7.03, 7.08, 7.07 and 7.00g/100ml for formulas 1, 2, 3 and human milk, respectively. The ash content was also higher in the prepared formulas than in human milk being 0.32, 0.32, 0.31 and 0.23g/100ml for formula 1, 2, 3 and human milk respectively. Nitrogen distribution. Infant formula contains whole milk proteins which may be make allergenic to the infants. Modified whey was used in infant formula to reduce the risk of allergenic. Formulas predominantly contained whey as a source of protein is considered to be more similar to breast milk in terms of protein composition (Thorkelsson et al., 1994). Table (7) recorded the nitrogen distribution of the prepared formula. The T.N was 291, 283, 255 and 183mg/100ml for formula 1, 2, 3 and human milk, respectively. N.P.N recorded 35, 38, 42.0 and 39mg/100ml for the same pervious order, which represents 12.02, 13.42, 16.47 and 21.53% of the T.N. The casein N recorded 132, 123, 107 and 61mg/100ml for formula 1, 2, 3 and human milk, in order. On the other hand, whey protein was 124.0, 122.0, 106.0 and 83.0mg/100ml for formula 1, 2, 3 and human milk, representing 42.61, 43.10, 41.56 and 45.36% of the T.N respectively.


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Table (6): Chemical composition and calculated energy value of proposed infant formulas and human milk (g/100ml) Protein/ Calculated Energy Energy value Type of milk T.S Fat Protein Lactose Ash ratio kcal/100ml Formula 1 Formula 2 Formula 3 Human milk

12.64a 12.62a 12.65a 12.42b

3.50c 3.57b 3.50c 4.03a

1.86a 1.79c 1.63b 1.20d

7.03a 7.08a 7.07a 7.00ab

0.32ab 0.32a 0.31b 0.23c

2.78a 2.67b 2.43c 1.79d

67.06c 67.61b 66.32d 69.07a


α-Lactoalbumin and β-lactoglobulin of the prepared formula recorded 39.1, 43.0, 40.2 and 35.0 & 18, 15, 10 and traces for formula 1, 2, 3 and human milk in sequence. Other nitrogenous components in the proposed formulas recorded 66.69, 60.2, 55.80 and 48.0mg/100ml, in the same previous order. Electrophoretic pattern Fig (2) shows that the protein bands of the proposed formulas matched with marker standard. Based on the intensity of the separated bands the prepared formulas contained higher amounts of β-lactoglobulin which was not detected in human milk. On the other hand, the prepared formula contains higher amount of lactalbumin than human milk due to adding of modified rennet whey during preparing of these formulas. Comparing the casein bands of the prepared formulas with the human milk it was clear that the prepared formulas contained condensed bands of caseins than human milk. From such analysis it could be conclude that the pattern of protein of prepared

formula was close to the protein pattern of human milk suggesting that these formulas to be easily digested by infants. Mineral contents. Table (8) shows that the Ca contents of formula 1, 2, 3 and human milk were 50, 55, 49 and 30mg/100ml respectively and P contents were 24, 26, 28 and 14mg/100ml in the same order. It was obvious that the Ca:P ratios were 2.1, 2.1, 1.75 and 2.1 in formula 1, 2, 3 and human milk, successively. The Ca:P ratios of formula 1 and 2 were the same as in human milk. However, The Ca:P ratios of all formulas are within the range given by most of the nutrition commission as they recommended the level namely: 1.1 to 2.0. The values of K, Na and Cl in the proposed formula were almost similar to that of human milk as it recorded 58, 53, 55 and 55mg/100ml for K; 15, 16, 18 and 15 for Na and the Cl contents were 43, 47, 40 and 43mg/100ml for formula 1, 2, 3 and human milk, respectively. These results are in accordance to that of SCF (1993), Wack et al. (1997) and LSRO (1998),

291a 283b 255c 183d

35.0d 38.0c 42.0a 39b

256a 245b 213c 144d

132a 123b 107c 61d

159ab 160a 148b 121c

Others (N)

