protein components of goat’s milk - … milk is a complex biological fluid produced by the mammary...

Protein Components of Goat’s MilkG.F. Greppi et al.

4 Protein Componentsof Goat’s Milk

G.F. GREPPI1,2, P. RONCADA2 AND R. FORTIN1

1Dipartimento di Scienze Cliniche Veterinarie, Facoltà di MedicinaVeterinaria, Università di Milano, Via Celoria 10, 20133 Milan, Italy;2Istituto Sperimentale Italiano ‘Lazzaro Spallanzani’, Località La Quercia,260207 Rivotta D’Adda, Italy

Introduction

Milk is a complex biological fluid produced by the mammary gland which hasthree main functions: (i) nutritional; (ii) immunological; and (iii) physiological.Recently, the physiological role of milk has been emphasized in research, andconsumer demand for high-quality products has increased. Consequently, thetransformation industry has started to market ‘high-quality milk’ with particularnutritional/dietary characteristics. In industrialized countries, milk proteins repre-sent 30% of total dietary protein intake and 75% of Ca intake (Meisel, 1993a,b,1997; Morand-Fehr, 1996; Reinert and Fabre, 1996; Spuergin et al., 1997; Restaniet al., 1999).

Several epidemiological studies have focused on the influence of cow’s milkconsumption on health, especially on cancer development. However, availabledata are contradictory, due to the complex interactions among milk components.Nevertheless, a positive linear relationship between cancer and energy intake orfat-rich milk consumption has been suggested (Attaie and Richter, 2000).

Milk contains lipid components which are well-known anticarcinogenicagents (Parodi, 1997). According to the World Cancer Research Fund, the lipidcomponents of milk and derived products increase the risk of two types of canceronly (kidney and prostate) (WCRF, 1997).

In an early study on the use of goat’s and ass’s milk, high infant mortalityoccurred due to the bad microbiological characteristics of the goat’s milk employed(Michel, 1910). As a consequence, for more than half a century goat’s milk wasnot recommended for human diets. However, since the 1980s several otherstudies and epidemiological research have highlighted the nutritional propertiesand the presence of minor components of pharmacological interest in goat’smilk, thus recommending its usage in human feeding (Domeniconi and Balzola,1980; Birkbeck, 1984; Parry, 1984; Darnton Hill et al., 1987; Awiatlo et al., 1990;Park, 1991; Houdebine, 1995; MacDonald, 1995; Albertini et al., 1999).

© CAB International 2008. Dairy Goats Feeding and Nutrition(eds A. Cannas and G. Pulina) 71

The US Food and Drug Administration has just concluded that food fromcattle, swine and goat clones and their progenies is as safe to eat as food fromanimals bred conventionally (Rudenko and Matheson, 2007). This decision onfood safety will certainly be greatly discussed in the very near future.

Biochemical Characteristics

Milk characteristics can be specifically associated with the environment and spe-cies. For example, there is a positive linear relationship between the logarithmof milk protein quantity and the body weight of the mother, which indicatesthat some mammalian species may produce insufficient milk for the newborn(Fig. 4.1).

Nowadays 4027 species are classified as mammals, but the milk compositionof only 200 species, 60 of these with fewer than ten samples, has been studied(Jenness, 1974, 1980). High intraspecific variations in protein percentage arecommonly observed, probably due to differences in genotype and/or in environ-mental conditions. A high number of scientific studies have demonstrated thatmilk protein content of 200 mammalian species can vary from 1% up to 20%.Milk is a colloidal heterogeneous system where mineral salts, lactose and somewhey proteins dissolved in water represent the continuous phase, while caseinmicelles represent the dispersed phase. Fat is present as tiny globules in this

72 G.F. Greppi et al.

mouse

Body weight

guinea pigmonkey

woman

horse

cow

goat

dog

rabbit

rat

Log

of p

rodu

ced

prot

eins

(g/

day)

Fig. 4.1. Relationship between body weight and produced proteins in mammals(logarithmic scale).

emulsion, the membranes of which are constituted mainly by a phospholipid–protein complex. The biochemical composition and characteristics of milk varyfrom species to species, reflecting the newborn’s nutritional needs (Jenness andSloan, 1970; Jenness, 1979; Davis and Collier, 1983; Targowski, 1983).

