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Abstract: Background: Human milk not only contains nutrients and antibodies, but also can be used as an indicator for levels of organic pollutants in human bodies. We developed a method for determining persistent organic pollutants (POPs) in the colostrum of women in preterm labor, the POPs of 36 colostrum samples have been examined. Methods: Thirty-six samples of colostrum from preterm women were extracted by acetone–acetonitrile, enriched and purified by solid-phase Florisil columns. The purified POPs were further separated by the capillary columns, and detected by the gas chromatography–electron capture detection (GC–ECD). Results: The average recovery rates of 6 types of organochlorine-based pesticides were 80.2%–112.1%, which represent the first 3 categories of the 12 species of POPs. The precise quantities detected were 3.85%–9.32% (the limits of detection were 0.03 μg/l to 0.08 μg/l), and the linear correlation coefficients were ≥0.9969. Of the 36 women tested, 10 (27.8%) were found to have colostrums containing traces of dichloro-diphenyl-trichloroethane (DDT), and 2 (5.56%) were tested positive for dieldrin. Conclusions: The combination of using GC–ECD proved to be both accurate and reliable, and this process proved to be both simple and time-effective. This method is applicable for determining the levels of POPs in organisms. [Copyright &y& Elsevier]

Gonadotropin-releasing hormone(GnRH)-associated peptide (GAP) is a 56-amino acid peptide found on the C-terminal of the GnRH (also called luteinizing hormone-releasing hormone) precursor and is assumed to be co-produced with GnRH. The purpose of this report is to demonstrate the presence of GAP immunoreactivity in bovine colostrum. Radioimmunoassay of acidified methanolic extracts demonstrated a concentration of GAP immunoreactivity of approximately 1.5 +/- 0.1 pmol/g dry skim bovine colostrum. Gel filtration (Sephadex G-10) and high-performance liquid chromatography of extracts containing GAP immunoreactivity showed it to be of low molecular weight and a high hydrophobic character. The presence of GAP immunoreactivity in bovine colostrum suggests that the GnRH precursor is synthesized and processed in mammary tissue itself.

The ribonucleosides adenosine, cytidine, guanosine, inosine and uridine as well as the modified components N1-methyladenosine and N6-carbamoylthreonyladenosine were characterized and determined quantitatively as minor constituents in raw bovine milk by use of an automated high performance liquid chromatography system. The studies have shown that except for the colostral phase the ribonucleoside levels are constant throughout the whole lactation period. That means, there is a typical ribonucleoside pattern which is assumed to be species-specific.

Human milk proteins have both nutritional and physiological roles for the breast-fed infant. While the biochemistry and developmental patterns for many whey proteins are well known, our knowledge of human casein and its subunits is still limited. We have recently developed a method to isolate casein from whey proteins in human milk and to separate the casein subunits by fast protein liquid chromatography. In this study we have applied this methodology to study the casein subunit pattern in preterm milk, colostrum, and mature milk. Casein concentration increased with lactation time, largely due to an increase in glycosylated forms of casein (kappa-caseins). Thus, the relative proportion of beta-casein to kappa-casein decreased during the lactation period. The patterns of phosphorylated and glycosylated casein subunits were found to vary during lactation, showing that both synthesis and posttranslational modification of beta- and kappa-casein are regulated by different mechanisms.

Vitamin B-6 status, assessed by plasma pyridoxal phosphate (PLP) concentrations, and vitamin B-6 concentrations in breast milk were examined in 47 lactating mothers supplemented with different amounts of pyridoxine.HCl (PN.HCl) during pregnancy and the first 6 mo of lactation. PLP concentrations in cord blood and maternal plasma at 2 d postpartum and vitamin B-6 concentration in colostrum were positively correlated with the amount of PN.HCl supplementation prenatally (r = 0.71, p less than 0.001; r = 0.74, p less than 0.001; and r = 0.78, p less than 0.001, respectively). Correlations between the amounts of PN.HCl supplementation postnatally and plasma PLP concentrations increased with the length of supplementation. Plasma PLP concentrations were also correlated with vitamin B-6 concentrations of milk samples, which were obtained on the same day as plasma. PN.HCl supplements between 2.5 and 4.0 mg/d (2.1-3.4 mg PN equivalents) ensured vitamin B-6 adequacy of the mother and maintained relatively saturated concentrations of vitamin B-6 in breast milk.