42.61c 43.10b 41.56d 45.36a

β-lacto globulin

124a 122b 106c 83.0d

α-lacto albumin

% WPN/T.N

Formula 1 Formula 2 Formula 3 Human

Whey (P.N)

Table (7): Nitrogen distribution of proposed infant formulas and human milk (mg/100ml). Total albumin (N) Type of T.N N.P.N P.N C.N N.C.N milk 39.1c 18a 66.9a a b 43 15 62b 40.2b 10c 55.8c d d 35.0 Traces 48.0d


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M

1

2

3

4

Fig (2): Poly acrylamide gel electrophoresis of human milk lane (1), formula 1 lane (2), formula 2 lane (3) and formula 3 lane (4) compared with the marker lane (M) The Mg, Fe, Zn, Cu and Mn were found as trace elements and were within the range given by Sievers et al. (1992), SCF (1993), LSRO (1998), AAP (1999), Hernell and Lönnerdal (2002), SCF (2001b), Abrams et al. (2002) and SCF (2003). The objective of replacing part of cow's milk fat with corn oil is to increase the poly unsaturated fatty acids, particularly linoleic acid which present in lower proportion in cows' milk than that of human milk and therefore, improve the absorption of fat by infant. Poor fat absorption may reduce the absorption of calcium and often cause hypocalcaemia (Widdowson, 1973).

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pH value, acidity and rennet coagulation time (RCT) Data presented in table (9) show the pH values and acidity of prepared infant formulas as well as human milk. The pH values recorded 6.45, 6.66, 6.60 and 7.10 while the acidity recorded 0.14, 0.15, 0.14 and 0.08% for formula 1, 2, 3 and human milk, successively. From the obtained results it could be concluded that the pH of the prepared formula is more or less the same and it was lesser than that recorded for human milk. Same observations were found for the acidity values but in the opposite direction. These results are in agreement with Abd El-Hakeem (1994).


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Table (8): Mineral contents of proposed infant formulas and human milk. Minerals mg/100ml Type of milk Ca

P

Ca : P K Na

Formula

50b

24c

2.1a

58a 15c

Formula 2

55a

26b

2.1a

53c 16b 47a

Formula 3

49c

28a

1.75b 55b 18a

40c

Human milk

30d

14d

2.1a

43b 0.055d 5.5a

55b 15c

Cl

Fe

Mg

Zn

Cu

Mn

0.47b

0.05a

0.0062c

0.062c 5.5a

0.46c

0.042c

0.0072b

0.085a 5.2c

0.52a

0.047b

0.0083a

0.30d 0.040d

0.0062c

43b 0.076b 5.3b


Rennet coagulation time (RCT). Table (9) shows the rennet coagulation time either without or with adding CaCl2. It was clear that the time of coagulation of the prepared formulas was long in the absence of CaCl2 as it recorded 215, 227 and 233sec. for formula 1, 2 and 3, respectively while human milk was not coagulated. Similar trend was found in case of adding CaCl2 as a coagulation activator, in the same time the rennet coagulation time almost reduced to 50% of the time and recorded 109, 111 and 120 sec for formula

1, 2, 3 while there was no clot in human milk. It was clear that either without or with adding CaCl2; human milk was uncoagulable by rennet and formed no clot. This is mainly due to lower casein and calcium content and higher pH and whey protein contents. The obtained results comply with the obtained results of some market infant formula (Shenana, et al., 2013). Also, the results are in accordance with Abd ElHakeem (1994).

