Liquid human milk fortifier significantly improves docosahexaenoic and arachidonic acid status in preterm infants.

We report the fatty acid composition of mother׳s own human milk from one of the largest US cohorts of lactating mothers of preterm infants. Milk fatty acid data were used as a proxy for intake at enrollment in infants (n=150) who received human milk with a powder human milk fortifier (HMF; Control) or liquid HMF [LHMF; provided additional 12mg docosahexaenoic acid (DHA), 20mg arachidonic acid (ARA)/100mL human milk]. Mothers provided milk samples (n=129) and reported maternal DHA consumption (n=128). Infant blood samples were drawn at study completion (Study Day 28). Human milk and infant PPL fatty acids were analyzed using capillary column gas chromatography. DHA and ARA were within ranges previously published for US term and preterm human milk. Compared to Control HMF (providing no DHA or ARA), human milk fortified with LHMF significantly increased infant PPL DHA and ARA and improved preterm infant DHA and ARA status.

Biochemical and white blood cell profiles of baboon neonates consuming formulas with moderate and high dietary long-chain polyunsaturated fatty acids.

BACKGROUND: Clinical chemistry and complete blood count (CBC) values were determined in 14 term baboons (Papio species) consuming formula with moderate or high levels of dietary long-chain polyunsaturated fatty acids (LCPUFA) from 2-12 weeks of age. METHOD: Neonates were randomized to three groups: C: Control, no LCPUFA; L: 0.33% docosahexaenoic acid (DHA)/0.67% arachidonic acid (ARA) (w/w); L3:1.00% DHA/0.67% ARA (w/w). Blood chemistries were assessed at 6 and 12 weeks and CBC parameters were measured at 2, 4, 8, 10, 12 weeks of age. RESULTS: Dietary LCPUFA had significant effects on serum triglyceride (C > L,L3) and calcium (L > C,L3). No other significant effects of diet were detected; pooled values are presented for all other parameters. CONCLUSION: These data provide longitudinal biochemical and white cell/platelet/immunological data on LCPUFA-fed baboons over the first 12 weeks of life. Data ranges are similar to reference data in cases for which values exist and hematological changes reflect trends observed during human neonatal development.

Negligible changes in piglet serum clinical indicators or organ weights due to dietary single-cell long-chain polyunsaturated oils.

Single-cell oils are currently included in human infant formula as sources of the long-chain polyunsaturates (LCP) docosahexaenoic acid (DHA) and arachidonic acid (AA) in many countries, but have not yet been approved for use in the USA. We prepared four bovine-milk-based formulas with AA/DHA=0, 34/17, 68/34 and 170/85 (mg per 100 kcal formula) provided by two commercial single-cell oils. These levels correspond approximately to 0, 1, 2 and 5 times the concentrations used in infant formulas and, due to greater consumption of formula per unit body weight, resulted in daily consumption of approximately 0, 3, 6 and 16 times those anticipated for human infants. All other dietary fat (47% of calories) was provided by a vegetable oil blend used in commercial human infant formulas. Domestic piglets were allowed to nurse with the sow for 24 h after parturition, then removed to individual cages and maintained on one of the four diets. At 30 days of age the piglets were sacrificed, and serum collected and organs weighed. With litters treated as a blocked variable, no significant differences among groups were found by analysis of variance for the following serum assays: alkaline phosphatase, alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood urea nitrogen (BUN), creatinine, albumin, glucose, cholesterol, triglycerides, and total protein. No significant differences were found for hematocrit or body weight. No significant differences were found among groups for weights of liver, brain, heart, lung, spleen, kidneys or lung, analyzed as absolute weight and as a fraction of body weight. Hematoxylin/eosin liver sections examined by light microscopy showed no abnormalities as evaluated by an independent pathologist. DHA content in liver and heart and AA content in heart showed significant dose-related accumulation (P<0.05) and confirmed enhanced tissue accretion of DHA and AA from both oils. We conclude that single-cell oils in formula consumed for 1 month in amounts up to 16-fold greater than proposed for human infants in the USA did not result in clinical chemistry or histopathologic indications of toxic effects in neonatal pigs.

