The effects of ionizing radiation on the alkaline phosphatase activity of milk have been studied under different environmental conditions. Twenty-five megarads were required to inactivate the enzyme completely. A combination of heat treatment and irradiation given either before or after heating showed that inactivation of the enzyme was greater when heating occurred after irradiation, this effect of the different forms of energy possibly indicating different mechanisms of heat inactivation. Varying the dose rate over the range 12–250 krad/min did not affect the amount of enzyme inactivated. After irradiation, activity declined during storage at 30°C but not at 5°C. Irradiation did not change the value of Km yet the Vmax fell, indicating that there are fewer active sites present after irradiation but that affinity of the remaining sites for the substrate remains unchanged.

The conductivity of raw and homogenized milks was measured using apparatus capable of high precision. The effect of fat content was examined and found to conform to the established behaviour for a polydisperse suspension of nonconductors in a homogeneous conducting medium. Homogenization of the milk over a range of pressures was found to have no measurable effect on the conductivity.

A routine method giving accurate results was required for the determination of the glyceride content of eluate fractions obtained in the silicic acid chromatography of lipid mixtures. The technique of determining glyceride glycerol by saponification, oxidation of the liberated glycerol to formaldehyde, reaction of the formaldehyde with chromotropic acid and measurement of the resulting chromogen at 570 mμ has been investigated. Procedures based on this principle have appeared in the literature but none of these has been found to be entirely satisfactory. Accordingly, a modification of the method has been developed that is capable of giving results of a high degree of accuracy.

Of several strains of lactic acid bacteria which produced nisinase, Streptococcus thermophilus, strain TJ, was the most active. The optimum pH for extraction of the nisinase from an acetone preparation of bacterial cells and for incubation of the solution containing nisin and nisinase was in the neighbourhood of 7·0. The proportions of nisin destroyed by the nisinase under various conditions were determined. Also, it has been shown that nisinase has no action on a number of antibiotics, including subtilin.

The effect of preservative tablets added to milk on its composition and density is discussed. A comparison between the S.N.F. content of 10-day composite samples of milk from individual herds after making due allowance for added preservative, and the weighted mean S.N.F. calculated on a daily basis, showed that the 10-day composite sample could be used successfully for estimating S.N.F. hydrometrically. A comparison of the weights of S.N.F. supplied when estimated by gravimetric and hydrometric methods was made for ten herds supplying a commercial plant over a period of 260 days. The mean difference was –0·14% of the gravimetrically estimated weight, S.D. 0·4, and the range was ±0·6%. The result is discussed and is considered of sufficient accuracy to justify, on a technical basis, using the hydrometric method of estimating S.N.F. for determining and, if necessary, paying for the S.N.F. supplied.

A simple system was developed, consisting of a solution of β-glycerophosphate, β-lactoglobulin and magnesium ions, in which alkaline phosphatase isolated from milk became strongly reactivated following heat treatment for 45 sec in boiling water. From 10 to 30% of the original enzyme activity reappeared after incubation at 37°C. Variations in the components of the system, and factors affecting it, were studied. Salts of Hg, Zn and Cd inhibited reactivation at low concentrations.

Milk which, after the same heat treatment, became reactivated to a much smaller extent (about 1%), was found to contain a dialysable, heat labile inhibitor whose presence is thought to be largely responsible for the low level of reactivation, though other factors, e.g. the suboptimal concentration of phosphate esters must be considered as contributory causes.

Milk contains also a non-dialysable, heat stable activator of the reactivation process, capable of replacing β-lactoglobulin in the simple system and active at very low concentration, e.g. 0·2% (v/v).

In the years 1959, 1960 and 1961 sections of the College herd of pedigree Guernsey cows were fed indoors on cut grass. The grass, cut twice each day, was fed in excess of appetite.

The weight of grass dry matter consumed varied significantly in each year with the percentage of dry matter in the grass. In one year only did the dry-matter consumption vary significantly with the percentage of crude fibre in the grass.

The weight of drinking water consumed varied significantly with the percentage of dry matter in the grass. The total water intake, i.e. drinking water plus uncombined water in the grass, remained almost constant at approximately 14 gal per cow per day.

The use of a concentrate supplement in 1960 and 1961 produced small increases in yield which were not statistically significant. If, however, the differences recorded were real, then they would have been economic in one year and not in the other.

The intake of starch equivalent, calculated from the chemical analyses of the grass, could not be reconciled with the performance of the cows.

Details are given of a histochemical technique suitable for the detection of alkaline phosphatase in cheese.

Lactoperoxidase activity has been measured in whey form fresh milk and from milk cultured with Str. cremoris or Str. lactis. It was found that with benzidine as hydrogen donor, the optimum H2O2 concentration in whey from fresh milk was 10–4M, whereas in whey from cultured milk it was greater than 10–2M. The lactic acid streptococci appeared to modify lactoperoxidase. The modified form of the enzyme resembled lactoperoxidase in its absorption spectrum, its inhibition by fluoride and cyanide, its chromatographic behaviour on IRC50 resin and its activation energy of heat denaturation, but differed in its heat and pH stability, and its optimum hydrogen peroxide concentration for reaction with benzidine or o-dianisidine.