Table (9): pH value, acidity and rennet coagulation time (RCT) of proposed infant formulas and human milk. Rennet coagulation time % (RCT) (sec.) Type of milk pH Acidity without CaCl2 with CaCl2 Formula 1 Formula 2 Formula 3 Human milk

6.45b 6.66b 6.60b 7.10a

0.14a 0.15a 0.14a 0.08b

215c 227b 333a No clotd

109c 111b 120a No clotd


Digestibility of proposed infant formulas. The rate of digestion by proteolytic enzymes of prepared infant formulas compared with the human milk is presented in Table (10). Formula No.1 showed a rate of peptic digestion similar to that of human milk as both recorded 134.62% increase in the O.D at the end of the digestion time. However, formula 2 and 3 showed slower

peptic digestion than human milk as they recorded 87.5 and 84.48% in the same period. The rate of tryptic digestion took the same trend of peptic digestion where formula 1 was similar to human milk, while formula 2 and 3 showed slower digestion. The rate of rennet digestion of the prepared formula was lower in formula 1 than the human milk. Formula 2 was more close to human milk.

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Table (10): Rate of peptic, tryptic and rennet digestion of proposed infant formulas and human milk. O.D / 500 mu /mg protein Time (min)

Type of milk 0

30

60

90

120

150

180

0.110a

0.122a

Net % increase increase

Peptic digestion Formula 1

0.052c

0.058c

0.075a

0.082a

0.093a

0.070 134.62a

Formula 2

0.056b

0.062b

0.069d

0.075c

0.083d 0.096bc 0.105ab

0.049

87.50b

Formula 3

0.058a

0.064a

0.071c

0.077b

0.085c

0.098b

0.107b

0.049

84.48c

Human milk

0.052c

0.058c

0.073b

0.082a

0.090b

0.110a

0.122a

0.070 134.62a

Tryptic digestion Formula 1

0.104c

0.110b

0.127a

0.134a

0.145a

0.162a

0.174a

0.07a

67.31b

Formula 2

0.107b

0.113a

0.120c

0.126d

0.134d

0.147d

0.156d

0.049c

45.79d

Formula 3

0.111a

0.113a

0.127a

0.131c

0.144b

0.151c

0.167c

0.056b

50.45c

Human milk

0.102d

0.108c

0.125b

0.132b

0.143c

0.160b

0.172b

0.07a

68.63a

0.115d 0.121cd 0.131c

0.142c

0.057d

67.06c

Rennet digestion Formula 1

0.085c

0.097c

0.104c

Formula 2

0.081c

0.099b 0.107bc 0.119b

0.122c

0.135b

0.146b

0.065b

80.25b

Formula 3

0.088b

0.097c

0.108b

0.117c

0.125b

0.135b

0.146b

0.058c

65.90d

Human milk

0.099a

0.105a

0.123a

0.135a

0.147a

0.154a

0.184a

0.085a

85.85a


Evaluation of the nutritional adequacy of the proposed infant formulas To evaluate the nutritional adequacy of experimental formulas differing in their protein fractions, healthy rats fed these formulas were compared with rats either fed on human milk or fed on conventional basal diet for growth. It is generally assumed that the nutritional needs of the animals are met when it shows normal growth. Growth performances of healthy infants were shown by Fomon et al. (1986) to be a very sensitive indicator of protein and amino acid adequacy of infants in addition to nitrogen balance.

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Changes of body weight (g) of rats. Data in Table (11) clear the changes in the body weight (g) of rats weekly and up to 7 weeks. From such data it could be concluded that during the first week and all over the experimental time the growth rate of the rats fed on the prepared formula was higher compared with the control groups (either negative or positive). The obtained data reflect the high body gain (g) (Table 12) during the experimental as it recorded 42.10, 52.14, 68.0, 69.60, and 77.03g for negative control, positive control, formula 1,2 and 3, respectively. On the other hand, the food efficiency ratio was higher in case


of prepared infant formula (Table 12) than the control. Nutritional parameters of rats as affected by feeding on different diet. The prepared infant formulas have been tested for their biological effects on the growth factors. Data in Table (12) show the nutritional parameters of different experimental groups of rats. The body weight gain recorded 42.10, 52.14, 68.00, 69.60 and 77.03g for

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negative control, positive control, formula1, 2 and 3, respectively vs food intake 111.80, 110.9, 111.9, 111.6 and 112.0g in the same sequence. The prepared formulas cleared high food efficacy ratio higher than the control groups whereas they were 0.38, 0.47, 0.61, 0.62 and 0.68 for negative control, positive control, formulas 1, 2 and 3, respectively. This may be due to constitute of the formula i.e. whey proteins which easy to digest. This was clear from the increase of food efficiency ratio with increasing added whey protein.

