Citrate in milk

Feed has relatively little effect on the concentration of most elements in milk because the skeleton acts as a reservoir of minerals. The level of citrate in milk decreases on diets very deficient in roughage and results in the Utrecht phenomenon , i.e. milk of very low heat stability. Relatively small changes in the concentrations of milk salts, especially of Ca, Pj and citrate, can have very significant effects on the processing characteristics of milk and hence these can be altered by the level and type of feed, but definitive studies on this are lacking. 

Effect of temperature on the salt balance of milk studied by capillary ion electrophoresis Calcium, sodium, chloride, phosphate and citrate in milk samples Sample filtered with cut-off at lOK, ultrafiltrate diluted, analyzed... 

It should be possible to determine experimentally the concentrations of anions such as HPO4 and Citrate in milk using ion-exchange resins or by nuclear magnetic resonance spectroscopy, but no such experimental work has been reported and available data are by calculation only. 

The relatively low concentration of citrate in milk (—8 mM) belies the importance of its metabolism in some cheeses made using mesophilic cultures (for reviews, see Cogan, 1985 Cogan and Hill, 1993). Citrate is not metabolized by L. lactis or L. cremoris but is metabolized by L. lactis subsp. 

Approximately 90% of the citrate in milk is soluble and is lost in the whey however, the concentration of citrate in the aqueous phase of cheese is approximately three times that in whey (Fryer et al., 1970), reflecting the concentration of colloidal citrate. Cheddar cheese contains 0.2 to 0.5% (w/ w) citrate which decreases to 0.1% at 6 months (Fryer et al., 1970 Thomas, 1987). Inoculation of cheesemilk with Lb. plantarum accelerated the depletion of citrate pediococci did not appear to utilize citrate (Thomas, 1987). 

Some species of the LAB group such as Leuconostoc mesenteroides subsp. cremoris, Leuconostoc mesenteroides subsp. dextranicum, and Lactococcus lactis subsp. lactis biovar diacetylactis, are known for their capability to produce diacetyl (2,3-butanedione) from citrate, and this metabolism appears especially relevant in the field of dairy products (Figure 13.4). Actually, selected strains belonging to the above species are currently added as starter cultures to those products, e.g., butter, in which diacetyl imparts the distinctive and peculiar aroma. Nevertheless, in particular conditions where there is a pyruvate surplus in the medium (e.g., in the presence of an alternative source of pyruvate than the fermented carbohydrate, such as citrate in milk or in the presence of an alternative electron acceptor available for NAD+ regeneration) (Axelsson, 2(X)9, pp. 1-72), even other LAB such as lactobacilli and pediococci can produce diacetyl by the scanted pyruvate (Figure 13.5). Thus, in addition to butter and dairy products, diacetyl can be present in other fermented foods and feeds, such as wine and ensilage (Jay, 1982). 

Mulzelburg ID (1979) An enzymatic method for the determination of citrate in milk. Austr J Dairy Teehnol 34 82-84... 

The opportunity of use of a ternary complex of ions Eu(III) with oxatetracycline (OxTC) and citrat-ions (Cit) for luminescent detection of OxTC in milk after chromatographic isolation is shown. 

Calcium and magnesium. Some calcium and magnesium in milk exist as complex undissociated ions with citrate, phosphate and bicarboante, e.g. Ca Citr-, CaP04, Ca HCOj. Calculations by Smeets (1955) suggest the following distribution for the various ionic forms in the soluble phase ... 

The murexide method measures Ca2+ only Mg2+, at the concentration in milk, does not affect the indicator appreciably. Calculation of Mg2 + concentration is possible when the total calcium and magnesium (obtained by EDTA titration) is known. This is based on the assumption that the same proportion of each cation is present in the ionic form, which is justifiable since the dissociation constants of their citrate and phosphate salts are virtually identical. 

The so-called Ling oxalate titration indicates that CCP consists of 80% Ca3(P04)2 and 20% CaHP04, with an overall Ca P ratio of 1.4 1 (Pyne, 1962). However, the oxalate titration procedure has been criticized because many of the assumptions made are not reliable. Pyne and McGann (1960) developed a new technique to study the composition of CCP. Milk was acidified to about pH 4.9 at 2°C, followed by exhaustive dialysis of the acidified milk against a large excess of bulk milk this procedure restored the acidified milk to normality in all respects except that CCP was not reformed. Analysis of milk and CCP-free milk (assumed to differ from milk only in respect of CCP) showed that the ratio of Ca P in CCP was 1.7 1. The difference between this value and that obtained by the oxalate titration (i.e. 1.4 1) was attributed to the presence of citrate in the CCP complex, which is not measured by the oxalate method. Pyne and McGann (1960) suggested that CCP has an apatite structure with the formula ... 

Milk contains a range of groups which are effective in buffering over a wide pH range. The principal buffering compounds in milk are its salts (particularly soluble calcium phosphate, citrate and bicarbonate) and acidic and basic amino acid side-chains on proteins (particularly the caseins). The contribution of these components to the buffering of milk was discussed in detail by Singh, McCarthy and Lucey (1997). 

For the purpose of this discussion, milk salts are considered as ionized or ionizable substances of molecular weight 300 or less. Ionizable groups of proteins are not included here, although, of course, they must be taken into account in a complete description of ionic balance and equilibria. Trace elements, some of which are ionized or partially so in milk, are considered in a later section of this chapter. Milk salts include both inorganic and organic substances thus they are not equivalent to either minerals or ash. The principal cations are Na, K, Ca, and Mg, and the anionic constituents are phosphate, citrate, chloride, carbonate, and sulfate. Small amounts of amino cations and organic acid anions are also present. 