For a long time, the use of milk in the diet of adults and, in particular, ofinfants has been studied. In comparison to the milk of other species, goat’s milkhas been considered a good source of proteins and amino acids (Tables 4.1–4.4).

In recent decades, milk has been considered as a message carrier to nursinginfants (Teschemacher et al., 1977; Teschemacher and Brantl, 1994; Teschemacher,1995) and adults as well. In the past, very few studies indicated the presence ofneurotransmitters or hormonal substances in milk (Blanc, 1982). On the otherhand, in the last few years, hundreds of works have shown the presence ofprolactin, melatonin, oxytocin, growth hormone, luteinizing hormone-releasinghormone, thyroid-stimulating hormone, vasoactive intestinal peptide, calcitonin,neurotensin and cholecystochinin in milk, indicating that milk can be consideredan exogenous endocrine system for the newborn (Mackle and Bauman, 1998).

Protein Components of Goat’s Milk 73

Proteins Lipids Lactose

PrimatesHuman 9–15 38–41 70–72Simian 16 40 70

RodentsRat 81 88 38

RuminantsGoat 29–31 35–45 41–44Cow 32–34 37–39 48

CetaceansDolphin 68 330 11Whale 109 423 13

Table 4.1. Milk composition of different species (g/l).

Human milk Cow’s milk Goat’s milk

Water (%) 87.6 87.3 87.5Dry residue (g/l) 11.7 12.5 13.6Dust (g/l) 2.0 8.0 8.9Total proteins (g/l) 10 34 33Carbohydrates (g/l) 70 48 51Total lipids (g/l) 38 37 29Non-protein N (g/l) 3.2 2.5 3.2Ca (mg/l) 33 125 124P (mg/l) 15 96 105

Table 4.2. Composition of human, cow’s and goat’s milk.



Total proteins 9–15 32–34 28–32Caseins 2.0–2.5 26–37 22–28aS1-Casein – 11–15 10aS2-Casein – 3–4 3b-Casein 1.5 9–11 11k-Casein 0.5 2–4 4

Whey proteins 6.3 5.8–6.5 5.5–6.5a-Lactoalbumin 1.9–2.6 0.6–1.5 1.2b-Lactoglobulin – 3–4 3.1Serum albumin 0.4 0.4 0.5Immunoglobulin 1.1 1.0 1.0Lactoferrin 1.7–2 0.1 0.02–0.2Lysozyme 0.04–0.2 – –

Table 4.3. Composition of the protein fraction of human, cow’s and goat’s milk(g/l).


Total proteins (g/l) 9–15 32–34 28–32Essential amino acids(mg/g protein)

Cysteine 20 9 9Phenylalanine 37 52 47Isoleucine 53 64 48Histidine 23 28 26Leucine 104 100 96Lysine 71 83 80Methionine 16 27 25Tyrosine 46 53 38Threonine 44 51 49Tryptophan 17 14 –Valine 51 68 61

Non-essential aminoacids (mg/g protein)

Aspartate 86 79 75Glutamate 190 208 209Alanine 40 35 34Arginine 36 37 29Glycine 22 21 18Proline 95 101 106Serine 61 56 49

Table 4.4. Protein amino acid composition of human, cow’s and goat’s milk.

These hormonal substances should be considered as passengers that use milk toreach their target organ (Choick, 1998).