Table (11): Changes of body weight (g) of rats weekly as affected by feeding on different diet. Animal Group diet Negative Positive Formula 1 Formula 2 Formula 3 Control* Control** Feeding time (weekly) 1St 7.17 9.50 10.00 10.50 11.67 2nd

9.50

11.50

14.67

16.17

14.17

3rd

4.83

6.33

10.83

11.00

11.83

4Th

5.50

6.83

8.17

9.67

10.00

5Th

6.33

7.67

10.00

8.67

12.67

6Th

3.33

4.33

4.83

4.33

9.17

7Th

6.00

6.00

9.50

9.33

7.50

* Negative control: Fed on basal diet. **Positive control: Fed Basel diet with human milk.

Table (12): Nutritional parameters of rats as affected by feeding different diet. Animal Group diet Negative Positive Formula 1 Formula 2 Control* Control** Characters

Formula 3

Initial Body weight (g)

105.50

109.16

114.0

106.00

104.80

Finial Body weight (g)

147.60

161.30

182.0

175.60

181.80

Body weight Gain (g)

42.10

52.14

68.0

69.60

77.03

Total food intake (g)

111.80

110.9

111.9

111.60

112.00

Food intake (g/day)

2.66

2.63

2.66

2.65

2.66

Food efficiency ratio

0.38

0.47

0.61

0.62

0.68

*Negative control: Fed on basal diet. ** Positive control: Fed on Basel diet with human milk.

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Laemmeli U.K. (1970). Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nuture 227:680. Lawry, O.H., Rosehrough, N. J., Forr, A. L., and Randal, L., R.J. (1951). "Protein measurement with the Folin–Phenol reagent ". J. Bio. Chem., 193:265. Lebenthal, E. (1975). Cows' milk protein allergy. Pediatr. Clin. N, Amer. 22:827. Lonnerdal, B.; Keen C. L. and Harley, L. S. (1981). Iron, cupper, zin and manganese in milk. Ann. Rev.Nutr.1, 149. LSRO (1998). (Life Sciences Research Office). LSRO Report: Assessment of nutrient requirements for Infant formulas. Center for Food Safety and Applied Nutrition Food and Drug Administration Department of Health and Human Services, Washington. Marshall, K.R. (1982). Industrial isolation of milk proteins: whey proteins, in developments in dairy chemistry, Vol.1: proteins,. (ed. P. F. Fox), Applied Science Publishers, London, P. P. 339. Nickerson, T.A. Vujicic I. F. and lin A.Y. (1975). Colorimetric estimation of lactose and its hydrolytic products .Journal of Dairy Science Vol. 59 No. 3, 386. Olsen, S.R. and Sommers, L.E. (1982). "phosphorus " pp.403. in A.L. page et al.(Eds.). Method of soil analysis, Part 22nd Ed., Amer. Soc. Madison, Wisconsin, U.S.A. Packard, S.V. Howard, A. Morris (1982). Effect of processing on whey protein functionality. Journal of Dairy Science V67 (11): 2723. Pescuma, M., Maria-Herbet, E., mozzi, F. DeValdez, F. G (2008). Whey fermentation by thermopile lactic acid bacteria: Evaluation of carbohydrates and protein content. Food Microbiology 25: 442. Picciano, M.F. (2001). Nutrient composition of human milk. Pediatric Clinics of North America, 48:53. Räihä, N. Minoli, I. and Moro, G. (1986). Milk protein intake in the term infant. I. Metabolic responses 75:136. Raiha, N.C.R., Fazzolari-Nesci, A. Cajozzo, C. Puccio, G. Monestier, A. Moro, G. Minol., I. Haschler, E. Bachmann, C., Van't