Mammary secretions obtained from four groups of sows at parturition and on days 7, 14 and 21 of lactation were defatted and assayed for total protein and insulin-like growth factor-I (IGF-I). Sows (n = 57) represented two breeds (Landrace and Duroc) and two genetic lines (selected for differences in sow productivity index, SPI) within each breed. Colostrum of Duroc sows was 4-6 fold and 30-60 fold greater in protein (P less than .001) and IGF-I (P less than .001) concentrations, respectively, than the corresponding day 7 milk from these sows. In contrast, the colostrum of Landrace sows was 2-3 fold and 30-50 fold greater in protein (P less than .001) and IGF-I (P less than .001) concentrations, respectively, than the corresponding day 7 milk. The IGF-I content in milk from Duroc sows did not differ among days 7, 14 and 21 of lactation, whereas the IGF-I content of day 7 milk from Landrace sows exceeded those for the corresponding 14 day and 21 day secretion (P less than .05). IGF-I concentration in days 14 and 21 milk was higher in Duroc (P less than .001 respectively) than Landrace sows. No significant differences in total protein or IGF-I content of mammary secretions were observed between the selected and control lines within each breed.(ABSTRACT TRUNCATED AT 250 WORDS)

Myoinositol concentration was studied in serum of 65 neonates and their mothers at the time of birth, in samples of various types of feedings for infants, and in serial serum samples of 15 premature infants receiving human milk, formulas for infants, or parenteral nutrition over a 3-wk period. At birth the serum concentration of myoinositol was greater in neonates than in their mothers (108 +/- 10 vs 52 +/- 6 mumol/L, respectively, means +/- SEM, p less than 0.01). In feedings for infants, the concentrations of myoinositol were significantly greater in human milk than in formulas or parenteral nutrition solutions (1840 +/- 451 vs 420 +/- 110 vs 100 +/- 8 mumol/L, respectively, p less than 0.001). Over a 3-wk period the serum concentration of myoinositol increased in infants receiving human milk but not in those receiving formulas or parenteral nutrition. Serum concentrations of myoinositol in neonates are greater than in adults and are directly influenced by myoinositol intake.

Colostrum, the initial postpartum secretion of the breast, ordinarily has a distinct yellow color due to carotenoids of its fat globules. This pigmentation progressively diminishes as milk production increases during the first week of lactation. Identity of these carotenoids was investigated by means of thin-layer chromatography, high performance liquid chromatography and spectral analysis. Alpha- and beta-carotene, lycopene and beta-cryptoxanthin were revealed as major chromogens. A component corresponding to lutein and/or zeaxanthin was also detected by both chromatographic techniques. Extracts of 23 saponified colostrum samples from 10 donors revealed considerable variation in total carotenoid concentration (0.34-7.57 micrograms/ml of colostrum). Multiparous mothers had greater mean colostrum carotenoid concentrations than did the primiparae, 2.18 +/- 1.94 vs 1.14 +/- 1.32 micrograms/ml, respectively. Seven of the eight primiparous donors' samples had little or no yellow color. These findings imply a difference in carotenoid transport by breasts that have lactated as compared to those that have not. The interrelation of carotenoids, lactation and breast cancer is discussed.

The occurrence and properties of PTH-related peptide (PTH-RP) in milk was investigated. PTH-RP was purified to homogeneity from human and bovine milk using heat and acid to precipitate milk proteins followed by ion exchange chromatography and reverse-phase HPLC. The peak of PTH-RP from HPLC was detected using a sensitive bone cell bioassay. A single band of peptide was detected on silver-stained polyacrylamide gels, which migrated as a 20-21-kDa macromolecule. PTH-RP isolated from either human or bovine milk had similar electrophoretic mobilities on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The partially purified bovine PTH-RP stimulated cAMP production in UMR106-01 and OK cell lines and elicited a concentration-dependent inhibition of sodium-dependent phosphate transport in OK cells. Incubation of milk extracts with an anti-PTH antiserum did not affect their bioactivity, whereas an antihuman PTH-RP 1-34 antiserum markedly reduced the cAMP response of UMR106-01 cells to the immunoabsorbed milk extracts. A PTH antagonist, norleu PTH 3-34, blocked the stimulation of cAMP production in UMR106-01 cells treated with milk extracts. PTH-RP immunoreactivity and bioactivity occurred in milk extracts of diverse animals from both eutherian and metatherian (marsupial) species. Porcine colostrum also had immunoreactive PTH-RP, although the levels were lower than the immunoreactive PTH-RP concentrations observed in milk samples collected at 7 and 14 days of lactation. Thus, a 20-21-KDa PTH-RP is secreted into milk where it could play a role in the development of suckling, newborn animals.