Knowledge of milk proteins has also increased enormously. For example,the functional meaning of some caseins and whey proteins has been identifiedand evidence for the importance of biopeptides (i.e. amino acid sequences inalimentary proteins) has been found. These peptides are inactive when they arepresent in their native protein, but they can be released in their active form afterproteolytic digestion (i.e. during gastric digestion or simply during any tech-nological treatment). Since the 1980s, many authors have described activebiopeptides derived from milk proteins which, in contrast with endogenousbiopeptides, have many biological functions (Yoshikawa et al., 1988). Sincethen, the nutritional importance of milk, such as in the diet of athletes, hasincreased. Athletes need to consume proteins with high biological value, in orderto: (i) maintain a positive N balance during physical activities; (ii) allow an increaseof muscular mass; and, occasionally, (iii) repair muscular lesions after training(Fig. 4.2). In these cases, since protein needs increase from twofold to threefoldwith respect to normal needs, kidneys may suffer due to high protein intake. Theconsequences from intense physical activity can be: (i) nutritional deficiencies; (ii)formation of free radicals that could also lead to immunological deficiencies; and(iii) a lack of Fe, during aerobic stress, with consequent limited performance. Ironintegration into the diet may have undesirable effects. Therefore, in all of thesecases, utilization of goat’s milk could be helpful (Domeniconi and Balzola, 1980).

Milk has two main protein fractions: caseins, which are predominant, andwhey proteins (Law and Brown, 1994). The qualitative and quantitative contentof each protein fraction depends on many factors: (i) physiological (lactation stage,lactation order); (ii) environmental (climate, hygiene); (iii) genetic (breed, gene-alogy); and (iv) nutritional (Polidori et al., 1991). More precisely, six main pro-teins are present in milk: four caseins (aS1, aS2, b and k) and two whey proteins(a-lactoalbumin and b-lactoglobulin) (Perez and Calvo, 1995). The protein frac-tion of goat’s milk, like that of other domestic ruminants, is constituted mainly bycaseins, which account for 80% of total proteins (Ambrosoli et al., 1988).


CARBOHYDRATES200–260 g/day

PROTEINS40–110 g/day

AMINO ACIDS GLUCOSE Acetyl-CoA

LIPIDS65–95 g/day

FATTYACIDS

TRIGLYCERIDES13 kgEnergy

GLYCOGEN0–0.5 kg

PROTEINS13 kg

Fig. 4.2. Energy production and protein metabolism in the athlete.

It has been recently demonstrated that mRNA levels for the four caseinsproduced in the mammary epithelial cells during lactation are almost equallyabundant (~25% each), while mature protein levels in milk differ greatly(Bevilacqua et al., 2006). In particular, translational efficiency for aS1- andb-casein is about four times higher than that for aS2- and k-casein, thus demon-strating that regulation of casein biosynthesis is also controlled at translationallevel.

The most abundant milk proteins are the three caseins (aS1, aS2 and b),which are sensitive to Ca. Casein biosynthesis is performed at cellular level underthe influence of external agents such as hormones (Figs 4.3 and 4.4). Casein is aglobular protein, present in milk as a colloidal suspension, having the aspect ofmicelles. It is characterized by a hydrophobic core and a charged polar hydro-philic part. The anionic regions of the polar part are responsible for sensitivity toCa2+ and for some physicochemical properties of this protein (Jaubert et al.,1999). Moreover, caseins are relatively hydrophobic, and under ionic condi-tions, such as those present in milk, they tend to associate tightly into theirtypical colloidal form (micelles). Micelles are constituted by a protein fractionand by a mineral component (Ca and phosphate). Caseins are a group of spe-cific phosphoproteins in milk which precipitate: (i) when milk is acidified to a pH


Progesterone

Glucocorticoids ProlactinInsulin

P

P

P(CHO)n

Ca2+

GolgiEndoplasmic reticulum

mRNADNA

Fig. 4.3. Casein biosynthesis. Following external signalling (i.e. hormones) andsignal transduction inside the nucleus, casein genes are transcribed into mRNAmolecules, which undergo post-transcriptional processing once inside the cytosol.Mature mRNA molecules are translated into single casein molecules which aretransferred inside the endoplasmic reticulum for post-translational modification.Casein micelle formation begins in the cis-Golgi with condensation of caseinmolecules; addition of Ca2+, possibly in the secretory vesicle, leads to maturationof the casein micelles into particles sufficiently dense to be seen in the electronmicroscope.

of 4.6–4.7 at 20°C; (ii) by enzymatic treatment (chymosin); or (iii) by ultracentri-fugation. Caseins are rich in proline, glutamate and glutamine, and poor inglycine and aspartate. A high amount of Ca and phosphate is required to makecasein soluble in milk in a stable form, so that it becomes a nutritional source ofamino acids.