The viscosity and flow properties of fresh and age-thickened separated sweetened condensed milk have been investigated using a rotating cylinder viscometer. The rheological properties, and the changes therein brought about by stirring, were studied by means of flow curves obtained by plotting stress against the corresponding rate of shear. Condensed milk during age-thickening is shown to develop thixotropic properties, with a high initial viscosity or ‘gel strength’ which, however, is easily reduced by stirring. On prolonged stirring the viscosity reaches a minimum (the ‘permanent’ viscosity), which becomes progressively higher as the age-thickening process continues. On storage subsequent to stirring there is only an insignificant recovery of the lost viscosity at room temperature, but a substantial recovery at higher temperature.

Sweetened condensed whey shows no age-thickening, and the casein-free supernatant liquids obtained by centrifuging dilutions of both fresh and age-thickened condensed milk have the same nitrogen contents and the same viscosities. This is interpreted to mean that the whey proteins play no significant part, directly or in association with casein, in the age-thickening process. Also the sedimentation rate of the casein during centrifuging is greater with age-thickened than with fresh condensed milk, which indicates an increase in casein particle size during agethickening.

The following working hypothesis is suggested to account for this behaviour. During age-thickening a slow irreversible change occurs in the size or shape of the casein micelles, probably by aggregation, which produces the rise in ‘permanent’ viscosity. These micelles orient themselves in such a way as to form a loose network enclosing some of the dispersion medium, thereby producing the large increase in the initial viscosity. On stirring, this network is broken down, and although it can reform in time at high temperatures, such recovery is greatly retarded at room temperature by the high viscosity of the aqueous phase containing the soluble constituents of the condensed milk.

The considerable falls in viscosity of sodium caseinate solutions under the action of rennet are shown to be due to changes in volume and shape (or either alone) of loosely coiled long chain molecules, resulting from the discharge of ionization (‘Third-type electro-viscous effects’). This reduction may be as well effected by small amounts of NaCl or by the separated decomposition products of the rennin-caseinate reaction. Even at the highest practicable caseinate concentrations (9–10% by weight) the solutions are truly fluid over the range of shear-rate of the Ostwald viscometers used

The initial rate of fall of viscosity is found to be proportional to the maximum (initial) specific viscosity, leading to an equation Xk1 = 1/τ where X the proportional viscosity loss, varies with the source of caseinate but is constant for all normal rennets; k1, the first-order reaction constant, varies with the rennet but is independent of the source of casein over a wide range; and τ, which has the dimensions of time, varies with both caseinate and rennet.

The parameter k1 may be found either from the exponential viscosity fall or calculated from the asymptotic initial slope and has been shown to serve as a useful index of rennet activity. This method of rennet testing does not depend on the keeping qualities of rennins, rennets, milks or caseinates. Details of the preparation of caseinates and of the methods for practical testing of commercial rennets are given.

A rapid spectroturbidimetric method for the determination of the fat content of homogenized ice-cream mixes using a simple filter absorptiometer has been devised. Turbidities are measured at two different wavelengths to allow for changes in the degree of homogenization. The standard deviation of the differences from the values obtained by the Werner-Schmid method was found to be 2·5% of the mean fat content value. The method also enables the mean fat globule diameter to be determined.

Details are given of the method used in this laboratory for the preparation of k-casein. The recovery is about 25% and the material has a purity, estimated from electrophoretic patterns, of about 90% with β-casein as the main contaminant. Higher temperatures and lower ion concentration caused precipitation of k-casein in the presence of calcium ions, 0·1m-acetate buffer at pH 6·5 being sufficient to stabilize a 0·5% solution in the presence of 0·01–0·2m-CaCl2 at 20 and 30°C but not at all calcium concentrations at 40°C. It was also found that solutions of para-k-casein did not aggregate in the presence of concentrations of electrolytes above about 0·25m.

The rate of release of non-protein nitrogen and decline in viscosity during the enzymic stage of gel formation in solutions of k-casein and rennin had similar apparent first order constants (0·087 ± 0·03 min—1 and 0·086 ± 0·021 min—1, respectively, at 25°C). A gel could be formed by rennin action in a solution containing as little as 6·25mg of the protein per litre. In the non-enzymic stage of gelling of k-casein solutions the calculated activation energy over the range of 20–40°C was much lower than that obtained from the non-enzymic stage of milk coagulation.

The rate of age-thickening of separated sweetened condensed milk increased with decreasing condensing temperature, over the range 55–22°C. Preheating the milk for 10 min at 85°C reduced the destabilizing effect of low condensing temperatures.