Evaluation of single-cell sources of docosahexaenoic acid and arachidonic acid: 3-month rat oral safety study with an in utero phase.

Owing to the presence of the polyunsaturated fatty acids (PUFA) docosahexaenoic acid (DHA) and arachidonic acid (ARA) in human milk and their important biological function, several authorities recommend that they be added to infant formulas. This study assessed the safety of an algal oil rich in DHA and a fungal oil rich in ARA, blended to provide a DHA to ARA ratio similar to human milk. The oil blend was incorporated into diets and fed to rats such that they received 3, 11 and 22 times the anticipated infant exposure to DHA and ARA. Low-fat and high-fat control groups received canola oil. Rats received experimental diets over a premating interval and throughout mating, gestation and lactation. Pups born during this period (F1) consumed treatment diets from weaning for 3 months. Physical observations, ophthalmoscopic examinations, body weight, food intake, clinical chemistry, neurobehavioural evaluations and postmortem histopathology of selected tissues were performed. No statistically significant, dose-dependent adverse effects were seen in reproductive performance or fertility, nor in the neonates from birth to weaning. Mid- and high-dose treated F1 animals exhibited increased white cell count, neutrophil count and blood urea nitrogen; increased liver and spleen weights (absolute and relative to body weight) also were observed. There were no corresponding microscopic findings. The clinical pathology and organ weight differences at these treatment levels represent physiological or metabolic responses to the test substance rather than adverse responses. These single-cell oils produced no adverse effects in rats when administered in utero and for 90 days at dietary levels resulting in exposures up to 22 or 66 times higher than those expected in infant formulas when extrapolated on the basis of diet composition (g/100 Cal) or intake (g/kg body weight), respectively.

Visual acuity and blood lipids in term infants fed human milk or formulae.

This multicenter, parallel group study determined plasma phospholipid and red blood cell (RBC) phosphatidylcholine and phosphatidylethanolamine fatty acids, plasma cholesterol, apo A-1 and B, growth and visual acuity (using the acuity card procedure) in term infants fed from birth to 90 d of age with formula containing palm-olein, high oleic sunflower, coconut and soy oil (22.2% 16:0, 36.2% 18:1, 18% 18:2n-6, 1.9% 18:3n-3) (n = 59) or coconut and soy oil (10.3% 16:0 18:6% 18:1, 34.2% 18:2n-6, 4.7% 18:3n-3) (n = 57) or breast-fed (n = 56) with no formula supplementation. Different centers in North America were included to overcome potential bias due to differences in n-6 or n-3 fatty acids at birth or in breast-fed infants that might occur in a single-site study. Plasma and RBC phospholipid docosahexaenoic acid (DHA, 22:6n-3) and arachidonic acid (AA, 20:4n-6), cholesterol and apo B were significantly lower in the formula- than breast-fed infants. There were no differences in looking acuity or growth among the breast-fed and formula-fed infants. No significant relations were found between DHA and looking acuity, or AA and growth within or among any of the infant groups. This study provides no evidence to suggest the formula provided inadequate n-6 or n-3 fatty acids for growth and looking acuity for the first 3 mon after birth.

Evaluation of single cell sources of docosahexaenoic acid and arachidonic acid: a 4-week oral safety study in rats.