A very important milk feature, especially in goat’s milk, is related to its con-tent of complex oligosaccharides, either in free form or conjugated to proteins. Infact, beneficial anti-inflammatory effects of such oligosaccharides, especially oninflammatory bowel disease, have been demonstrated (Daddaoua et al., 2006).

Milk Protein Polymorphisms

Aschaffenburg and Drewry (1955) reported, for the first time, the presence oftwo variants of b-lactoglobulin in cow’s milk. Since then, many other polymor-phisms have been identified both in caseins and in whey proteins. In the pastdecade, molecular and biochemical characterization of protein polymorphismshas been performed and applied to the genetic improvement of animals and theimprovement of milk characteristics, as described in detail in the reviews ofNg-Kwai-Hang (1997, 1998). Many protein variants were correlated with differ-ences in casein or whey protein composition as well as in milk physicochemicalproperties related to caseification (Martin et al., 2002).

In the last 20 years, electrophoretic methods have been widely used to iden-tify genetic variants of proteins (Russo et al., 1981, 1986; Martin, 1993; Martinet al., 1999). The molecular origin of milk protein polymorphisms is the substitution


Exon

Mature protein

Stop

Intron 18size 81–8500 bp

Fig. 4.4. Molecular aspects of biosynthesis.

or deletion of amino acids in the protein chain (Table 4.5). Alleles associatedwith reduced protein synthesis have been reported by many authors (see thereview of Valentine, 1998, for molecular aspects). These modifications result inchanges of milk physicochemical properties such as charge, superficial hydro-phobicity and molecular size and shape (Russo et al., 1981, 1986; Addeo et al.,1987, 1989; Grosclaude et al., 1987; Dall’Olio et al., 1988, 1989; Brignon et al.,1989; Di Gregorio et al., 1989). Such modifications have been identified throughthe use of electrophoretic and chromatographic techniques (Addeo et al., 1988).Each protein fraction shows different genetic variants (Mercier and Vilotte,1993). These differences in the primary structure of a protein can deeply modifyits molecular characteristics. The ratios between various protein fractions canvary and are regulated by structural genes (Parma et al., 1999a,b; Rando et al.,2000). Since each one of the four types of casein is polymorphic, differentgenetic variants can be identified. Polymorphisms can be attributed to two mainfactors: (i) genetic variants; and (ii) post-translational modifications, due to a dif-ferent localization of phosphorylation or glycosylation sites in the amino acidicsequence of the primary structure (Bouniol et al., 1993, 1994; Ramunno et al.,1994; Klose, 1999). Intrinsic electrophoretic heterogeneity of casein fractions isdue to the presence of incompletely phosphorylated or glycosylated sites in theseproteins (Bevilacqua et al., 2002; Martin, 1993; Ramunno et al., 2004).

The aS1- and aS2-caseins differ in their content of phosphoric groups, whilek-casein differs in its content of glucidic and phosphoric groups (Boulanger et al.,1984; Leroux et al., 1992, 2003; Iametti et al., 1996). b-Casein heterogeneityderives from incomplete saturation of its phosphorylation sites (Chianese et al.,1993).

The most studied goat’s milk polymorphism has been regarding aS1-casein(Bouniol et al., 1993; Remeuf, 1993; Jansa-Pérez et al., 1994; Remeuf et al.,


Protein Variant Amino acid position

aS1-Casein 14–26 53 59 192A MissesB Ala Gln GluC GlyD ThrE Lys Gly

aS2-Casein 33–47 50–58 130A Glu Ala ThrBC Gly Thr IleD Misses

Ala, alanine; Gln, glutamine; Glu, glutamic acid; Gly, glycine; Ile, isoleucine; Lys, lysine; Thr,threonine.

Table 4.5. Milk protein genetic variants.