Antimicrobial proteins in maternal and cord sera and sequential samples of human milk were studied in a group of 60 Chinese women to determine the degrees of passive immunity transferred from women of different nutritional status to their infants. Maternal malnutrition was characterized by low values for wt/ht2 and serum total protein and albumin. Maternal immunoglobulin (IgG) concentrations were not significantly different between well- and malnourished groups prepartum but were significantly different postpartum. Mean concentrations of cord IgG and lysozyme from well- and malnourished groups were not statistically different. During the first 7 d of lactation and most stages thereafter, mean concentrations of IgA; complements C3 and C4, and lysozyme in milk from the malnourished group were only half of those of the well-nourished group. Antimicrobial proteins transferred via milk to newborns may be influenced by the mother's nutritional status.

The composition of colostrum and milk and of sows fed either 3.4 kg or 1.0 kg daily of a conventional sow diet during the last 14 days of gestation was investigated. The fat content of colostrum of the very restrictedly fed sows was higher compared with the standard fed sows. There was a significantly decrease in the crude protein and immunoglobulin contents of colostrum during the first 4 h post farrowing. The milk composition 14 days after farrowing was not affected by the late pregnancy feeding level. The composition of milk was significantly correlated with the 3 week litter weight. The colostrum IgG content was not correlated to the mortality of piglets during the first week post favouring. The content of crude protein and immunoglobulins in the milk was not correlated with the frequency of post weaning diarrhoea.

A colostrum sample was collected within 24 h after foaling from 27 mares and from 10 other mares a milk sample was collected several weeks post partum. Immunoglobulin G concentrations were determined quantitatively by radial immunodiffusion and semi-quantitatively using a commercial latex agglutination test. Total protein, specific gravity, viscosity and refractive index were determined and their relationships to the immunoglobulin G concentration analysed. All parameters correlated with the immunoglobulin G concentration. The latex agglutination test divided the colostrum samples into three groups with different means for immunoglobulin G and total protein concentrations. Specific gravity and the latex agglutination test were found to be the methods best suited for on-farm evaluation of colostrum quality.

Presence of specific secretory immunoglobulin A (sIgA) against pathogenic factors of Escherichia coli related with diarrheal disease was studied in colostrum and breast-milk samples obtained longitudinally from a cohort of rural Mexican women. Levels of sIgA against heat-labile enterotoxin, Shiga-like toxin, colonization factors antigens I, II and E8775 and adherence to HEp-2 cells were detected in samples obtained from 54 rural women during the first year of lactation. Although production of specific sIgA against these pathogenic factors was almost universal it was not constant, even in the same woman. The results reflect a definite mother-infant relationship during this period. The data support the thesis of using breast-milk as a vaccination vehicle against diarrhea associated with specific organisms during the first year of life of infants born in developing areas of the world.

Synthesis of beta-lactoglobulin and alpha-lactalbumin by explants of ovine mammary gland and evolution of concentration of these proteins in cow and sheep colostrum and milk throughout early lactation have been studied. The evolution of both proteins was similar in cow and sheep species. The highest concentration was found in the first milking (19 and 2 mg/ml for beta-lactoglobulin and alpha-lactalbumin, respectively). Then, levels of beta-lactoglobulin decreased sharply and those of alpha-lactalbumin slowly during the first days of lactation, reaching stable values during the second week postpartum (4 and 1.5 mg/ml). The concentration ratio beta-lactoglobulin/alpha-lactalbumin was four times greater in colostrum than in mature milk. On the other hand, synthesis of these proteins represented about 25-30% of the synthesized total soluble proteins. The synthesis ratio beta-lactoglobulin/alpha-lactalbumin in explants obtained at 12 and 30 hours postpartum was found to be 3.5 and 1.7. These results indicate that the synthesis and secretion of beta-lactoglobulin are comparatively greater than those of alpha-lactalbumin during colostral period, suggesting that beta-lactoglobulin could have some specific function during this period.