Changes take place after condensing which have the effect of decreasing the rate of age-thickening during subsequent high-temperature storage. These changes are complete in about 2 weeks at room temperature but take longer at lower temperature.

The hypothesis is put forward that the stabilizing effects of a moderately high condensing temperature, and of cool storage after condensing, are connected with the precipitation of insoluble salts, especially calcium phosphate. The effect of preheating is ascribed to the formation of nuclei of precipitated calcium phosphates which facilitates further precipitation during and after condensing.

Experiments on age-thickening at a series of different temperatures with four batches of commercial condensed milk gave mean values for the temperature coefficients of increase in ‘initial’ and ‘permanent’ viscosity of 4·9 and 7·5 per 10 degC, respectively. The difference between these values is thought to support our previous conclusion that two different processes are involved.

Age-thickening of sweetened condensed separated milk can be greatly reduced by removing part of the diffusible salts by dialysis of the milk before processing. This treatment is still effective after high-temperature preheating, which indicates that denaturation of the whey proteins does not influence age-thickening independently of the milk salts.

When added at the level of 20 m-equiv./kg of condensed milk to the milk before condensing, those polyvalent anions (phosphate, oxalate, citrate, ethylenediaminetetra- acetate) which have the property of forming undissociated or insoluble calcium salts greatly increase the rate of age-thickening. Fluoride, which forms an insoluble calcium salt but is monovalent, has the opposite effect.

It is suggested that both natural and added polyvalent anions destabilize the condensed milk by forming linkages between the calcium atoms of the calcium caseinate micelles, producing the larger, more asymmetric units which were postulated in Part I to account for the viscosity changes during age-thickening. Fluoride would have the opposite effect by combining with the calcium, and so preventing such linkage formation by the polyvalent anions.

When added to the milk at the same level as the anions, divalent cations, especially calcium and zinc, reduce the rate of age-thickening. It is suggested that this is due to removal of milk-serum anions, particularly phosphate and citrate, as insoluble or undissociated salts.

The effect of both anions and cations added after condensing is almost the opposite of that produced by the same ion before condensing, with the exception that phosphate increases age-thickening in both cases. There is as yet no clear explanation for this observation.

The ‘metallic compound’ previously reported as an important cause of off-flavour in oxidized dairy products has been identified as oct-l-en-3-one. The compound was found to have a flavour threshold value of one part in 109 in butterfat and one part in 1010 in water. This compound was also isolated from oxidized safflower oil.

Spray-dried whole milk powders have been prepared containing 0·01% of four flavones, propyl and dodecyl gallates, butylhydroxyanisole (BHA) and nordihydroguaiaretic acid (NDGA). The powders were stored at 20 and 37°C and examined at intervals of 25 days for taste of the reconstituted milk, peroxide value of the fat and oxygen absorption of the powder. Three of the four flavones, 7,8-dimethoxy-3,2',5'-trihydroxyflavone, 6-ethyl-3,7,2',5'-tetrahydroxyflavone and 6-dodecyl-3,7,2',5'-tetrahydroxyflavone improved the keeping quality of the powder, the last-named compound being the most effective, but 3,7,8,2'5'-pentahydroxyflavone was without antioxidative properties in this system. These results are briefly discussed with respect to substitution within the flavone molecule.

Dodecyl gallate was a very effective antioxidant for spray-dried whole milk powder and propyl gallate and NDGA, whilst affording good protection, were less so; BHA did little to improve keeping quality.

The best antioxidant of the flavones investigated, 6-dodecyl-3,7,2',5'-tetrahydroxyflavone, appeared to be almost as effective as propyl gallate and NDGA in these milk powders.

The freezing point and pH of milk ultrafiltrate were different from the values obtained for the original milk. It was estimated that from 3·2 to 4·6% of the soluble material in milk was retained by the ultrafiltration membrane. Comparison of the composition of ultrafiltrate and rennet whey showed that the soluble material retained consisted mainly of calcium and citric acid. A similar effect was observed with dialysis, although the quantity of soluble material retained was much less. Rennet whey was considered more representative of milk serum than either ultrafiltrate or diffusate.

A study was made of the effect of various factors which may influence the calcium content of milk. The basic data consisted of the calcium content of 20840 samples of individual milks, taken once a month, during the course of 2300 lactations of Montbeliarde cows. For each lactation, a weighted average figure for the calcium concentration was calculated, taking into account the milk quantities recorded on each sampling day. The calcium content has been expressed in g/kg of milk.

The stage of lactation caused significant variations in calcium content mainly during the first month and the last three months. Expressed as percentages of the arithmetic mean for the whole lactation period, these variations were always within the limits —4 to +8% (Fig. 1).

The season of calving did not greatly affect the weighted lactational average content of calcium but caused some important changes in the average value with advancing lactation (Fig. 2). The minimum values were always observed in July, whatever the month of calving (Fig. 3).

The age of the cow, as well as the age at first calving (Table 1), had very small effects on the calcium content.