1995; Feligini et al., 1995, 1996, 1998; Chianese et al., 1996; Pena et al., 1998;Martin et al., 1999; Iametti et al., 2000). The gene encoding this protein is amajor-effect gene, due to the presence of alleles which are responsible for highdifferences in the amount of such protein in milk (Parson and Heflick, 1997). Atchromosomal level, seven alleles are associated with the codification of threedifferent quantities of aS1-casein in milk: (i) aS1-Cn A, aS1-Cn B and aS1-Cn C,so-called ‘strong alleles’, are associated with a production of 3.6 g/l per allele;(ii) aS1-Cn E, so-called ‘medium allele’, is associated with a production of 1.6 g/lper allele; (iii) aS1-Cn D and aS1-Cn F, so-called ‘weak alleles’, are associated witha production of 0.6 g/l per allele; and (iv) aS1-Cn 0, which seems to be a ‘nullallele’, whose homozygous form does not synthesize any kind of casein (Brignonet al., 1990; Pierre et al., 1998a,b). Therefore, milk produced by homozygousindividuals AA, EE, FF and 00 would have 7.2, 3.2, 1.2 and 0 g aS1-casein/l,respectively (Tables 4.6 and 4.7). Some examples of the main electrophoretictechniques used in the analysis of the protein profile of goat’s milk are reportedin Figs 4.6–4.9 (Jaubert and Martin, 1992; Cattaneo et al., 1996; Recio et al.,1997; Roncada et al., 1997, 2002a,b, 2003a,b,c, 2004; Murakami et al., 1998;Bini and Roncada, 1999; Roncada and Greppi, 1999).


aS2-Casein

aS1-Casein

b-Casein

k-Casein

Fig. 4.5. SDS-PAGE of milk caseins. Milk proteins can be separated andidentified by molecular mass (molecular weight; in kDa). One method of separatingproteins is by polyacrylamide gel electrophoresis. This method essentially separatesthe proteins by molecular mass, with the largest proteins migrating more slowly inthe gel and remaining nearer the top and the smaller proteins migrating more rapidlytowards the bottom of the gel. The relative size of the caseins is ~25–35 kDa.


Casein

Genotype aS1 aS2 b k Other

Medium 28.35a 0.62a 53.74b 15.76 1.62a

Weak 5.20b 21.16b 49.01a 19.15 3.92b

Null 1.61c 23.98b 52.87b 17.07 4.46b

a,b,cValues in a column with different superscript letters were significantly different (P ≤ 0.05).

Table 4.6. Individual casein content (%) per genotype.

Whey protein

Genotype b-Lg a-La SA Lf Other

Medium 43.8 21.8 9.0 2.6 22.8Weak 45.8 23.3 7.7 1.5 21.7Null 50.0 23.1 7.0 1.5 18.4

b-Lg, b-lactoglobulin; a-La, a-lactoalbumin; SA, serum albumin; Lf, lactoferrin.

Table 4.7. Major whey protein content (%) per genotype.

Fig. 4.6. Isoelectric focusing analysis of casein samples of goat’s milk (pH 7–4from top to bottom) (in the low part, aS1 variants).

Compared with the other genotypes, milk with strong genotype at theaS1-casein locus is well suited for dairy transformation due to the following prop-erties: (i) higher Ca content, therefore higher propensity for enzymatic coagula-tion; (ii) lower casein micelles diameter, therefore higher coagula consistency;(iii) lower activation period of coagulation process, i.e. the time between curdleaddition and coagula hardening; and (iv) faster formation of coagula.

Even if quantitative differences between genetic variants of single proteins arealmost always small (Di Luccia et al., 1990; Grousclaude et al., 1994; Noè, 1995;Lamberet et al., 1996; Enne et al., 1997; Bramanti et al., 2003; Noè and Greppi,2003), they might directly or indirectly influence milk technological and nutritionalproperties (Ambrosoli et al., 1988; Addeo et al., 1989; Heil and Dumond, 1993;Remeuf, 1993; Pirisi et al., 1994; Vassal et al., 1994; Delacroix-Buchet et al.,1996; Buchin et al., 1998; Trujillo et al., 1998; Clark and Sherbon, 2000). Themain studies on goat casein polymorphisms are reported in Table 4.8.

b-Casein is the most abundant of the caseins on SDS-PAGE (Fig. 4.5), giv-ing rise to two bands, b1 and b2. These two isoforms (b1 and b2) differ from eachother due to the presence of six and five phosphoric groups, respectively(Galliano et al., 2004).