Lactoferrin content of colostrum obtained from cows within 24 h after parturition was measured using a single radial immunodiffusion test and was compared among cows of two dairy breeds (Holstein-Friesian, Jersey) and two beef breeds (Japanese Black and Japanese Brown). Average lactoferrin content in colostrum of dairy breeds was 2 mg/ml and in colostrum of beef breeds was .5 mg/ml. Lactoferrin content of colostrum due to lactation number was also different among breeds. In dairy breeds, multiparous cows had lactoferrin content two to three times higher than that of primiparous cows; beef breeds showed no obvious differences between lactation years. Lactoferrin content also varied considerably within breed. In beef breeds, half the cows had values of nearly zero. Transferrin content in colostrum was fairly constant (.9 mg/ml) and was not as variable among and within breeds. There was no correlation between lactoferrin and transferrin contents in colostrum. Examination of cows lacking lactoferrin suggested that transferrin plays an important role as an iron carrier from a cow to her newborn calf.

Immunoaffinity columns made with monoclonal antibodies to either human or bovine lactoferrins were prepared to isolate human lactoferrin or bovine lactoferrin from milks by a single chromatographic step. Recoveries of human lactoferrin and bovine lactoferrin were 98 and 97%, respectively. The human lactoferrin recovered from defatted human colostrum was 98% pure with 93% iron-binding capacity. Amount of recovered bovine lactoferrin, as well as purity and iron-binding capacity, varied widely depending on the source of bovine milks and pretreatments (particularly pasteurization temperature). The best source to isolate bovine lactoferrin was raw skim milk yielding a protein 97% pure and with a 99% iron-binding capacity. Thus, immunoaffinity chromatography provides an effective one-pass isolation of highly pure human or bovine lactoferrin with reasonable recovery and iron-binding capacity.

Growth-promoting activity in bovine colostrum has been detected as the capacity to stimulate protein synthesis in L6 myoblasts. By using this assay as a measure of bioactivity, a growth factor has been purified to near homogeneity from centrifuged colostrum by a series of steps including acid extraction, chromatography on sulphopropyl-Sephadex, followed by adsorption to, and elution from, C18 columns using acetonitrile and propan-1-ol gradients. The purified growth factor has a low solubility at neutral and alkaline pH and has an Mr of 7800 by gel-permeation chromatography. Sequence analysis of the first 30 amino acids from the N-terminus indicated complete identity in this region with human insulin-like growth factor-1. Accordingly we conclude that the purified growth factor is bovine insulin-like growth factor-1.

Serum myoinositol (henceforth called inositol) concentration was measured in 12 preterm infants (birth weight 800-1,700 g, gestational age 27-32 weeks) from birth to 10 postnatal weeks. The diet was analyzed for inositol concentration and the daily intake was correlated with serum inositol. There were striking differences in the inositol concentration of infant feedings: preterm colostrum 4.22 +/- 0.51 mM; term colostrum 2.91 +/- 0.21 mM; mature milk 1.81 +/- 0.20 mM; infant formulas 0.09-0.39 mM; parenteral nutrition 0.15 +/- 0.13 mM. The high fetal serum inositol often fell during the first 2 weeks. After 2 weeks, serum inositol correlated significantly with inositol intake (R = 0.601, p less than 0.004). On breast milk serum inositol concentrations (0.56 +/- 0.07 mM) were higher than on formula feedings (0.36 +/- 0.03 mM). Since according to animal studies inositol is important during prenatal growth and differentiation, the present data justify further study on importance of dietary inositol in preterm infants.

40% of the primary structure of the cow colostrum proteinase inhibitor (CTI) is homologous with the structure of the trypsin kallikrein inhibitor (TKI) from bovine organs; the positions of the reactive lysine residues are also the same in both inhibitors. Both CTI and TKI were modified by carbamoylation and the fully labeled derivatives were isolated by ion-exchange chromatography. The effect of the modification on the antitryptic and antichymotryptic activity of both inhibitors was investigated. The antichymotryptic activity of both inhibitors is not decreased after the modification. The antitryptic activity of modified TKI is retained, yet the dissociation constant of the complex of the modified inhibitor with trypsin is considerably increased; nevertheless, modified TKI is a good trypsin inhibitor. The antitryptic activity of modified CTI is hardly detectable. We explain this difference in the behaviour of both inhibitors by a replacement of basic residues Arg-17 and Arg-39 in TKI by neutral amino acids Ala-20 and Gln-42 in CTI.