Not all of the salt constituents are found in the dissolved state in milk. Calcium, magnesium, phosphate, and citrate are partitioned between the solution phase and the colloidal casein micelles (see Chapter 9 for the composition and structure of these micelles). For analytical purposes, partition of the salt constituents can be achieved by equilibrium dialysis or by pressure ultrafiltration. In the latter technique, pressures must be limited to about 1 atmosphere to avoid the so-called sieving effect (pushing water through the filter faster than the dissolved components (Davies and White 1960). 

Ethanol and a long list of carbonyl compounds and aliphatic acids occur in fresh milk (Table 1.5). Some of them have been detected in only a few of the samples in which they were sought. Techniques for detecting such compounds include derivatization with 2,4-dinitrophe-nylhydrazine and various methods of volatilization, extraction, and chromatography (Harper and Huber 1956 Morr et al. 1957 Harper et al. 1961 Wong and Patton 1962 Scanlan et al. 1968 Marsili et al. 1981). The sum of the concentrations of acids listed in Table 1.5 is only 1-3 mmol/liter, compared to the citrate concentration of 10 mmol/liter. Oxalate has been reported to occur in milk (Zarembski and Hodgkin-son 1962) on the basis of a certain colorimetric reaction, but positive identification has not been made. 

The presence of sugars and salts can also affect the rate of mutarotation. Although the effect is small in dilute solutions, a combination of salts equal to that found in solution in milk nearly doubles the rate of mutarotation (Haase and Nickerson 1966). This catalytic effect is attributed primarily to the citrates and phosphates of milk. The presence of high levels of sucrose, on the other hand, has the opposite ef-... 

In principle, it would be logical to combine plots of the buffer index curves of each of the buffer components of milk and thus obtain a plot which could be compared with that actually found for milk. It is not difficult, of course, to conclude that the principal buffer components are phosphate, citrate, bicarbonate, and proteins, but quantitative assignment of the buffer capacity to these components proves to be rather difficult. This problem arises primarily from the presence of calcium and magnesium in the system. These alkaline earths are present as free ions as soluble, undissociated complexes with phosphates, citrate, and casein and as colloidal phosphates associated with casein. Thus precise definition of the ionic equilibria in milk becomes rather complicated. It is difficult to obtain ratios for the various physical states of some of the components, even in simple systems. Some concentrations must be calculated from the dissociation constants, whose... 

Although citric acid is present in milk in small amounts (0.07-0.4%), it is a required substrate for production of desirable butter-like flavor and aroma compounds in cultured products. Because seasonal variation in the citrate content of milk is sufficient to affect the flavor of cultured products (Mitchell, 1979), milk may need to be supplemented with citrate to produce cultured products with consistent flavor. Citric acid is metabolized by many organisms found in milk, including S. lactis subsp. diacetylactis, Leuconostoc spp., Bacillus subtilis, various lactobacilli, various yeasts, coliforms, and other enteric bacteria. 

Minerals The main mineral constituents in milk are calcium and chlorine, magnesium chloride, phosphate, and citrate. Minerals in milk are mainly present as soluble salts or in colloidal form associated with caseins. Their concentrations may vary enormously. Thus, the minerals present in milk can be classified according to their concentration level as major and minor elements, with small quantitative contributions from trace and ultratrace elements. The total content of minerals in mammalian milks should correspond to the growth requirements of each biological species. Accordingly, the mineral total content in cow milk is four times higher than in human milk. 

Distribution of Cu in milk has been studied by SEC coupled to ICP-AES [12, 14], and ICP-MS [15, 17-19], or even using electrothermal atomization atomic emission spectrometry (ET-AAS) for the detection [20]. Breast milk Cu seems to be distributed all over the biocompounds from high (caseins, immunoglobulins, lactoferrin, and serum albumin) to lower-molecular-weight ligands (lac-toalbumin, peptones, free aminoacids, citrates, etc.). The distribution patterns of Cu have been shown to be very different in mature milk and colostrum [18] (Fig. 17.3). 

Milk products Determination of iodide in whole milk chloride and/or sodium in butter lactate, pyruvate and citrate in cheese... 

Complexones such as EDTA (complexone III) [1-3] and DCTA (complexone IV) [4,5] are suitable eluents, but other complexing agents, such as citrate [3,6] and sulphate [7] are also applied. Barium has been separated from strontium and other metals by cation-exchange chromatography using mixed HCl-organic solvent eluents [8]. Strontium has been enriched and determined in sea water [5] and in milk [2]. 

Metabolic alkalosis may also be generated by the gain of exogenous alkali. This may be seen as a result of bicarbonate administration or from the infusion of organic anions that are metabolized to bicarbonate, such as acetate, lactate, and citrate. The milk-alkali syndrome was historically a common cause of metabolic alkalosis in patients with peptic ulcer disease secondary to the ingestion of large quantities of milk products and antacids. This syndrome has become increasingly uncommon with the advent of alternative effective therapies for dyspeptic syndromes. 

Citric Acid. Thunberg (T21) has summarized the steps which led to the identification of citric acid in urine. Found in lemon juice by Scheele in 1784, citric acid was considered a typical plant acid until its presence was shown in milk in 1888. Classified among the normal metabolites by Thunberg since 1910, this acid was found first in animal urine after administration of citrate, then in normal human urine by Amberg and McClure (A6). Later on, it was found also in normal human plasma by Benni et al. (B9). 

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