Some recent studies have demonstrated that the milk of Italian goat breedshas a b-null allele (b-Cn 0) associated with the aS1-Cn A allele (Ramunno et al.,


Fig. 4.7. Electrophoretogram of goat’s milk in two-dimensional electrophoresis(pH 4–8 from left to right, molecular weight 220 kDa to 10 kDa from top to bottom).Protein spots have been identified by matrix-assisted laser desorption/ionization–time-of-flight mass spectrometry.

1996; Cosenza et al., 2003). This b-null allele reduces the content of b-casein,but it is also associated with an increase of aS1- and total casein. Moreover, com-pared with heterozygous individuals, milk produced by homozygous b-Cn 0/0individuals has: (i) coagulation time three times higher; (ii) coagula with reducedconsistence; and (iii) lower yield of caciotta cheese (Chianese et al., 1993; Penaet al., 1998).


4020

a

Abs

orba

nce

at 2

80 n

m (

AU

)

b

d

c

Fig. 4.8. Reversed-phase high-performance liquid chromatograms of differentvariants of aS1-casein (A, B and F); a = k-casein, b = aS2-casein, c = b-casein.

Studies on aS2-casein are not as numerous as those on aS1-casein. aS2-Casein has three known genetic variants, A, B and C (Bouniol et al., 1994),which have only recently been identified through electrophoretic techniquessuch as isoelectric focusing and SDS-PAGE. Alleles differ from each other only ina single amino acid substitution which has ties with a phosphorylation site. Thelink between aS1-Cn and k-Cn does not seem to be as strong as that betweenaS1-Cn and b-Cn (Chang et al., 1993).

A genetic polymorphism at the k-Cn locus was also found in k-casein (DiLuccia et al., 1990; Angiolillo et al., 2002; Yahyaoui et al., 2003; Prinzenberget al., 2005). This casein has two known variants, A and B, which probably differ


aS2

A21

4A

214

aS1

g1

g2 g3

S S S

b1

b2

k

b2 b1

k

aS1

0.02

0.06

0.04

0.02

0

0.04

0.06

0.08

0.10

0

Fig. 4.9 Capillary electrophoresis of goat’s milk (upper) and plasmin-treated goat’smilk (lower); S indicates casein peptides.


Breed Country Obs. Year A B C D E F 0

Alpinea F 213 1994 14 5 1 34 41 5Alpineb I 38 1994 3 26 1 5 42 10 8Alpinec I 40 1993 – – – – – – 23Alpined I 80 1991 – – – – 35 59 6Alpinee I 26 1995 13 15 – – 9 40 21Alpinef I 113 1995 11 29 – – 16 14 30Alpineg I 88 2005 43 20 0.6 0.6 11 23 1Blonde Ae I 85 1996 1 3 – – 5 33 58Canarianh S 74 1991 28 32 – – 20 – 20Corsei F 106 1989 6 13 – – 14 59 8Frisiane I 36 1995 – 3 – – 5 37 55Frisianc I 70 2006 13 0.7 – – 20 56 11Garganicb I 48 1994 39 41 5 4 – – –Garganicd I 54 1991 61 – 37 – – 2 –Garganich S 77 1991 8 25 – – 62 5 –Malaguenianh S 56 1991 – 25 – – 70 5 –×Maltesiane I 177 1995 21 38 – – 4 11 27Maltesiand I 81 1991 33 – 28 – – 11 27Maltesianb I 372 1994 46 16 1 – 5 14 3×Sardiniane I 181 1995 20 53 – – 5 14 8Ionicb I 77 1994 39 27 8 – 5 1 4Marroche M 82 1995 21 52 4 – 9 7 6Orobicf I 70 1995 4 – – – 2 47 45Orobicc I 66 2006 – 0.8 – – 0.8 96 2Payoyah S 39 1991 4 14 – – 82 – –Poiteviniani F 209 1989 5 35 – – 45 14 –Poitevinianj F 302 1995 4 36 – – – 43 9Rovei F 147 1989 12 5 0 0 62 10 11Saanena F 159 1994 7 6 0.3 – 49 46 –Saanend I 70 1991 5 35 – – 45 14 –Saanene I 50 1995 10 8 – – 18 10 54Sardiniane I 115 1995 11 59 – – – 7 14Sirianb I 241 1994 50 17 – – – 6 16Tunisiane T 86 1995 31 41 1 – 5 5 17Verzaschesee I 57 1995 4 34 – – – 7 48Verzaschesek I 54 1999 3 30 – 2 5 49 11Verzaschesec I 67 2006 – 4 – – 20 75 0.7