Colostral and milk samples were collected from 6 primiparous sows on postpartum days 0, 9, 18, and 27. Insulin content was measured and epidermal growth factor (EGF) concentration was determined by use of radioimmunoassay and radioreceptor assay, respectively. Total milk protein concentration was measured colorimetrically. Concentrations of insulin, EGF, and protein in the colostral samples were significantly (P less than 0.01) higher than those found in the milk. Milk samples obtained from postpartum days 9, 18, and 27 did not differ significantly in insulin, EGF, or total protein content.

Colostrum removed from mammary glands of parturient cows before letdown contained no greater concentration of immunoglobulins than colostrum collected after letdown and complete milking. Greater absorption of colostral immunoglobulins in calves suckling their dams initially postpartum over those hand fed by buckets or bottles is not from higher concentration of immunoglobulins in the gland and teat cistern. Colostral concentrations of three immunoglobulin classes (G, M, and A) obtained from one partial and six consecutive complete milkings at 12-h intervals postpartum decreased at different rates over time or number of milkings. Potential prophylactic value is discussed of continued lacteal secretion of immunoglobulins A and M available to the calf after intestinal closure to systemic absorption of colostral immunoglobulins.

Reported contents of fluoride (F) in human milk vary considerably. The aim of this study was to determine the F content in human milk under different levels of F intake using a siliconfacilitated microdiffusion technique, which had a good accuracy and precision. The mean F concentration of colostrum from mothers in a 1.0 ppm and a 0.2 ppm F area was 0.36 +/- 0.02 mumol/l (+/- SEM) and 0.28 +/- 0.02 mumol/l, respectively. The mean F concentration of mature milk from a 1.0 ppm F area was 0.37 +/- 0.04 mumol/l. Within the 1.0 ppm F area, the intra- and interindividual differences in F concentration were very small. No statistically significant difference in milk F concentration between the two areas was found. Consequently, breastfed infants living in a 1 ppm or a 0.2 ppm F area will have an approximately equal F intake of 5-10 micrograms per day, in spite of great differences in F intake among the nursing mothers.

Sephacryl S-300 Superfine is a new gel filtration medium that consists of allyldextran crosslinked with N,N'-methylenebisacrylamide. It was shown to have a separation range for proteins that extends to 1--2 x 10(6) dalton. The gel filtration properties were characterized by fractionation of human serum and of bovine colostrum. Thyroglobulin from pig could be isolated in one step in a very pure form. The flow properties were characterized by the relationship between flow rate and pressure drop. It was shown that Sephacryl has very good flow properties in comparison with Sephadex G-200 and that it can be used in large-scale columns.

As revealed by agarose gel electrophoresis and immunoassay, followed by staining with Sudan black, lipoproteins occur in the alpha-, pre-beta- and beta-regions of the serum of fetal, newborn, neonatal, juvenile and adult pigs. These lipoproteins undergo characteristic quantitative changes during development. After the onset of colostrum ingestion, a lipid-stainable zone appeared in the gamma-region of piglet's serum and sow colostrum/milk. The levels of total serum lipids, cholesterol, high and low density lipoproteins were low at birth in comparison to those of the adult pig, but increased to higher levels after the onset of colostrum and milk ingestion. After 3-8 weeks, the total lipids, cholesterol and low density lipoprotein values had decreased to adult level but the high density lipoprotein values remained high in comparison to the adult pig. It is discussed that the changes found in lipoprotein and lipid patterns in serum of the developing pig reflect changes in cellular metabolism and nutritional needs of the organism. It is suggested that the perinatal pig might be a useful model for studying the development of atherogenic diseases in man.

Samples of colostrum were taken from 19 subjects before and after infant feeding, 3 and 5 days post partum. Milk samples were analyzed by argentation and gas chromatography for 30 fatty acids contained in the lipid fraction. Although small differences were observed between day 3 and day 5 samples, the fatty acid composition of the milk did not vary on either of the days between samples taken before and after nursing. It is concluded that a single sample taken any time during the feeding is representative of the entire nursing.

