R.D. McCarthy

Mammalian metabolism of straight-chain saturated hydrocarbons.

Abstract The metabolism of straight-chain saturated hydrocarbons has been demonstrated in the goat, rat and chicken. Studies with [1-14C]octadecane and [1-14C]hexadecane shwoed that the hydrocarbons were converted directly to fatty acids of the same chain length. Since both the initial substrates were symmetrical molecules with a methyl group on either end, the reaction must be omega oxidation. The studies have suggested a widespread occurrence of the oxidation in various tissues, and among widely divergent animal types. Although, only tracer quantities of hydrocarbons were administered, many intriguing questions for further study have been indicated.

Biosynthesis of secretory lipids from [2-14C]acetate by bovine mammary cells in vitro.

Bovine mammary cells were cultured in vitro and their capacity to synthesize lipids peculiar to the bovine mammary gland was studied using [14C]cetate as substrate. Primary cultures of bovine cells avidly used exogenous acetate (70%) and rapidly incorporated it into fatty acids. The composition and specific activities of the fatty acids produced were similar to those normally synthesized by the intact bovine mammary gland. These de novo synthesized fatty acids were rapidly incorporated into all the lipid classes particularly into the triglycerides. The specific activity-time curves suggested that the triglycerides were produced by the acylation of 1,2-diglycerides with endogenously synthesized fatty acids. The cultured cells actively secreted labeled triglycerides which were compositionally similar to those isolated from bovine milk. In vitro cultures of bovine cells showed an attenuation in lipogenesis from acetate when maintained in culture for periods beyond 72 h. Whereas primary cell cultures incorporated newly synthesized fatty acids into triglycerides for secretion, older subcultures of mammary cells fabricated phospholipids apparently to meet the structural demands of cell proliferation.

Lipid composition and secretory activity of bovine mammary cells in vitro

Abstract The lipid content and composition of collagenase dispersed mammary cells was determined Individual cells contained approx. 300 pg lipid per cell which decreased as the cells aged. The neutral lipid fraction contained triglycerides (56%), sterols (22%), and free fatty acids (13%) as major components. Phosphatidyl choline was the major polar lipid fraction (52%). A trace quantity of phosphatidic acid was tentatively detected in the cellular lipids. Active secretion of fat droplets was observed by phase microscopy. The cells secreted from 0.6 to 7.5 pg lipid per cell per h during the initial 3 days in culture after which secretion diminished. The secreted lipid was composed predominantly of triglyceride whose fatty acid composition was similar to that of bovine milk fat.

Origins of the cholesterol in milk.

Studies were conducted to investigate the origin of milk cholesterol in the ruminant. In the first experiment, [1-14C] sodium acetate was infused into one side of the udder of a lactating goat via the teat canal whereas in the second, [1,2-3H] cholesterol was injected intravenously and concurrently with a [14C] acetate intrammamary infusion. In both experiments, blood and milk samples were collected at intervals for 6 days postinjection. Maximum unesterified cholesterol specific activity (sp act) in whole milk appeared at 78 hr after intravenous injections of3H cholesterol and within 3–7 hr after infusion of [14C]acetate. Virtually all the tritium in milk was associated with unesterified cholesterol. The sp act of14C-labeled cholesterol was only 20% of gland-synthesized decanoic acid. Decanoic acid is known to be completely synthesized in the mammary gland, and, like cholesterol, acetate is its precursor. The results indicate that, although some milk cholesterol is synthesized in the mammary gland, it is derived principally from serum cholesterol. The data show also that serum cholesterol equilibrates with membrane cholesterol of the lactating cell prior to its secretion in milk.

Metabolism of phospholipid in mammary gland: I. The supply of phospholipid for milk synthesis in the rat and goat

Data presented in this study demonstrate that under normal physiological conditions milk phospholipids in the rat and the goat originate predominantly, if not totally, by de novo synthesis within the mammary gland. Evidence to support this has been obtained for the goat by measurement of P32-phosphate incorporation into milk phospholipids, and in the rat by measurement of P32-phosphate incorporation and by feeding radioactive phospholipid to measure the incorporation of serum phospholipids into milk. The latter experiment showed that the fatty acid portion of the dietary phospholipid can readily be utilized by the mammary gland for triglyceride synthesis, but that the contribution of the serum phospholipid “backbone” to milk phospholipid is minimal.

Effect of milk constituents on hepatic cholesterolgenesis

Two preparations active in reducing hepatic cholesterol biosynthesis were isolated from bovine skim milk. One of the inhibitors was in the dialysate and was identified as orotic acid (OA). The other inhibitor, present in the retentate, was not identified. Orotic acid appears to act by inhibiting cholesterol biosynthesis before the formation of mevalonate, whereas the retentate inhibitor exerts its effect beyond the formation of mevalonate in the biosynthetic pathway. Human milk also inhibited the incorporation of both labeled acetate and mevalonate into cholesterol by rat liver. Orotic acid was not detectable in human milk samples employed in this study. Administration of [6-14C]orotate to rats revealed its conversion to uracil in the liver. Subsequent work demonstrated that uracil had inhibitory activity on hepatic cholesterol biosynthesis similar to that of orotate when incubated with rat liver slices.

The serum lipoproteins as a source of milk cholesterol

Amsterdam ods employed in [l] : VLDL, very low density lipoproteins, and chylomicrons p<l.O06; LDL, low density lipoproteins, 1.006(0< 1.063; HDL, high density lipoproteins, l.O63<p< 1.210. Blood serum (5.65 ml) was diluted to 8 ml M NaCl containing 0.1 mg ml for of the ed to the of Folch et al. up 1 v/v) and portions of by thin on plates of silica G (Supelco, Bellefonte, Pennsylvania, U.S.A.). The solvent system was petroleum ether, ethyl ether and acetic acid 180:2: (v/v/v). The developed plates were placed briefly in an iodine to facilitate by scraping in scintillation by the of Searcy of the a mock serum solution (0.195 NaCl) for 24 h at to remove

Multi-origins of milk serum albumin in the lactating goat

L-[U-14C]Leucine was infused into the right-hand mammary glands of lactating goats. Milk from both glands of the animals was sampled at intervals for 36 h. After 3 h the specific activity of milk serum albumin from the infused glands was more than six times that from the non-infused glands. The specific activity of milk serum albumin was considerably lower than that of alpha-lactalbumin or beta-lactoglobulin which are exclusively synthesized by mammary secretory cells. Following the intravenous injection of 125I-labeled serum albumin, maximum specific activity of this protein appeared in milk in 12 h. The specific activity of serum albumin in milk attained no more than 45% of the specific activity of the serum albumin in blood. It is concluded that milk serum albumin has multiple origins and that a portion of it, at least (10-20%), is made in the mammary gland.

The Modulating Effect of an Inhibitor of Cholesterolgenesis Present in Bovine Milk Upon the Synthesis of Cholesterol, Dolichol and Ubiquinone *

Bovine milk contains two inhibitors of hepatic cholesterol genesis. One of these, identified as orotic acid, influences the early segment of the cholesterol biosynthetic pathway and suppresses the conversion of acetate to mevalonate. In this study the other inhibitor was shown to curtail the formation of compounds past farnesyl pyrophosphate on the squalene-cholesterol branch of the pathway. Thus cholesterol synthesis may be suppressed while the production of two other products of the branched pathway, dolichol and ubiquinone, is allowed to continue. The possible role of these ingested regulators in the metabolism of the young until they achieve sufficient development is discussed.