F, France; I, Italy; M, Morocco; S, Spain; T, Tunisia.Data source: aGrosclaude et al. (1994); bChianese et al. (1994, 1996); cCaroli et al. (1993, 2006);dRamunno et al. (1991); eFeligini et al. (1998); fMeggiolaro et al. (1995); gBudelli et al. (2005);hJordana et al. (1991); iMahé and Grosclaude (1989); jRicordeau et al. (1995); kIametti et al. (1999).

Table 4.8. Allelic frequencies at aS1-Cn in different goat breeds.

in just a single amino acid residue. The physicochemical properties of k-caseinare characterized by solubility in the presence of Ca2+ and sensitivity tochymosin. Its content in milk influences coagulation, since this phase starts onlywhen all k-casein has been hydrolysed. Even if a strong association betweenaS1- and b-casein has not yet been demonstrated, it has been hypothesized thatmany correlations exist between all casein loci for goats, as already found for cat-tle (Parma et al., 1999a,b, 2003a,b).

Allergic Reaction and Milk Intolerance

Alimentary intolerance is an abnormal reaction by the organism that occurs afterfood ingestion, regardless of the pathogenetic mechanism involved. Allergy isdefined as the sum of clinical symptoms (e.g. rhinitis, asthma and eczema) whichappear after contact between an ‘antigen’ (e.g. dust mites, pollen, cow’s milk,pig’s meat, barley) and a specific antibody of the ‘epsilon’ (IgE) class ofimmunoglobulins. The IgEs are commonly attached to the surface of differentcell types such as mastocytes and polymorphonucleate basophiles. Contactbetween an antigen and its specific IgE leads to the production of mediators,such as histamine, which cause an excessive immune response, from whichderives the term hypersensitivity. Free histamine is present in considerable quan-tities in tuna, mackerel, other dark meat fish and in seasoned cheese. Otherfoods, such as strawberries, tomato and albumen, cause the release of histaminethrough particular mechanisms which are also valid for lecithins contained infruit, cereals and many vegetables (Carbonaro et al., 1996; Bellioni-Busincoet al., 1999; Cartoni et al., 1999).

If the above-mentioned and many other biologically active agents (e.g. anti-oxidants, antimicrobials, toxins, microorganisms) are present in great quantities,they can potentially induce a great variety of symptoms, whose manifestationdepends on the sensitivity of each individual. For example, subjects with hyper-active gastrointestinal and respiratory mucus membranes or with irritable skinhave a low threshold and thus also react in the presence of low amounts of anti-gen (biologically active agent in food), showing symptomatology typical of ali-mentary allergy.

Cow’s milk contains at least 20 different proteins, but only five are relatedto allergy: (i) serum albumin, g-globulin and a-lactoalbumin, which are thermo-labile and thus easily deactivated; and (ii) b-lactoglobulin and casein, which arethermostable and resistant to the proteolytic action of the digestive apparatus.In the case of infants, the structure of any milk protein ingested would remainunaltered, i.e. amino acids are not removed from its peptide chain, due to insuf-ficient enzymatic activity of proteases or lack of a specific enzyme in the intestine.Therefore, milk protein would be recognized as a foreign body (antigen). Thequantity of absorbed antigenic material depends on anatomic barriers (mucus,epithelium) and/or its removal by immunological barriers, as a consequence ofthe combination of antigens with secretory immunoglobulins (IgA) on the surfaceof the epithelium. This is an important protective action which reduces the aller-genic penetration, since antigens (milk proteins) do not have access to IgE