Trypsin inhibitor from cow colostrum has been purified by affinity chromatography of colostral proteins on insolubilized trypsin. The method described compares favourably, in both simplicity and yield, with previous methods developed for the isolation of this inhibitor. Gel electrophoresis followed by characterization of antitrypsin activity allows the demonstration of four molecular forms of bovine colostral trypsin inhibitor in both crude colostral whey and purified preparations of the inhibitor. Immunoelectrophoresis of each of these materials with antisera specific for this inhibitor reveals a single precipitation line of broad anodic mobility. By immunodiffusion tests, the precipitation lines in preparations of purified inhibitor and colostral whey appear immunologically identical. In contrast, absence of crossed reactivity was observed between bovine colostral trypsin inhibitor and trypsin inhibitors of bovine serum. This strongly suggests the high specificity of this inhibitor as a colostral and milk constituent.

Protease inhibitors (inhibiting trypsin, chymotrypsin, and elastase) were demonstrated in human milk from birth to 4 months after delivery. No pepsin inhibitor was found. The protease inhibitors were localized in the alpha 1-region--inhibiting trypsin, chymotrypsin, and elastase--and in a more cathodal region--inhibiting chymotrypsin--in agarose gel electrophoresis of human milk. alpha 1-antitrypsin and antichymotrypsin were demonstrated by crossed immunoelectrophoresis. Electroimmunoassay showed the concentration of alpha 1-antitrypsin in 1st day milk to be 10.9% and the concentration of antichymotrypsin was 116% of that of adult serum. The concentrations decreased during the 1st wk; from 1 wk to 4 months they were 1.6% for alpha 1-antitrypsin and 3.8% for antichymotrypsin of those of adult serum. One ml of human milk from the first 3 days inhibits, by enzymatic methods, 0-150 microgram (mean value, 70 microgram) trypsin. Milk from 1 week after delivery and later inhibits 0-65 microgram (mean value, 38 microgram) trypsin per ml.

Postpartum, presuckle, colostrum samples were collected from 100 mares. Colostral specific gravities significantly correlated (r = 0.9) with colostral immunoglobulin (Ig)G concentrations. Foal serum IgG concentrations highly correlated (r = 0.82) with specific gravities of the colostrum each foal ingested. Eight of 48 foals (17%) had serum IgG concentrations less than 400 mg/dl. The dams of these 8 foals had colostral sp gr less than 1.06 and colostral IgG concentrations less than 3,000 mg/dl. Foals had serum IgG concentrations greater than 520 mg/dl 24 hours after parturition, when the colostral specific gravity of the dam was greater than or equal to 1.06. Effects of breed on colostral specific gravity, colostral IgG concentrations, foal serum IgG concentrations, and mare serum IgG concentrations were not significant.

A new one-step procedure for isolating lactoferrin from human whey using Cibacron Blue affinity chromatography is described. Characteristics of the isolated lactoferrin include pI 5.5-6.1 and differential migration on SDS-polyacrylamide gel electrophoresis of apo and iron saturated lactoferrin. The concentration of lactoferrin in breast milk varies with time after parturition but the iron saturation remains relatively constant with time.

Trypsin inhibitor from sow colostrum was isolated by ion exchange chromatography on DEAE-Sephadex A-50 followed by gel filtration chromatography on Sephadex G-100 and affinity chromatography. Antiserum against sow colostrum trypsin inhibitor was produced by immunization with the purified inhibitor, and made specific by absorption with normal porcine serum. The specific antiserum was used for immunoquantitation by single radial immunodiffusion (SRI). In sow colostrum whey, good agreement was found between the results obtained by SRI and the total trypsin-inhibiting activity as determined by radial diffusion in a casein-containing agarose gel (r = 0.97, n = 10). In sow's milk there was only a very low inhibiting activity, and no colostral inhibitor was demonstrable by SRI. Also in baby-pig urine agreement was found between the two methods (r = 0.97, n = 14). In baby-pig serum such an agreement was not seen, undoubtedly becuase of the presence of genuine serum trypsin inhibitors. By the SRI technique it is possible specifically to determine the colostral inhibitor even in the presence of other trypsin inhibitors.