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‫خلطات جديدة مقترحة أللبان األطفال من اللبن البقري ومدعمة بالشرش المعدل‬ ‫ـــــ‬ ‫تم تحضٌر الشرش فً المعمل بتجبن خلٌط من اللبن البقري والجاموسً ( ‪ ) 1:1‬بالمنفحة بحٌث ٌتم‬ ‫التجبن فً ‪ 30‬د‪ .‬بعد تصفٌة الشرش تم تعدٌله بإزالة مركب ‪ β -lactoglobulin‬تم تحلٌل الشرش المعدل بعد‬ ‫إزالة ‪ β -lactoglobulin‬وإتضح انه خالً منه تقرٌبا وكانت نسبة األمالح معظمها منخفضة عنها فً‬ ‫الشرش غٌر المعدل مما ٌقارب وجودها فً لبن االم‪.‬‬ ‫أ ‪ -‬إستخدم الشرش المعدل في تكوين خلطات مقترحة من البان االطفال‪.‬‬ ‫تم تحضٌر ثالث تركٌبات أو خلطات من الشرش المعدل بعد خلطه مع اللبن البقري وإضافة‬ ‫الالكتوز لتصل النسبة الً ‪ %7‬والدهن الً ‪ %3,5‬باستخدام زٌت الذرة واللٌسٌثٌن كمستحلب للدهن بنسبة‬ ‫‪ %0,5‬مع لبن االم كنترول على النحو التالً ‪:‬‬ ‫‪ 1:1= F1‬لبن بقري الً الشرش المعدل‪ 2:1,5= F2 .‬لبن بقري الً الشرش المعدل‪ 3:2 = F3 ..‬لبن‬ ‫بقري إلً الشرش المعدل‪.‬‬ ‫كل المعامالت السابقة تم تسخٌنها الً ‪°85‬م والتبرٌد الفجائً ثم إضافة الفٌتامٌنات الذائبة فً الدهن‬ ‫والذائبة فً الماء وتم تحلٌلها ومقارنتها بلبن االم‪ .‬وكانت النتائج المتحصل علٌها كاآلتً ‪-:‬‬ ‫‪ - 1‬التركٌب الكٌمائً للخلطات المقترحة كانت متقاربة مع لبن االم ماعدا البروتٌن الذي لوحظ انخفاضه فً‬ ‫لبن االم‪.‬‬ ‫‪ - 2‬لوحظ عدم تجبن لبن األم باستخدام المنفحة‪.‬‬ ‫‪ - 3‬كان معدل الهضم بواسطة اإلنزٌمات متقارب جدا فً الخلطات المقترحة ولكنها أعلً قلٌال فً لبن االم‪.‬‬ ‫‪ - 4‬تم عمل اإللكتروفورسٌس مع مقارنته بلبن األم والشرش قبل وبعد التعدٌل لمالحظه غٌاب‬ ‫البٌتاالكتوجلوبٌولٌن وتفرٌد األجزاء المختلفة للبروتٌن للخلطات المجهزة مقارنة بلبن االم‬ ‫ب ‪ -‬تم تقييم ألبان األطفال المقترحة كغذاء مناسب ‪:‬‬ ‫بتغذٌة بعض فئران التجارب علٌها ومقارنتها بالمغذاه علً لبن األم‪ .‬وخلصت النتائج إلً ما ٌلً‪:‬‬ ‫‪ ‬الوزن المتحصل علٌه فً فترة التجربة ( ‪ 7‬أسابٌع) كان أعلً فً حالة التغذٌة علً األلبان المقترحة عن‬ ‫الكنترول‪.‬‬ ‫‪ ‬كمٌة الغذاء المستهلكة بواسطة الفئران فً فترة التجربة كانت غالبا متماثلة‪.‬‬ ‫‪ ‬مدي كفاٌة الغذاء كانت متقاربة بالتغذٌة علً التركٌبات المقترحة أللبان األطفال وكانت أعلً منها فً‬ ‫الكنترول وكانت أفضلها هً ‪F3‬‬

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