and IgG. If these barriers are weakened, by inflammatory processes or by lack ofIgA, the penetration of high quantities of antigens can cause sensitivity andallergy in atopic individuals. In general, the newborn is not intolerant to maternalmilk, excluding those affected by serious metabolic diseases (phenylketonuria,galactosaemia) or by diseases that may require interruption of milking (icterus, diar-rhoea, constipation). However, in some cases cow’s milk is not tolerated bybreast-fed children as some of its thermostable protein components(b-lactoglobulin and casein) are absorbed almost intact and recognized as ‘for-eign’ proteins by hypersensitive subjects. Cow’s milk intolerance is convention-ally defined as cow’s milk protein allergy (CMPA) or cow’s milk proteinintolerance, and can affect 2–4% of the population. In paediatrics, the estimatedfrequency of cow’s milk intolerance ranges from 1 to 8%, while that of atopicpredisposition in the entire population varies between 15 and 20%. Cow’s milkallergy is the first cause of digestive allergies in industrialized countries.

The CMPA can be of immunologic or non-immunologic origin. Amongnon-immunologic intolerances lactose intolerance is the best known, being causedby a lack of intestinal lactase. However, not all problems and symptomatologiesfound in sensitive individuals following ingestion of certain foods are caused byalimentary allergy (i.e. proved or likely to be of immunologic aetiology). In fact,in many cases such disturbances are related to alimentary intolerance, pseudo-allergy or idiosyncrasy.

CMPA encompasses different types of allergies: (i) type I, with immediatehypersensitivity reaction which can lead to anaphylactic shock, nettle rash,asthma crisis and serious digestive problems; (ii) type III, with complex immuno-logic reactions characterized by enteropathies and internal haemorrhages; and(iii) type IV, with a retarded hypersensitivity which causes chronic diarrhoea.Clinical symptoms of CMPA are very different. They can be acute or delayed,and digestive or extradigestive. Even if the risk of anaphylactic shock is rare, it canaffect 10% of CMPA individuals. Digestive problems can be acute or chronic,while described extradigestive symptoms are mainly related to the skin (dermati-tis, nettle rash) or the respiratory tract (asthma). Nevertheless, renal problems,blood alterations, and central nervous system or neurological alterations mayalso occur, at a lower frequency.

The fact that the clinical symptomatology of CMPA is protein-related hasresulted in a shortage of specific diagnostic tools. For example, total and specificIgE is a diagnostic tool only for type-I CMPA, while intestinal biopsy and perme-ability test are difficult to perform and are suitable only for enteropathic forms.There are predisposing factors that can favour the appearance of CMPA symp-toms, which are correlated with: (i) some degree of hereditability, i.e. allergic rel-atives; (ii) early feeding with cow’s milk (fresh or powdered), during the very firstdays of a newborn’s life; and (iii) development of gastroenteritis within the first3 months of life, after which individuals are considered to be at high risk. Asalready noted, an inflammatory intestinal disease increases epithelial permeabil-ity and, as a consequence, the risk of reaction due to intolerance or allergy. Morethan a quarter of newborns who suffered gastroenteritis in their first months oflife showed CMPA symptomatology afterwards. In addition, a relevant percent-age of children who suffered from gastroenteritis within the first year and up to


3 years of life also show CMPA symptoms later on. However, whereas milk isessential in the first few months of life, it can be replaced by other food productsas children get older. The few studies conducted on the use of goat’s milk inhuman feeding are encouraging (Dore et al., 1993). Moreover, goat’s milk hasbeen reported to be one of the best alternatives for the production of hypo-allergenic formulas for infants in case of cow’s milk allergy (D’Auria et al., 2005).Very recent studies in this field have demonstrated that goat’s and sheep’s milkallergy differs from cow’s milk allergy because it affects older children andappears later. Cow’s milk products do not cause any clinical manifestation inpatients allergic to goat’s and sheep’s milk, whereas goat’s and sheep’s milkcause cross-reaction in patients allergic to cow’s milk (Ah-Leung et al., 2006).

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protein components of goat’s milk - … milk is a complex biological fluid produced by the mammary...

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