Colostrum was inoculated with Streptococcus lactis or yogurt culture or preserved with .1% (vol/vol) formalin in two separate experiments. All preparations then were stored at ambient temperature for 24 days. With increasing storage time, a larger proportion of the total colostrum nitrogen was not precipitated in 10% (wt/vol) trichloroacetic acid. By day 24, this represented 30 to 35% for the fermented samples and 10 to 15% for the formalin preserved samples. The majority of this nonprecipitable nitrogen was amino acids and small peptides. Most of the nitrogen in colostrum even after 24 days of storage should be nutritionally useful to the calf because even the nonprecipitable portion is amino acids and peptides.

Bovine beta 2-microglobulin (beta 2-m), the light chain of the histocompatibility antigen, was isolated in crystalline form from colostrum. Previous studies from this laboratory on the solution properties of this protein suggest the existence of a time-dependent multiple aggregation phenomenon. To clarify the molecular states of beta 2-m, its solution properties were studied by ultracentrifugation and spectropolarimetry. Sedimentation equilibrium experiments at pH 5.0 (0.08 M NaCl, 0.02 M sodium phosphate) at concentrations less than 0.3 mg/ml give Mr = 11,800. From sedimentation velocity results, we conclude that bovine beta 2-m is a much more symmetrical and compact molecule than either guinea pig or human beta 2-m. At concentrations above 0.4 mg/ml under the same conditions, sedimentation equilibrium experiments show that a monomer to tetramer reversible self-association occurs. Also, the tetramerization increases with decreasing temperature. beta 2-Microglobulin undergoes an irreversible temperature-dependent association to a much larger aggregate over a period of 7 days, as evidenced by sedimentation equilibrium and velocity results. The rate of this aggregation decreases as the pH approaches the isoelectric point (pH 7) from either side. Furthermore, circular dichroism measured at pH 5.0 under time-dependent aggregating conditions showed a marked increase in the percentage of disordered structure, leading to the conclusion that this effect is a denaturation phenomenon.

The trypsin-inhibitor capacity of bovine milk was seen to increase in mastitis. This capacity showed good correlation with the California Mastitis Test score, somatic cell count (Counter method) and bovine serum albumin (BSA) content (radial immunodiffusion). The elevated antitrypsin level proved to be a safer indicator of mastitis than BSA alone. Using gel-filtration chromatography, the major trypsin-inhibitor activity of mastitic milk and plasma was eluted with BSA, indicating a similar mol. wt (70 000). This fraction showed electrophoretic polymorphism. A large molecular weight fraction cross-reacted immunologically with human alpha 2-macroglobulin. The antitrypsin activity in mastitic milk was clearly different from the principal activity in colostrum, which had a mol. wt of about 13 000.

Milk pyroglobulin was capable of passing through the "Visking" 8/32 cellulose membrane, when dialysed against deionized water but not against saline. Gel filtration analysis of the protein suggested its dissociation into subunit molecules in deionized water. Amino acid composition revealed high contents of proline, glutamic acid, valine and leucine. The protein possessed a biological activity to increase the permeability of skin vessels when injected intradermally. The colostral whey which was experimentally freed from heat-coagulable substances contained as major components proteins which are antigenically related to the pyroglobulin.

Use of a microwave oven to thaw frozen colostrum was evaluated. Colostrum was collected from nine cows, four of which were immunized to produce specific colostral antibodies. Colostrum from each cow was frozen, subsequently thawed, and pooled. One-liter aliquots of the pooled colostrum were frozen and assigned randomly to three thawing treatments. Colostrum was thawed using one of three regimens: 10 min in a microwave oven at full power (650 W), 17 min in a microwave oven at half power (325 W), and 25 min in 45 degrees C water. Colostrum thawed in the microwave oven was slightly coagulated and had lower volume and total protein content than colostrum thawed in water. Casein and pH were not different among treatments. Both concentration and total content of immunoglobulin A were higher in the control than in microwave treatments. Neither amount nor concentration of immunoglobulin G and immunoglobulin M were different among treatments. Immunological activity, measured by a hemolytic test, was lower for microwave treatments than the control but did not differ between microwave treatments. Frozen colostrum thawed in a microwave oven should provide a reasonable source of colostrum when fresh high quality colostrum is not available.