Docosahexaenoic acid (DHA) and arachidonic acid (ARA) are secreted in human milk and consumed by the nursing neonate but are not present in infant formulas currently available in the US. Supplementation of formulas with DHA and ARA may be particularly important for premature infants, who have less accretion of these fatty acids in utero than term infants. Some experts suggest that DHA and ARA should be added to infant formulas. Common sources of these fatty acids (e.g. fish oils, egg yolk lipids) are not optimal for infants in that they contain disproportionate amounts of other fatty acids. This 4-wk study examined the safety of a high-DHA algal oil and a high-ARA fungal oil, blended so that the DHA:ARA ratio approximates that in human milk. Rats were fed the blend at levels representing three, 11 and 22 times the anticipated infant exposure. Control animals were fed either a high-fat diet (13.1%, w/w; equivalent to the fat content of the treated groups) or a low-fat diet (5%, w/w). There were no treatment-related differences in body weight, food intake, organ weights, haematology or clinical chemistry. Thus, this study indicates that a blend of algal and fungal oils is a safe source of DHA and ARA as it produced no adverse effects in rats when administered for 4 wk at levels up to 22 times the expected infant exposure.

Dietary triacylglycerol structure and saturated fat alter plasma and tissue fatty acids in piglets.

Human and pig milk triacylglycerols contain a large proportion of palmitic acid (16:0) which is predominately esterified in the 2-position. Other dietary fats contain variable amounts of 16:0, with unsaturated fatty acids predominantly esterified in the 2-position. These studies determined if the amount or position of 16:0 in dietary fat influences the composition or distribution of liver, adipose tissue, lung, or plasma fatty acids in developing piglets. Piglets were fed to 18 d with sow milk or formula with saturated fat from medium-chain triglyceride (MCT), coconut or palm oil, or synthesized triacylglycerols (synthesized to specifically direct 16:0 to the 2-position) with, in total fatty acids, 30.7, 4.3, 6.5, 27.0, and 29.6% 16:0, and in 2-position fatty acids, 55.3, 0.4, 1.3, 4.4, and 69.9% 16:0, respectively. The percentage of 16:0 in the 2-position of adipose fat from piglets fed sow milk, palm oil, and synthesized triacylglycerols were similar and higher than in piglets fed MCT or coconut oil. Thus, the amount, not the position, of dietary 16:0 determines piglet adipose tissue 16:0 content. The effects of the diets on the plasma and liver triacylglycerols were similar, with significantly lower 16:0 in total and 2-position fatty acids of the MCT and coconut oil groups, and significantly higher 16:0 in the plasma and liver triacylglycerol 2-position of piglets fed the synthesized triacylglycerols rather than sow milk or palm oil. The lung phospholipid total and 2-position 16:0 was significantly lower in the MCT, coconut, and palm oil groups, but similar in the synthesized triacylglycerol group and sow milk group. The lung phospholipid total and 2-position percentage of arachidonic acid (20:4n-6) was significantly lower in all of the formula-fed piglets than in milk-fed piglets. The physiological significance of this is not known.

Stereospecific analysis of triacylglycerols rich in long-chain polyunsaturated fatty acids.

Six oils of marine, algal, and microbial origin were analyzed for stereospecific distribution of component fatty acids. The general procedure involved preparation of sn-1,2-(2,3)-diacylglycerols by partial deacylation with ethylmagnesium bromide or pancreatic lipase, separation of X-1,3- and sn-1,2(2,3)-diacylglycerols by borate thin-layer chromatography, resolution of the sn-1,2- and sn-2,3-enantiomers by chiral phase high-performance liquid chromatography following preparation of dinitrophenylurethane derivatives, and determination of the fatty acid composition by gas chromatography. Unexpected complications arose during a stereospecific analysis of triacylglycerols containing over 33% of either 20:4 or 22:6 fatty acids. The sn-1,2(2,3)-diacylglycerols made up of two long-chain polyunsaturated acids migrated with the X-1,3-diacylglycerols and required separate chiral phase resolution. Furthermore, the enzymatic method yielded sn-1,2(2,3)-diacylglycerols, overrepresenting the polyenoic species due to their relative resistance to lipolysis, but prolonged digestion yielded correct composition for the 2-monoacylglycerols. The final positional distribution of the fatty acids was established by pooling and normalizing the data from subfractions obtained by normal- and chiral-phase separation of diacylglycerols. The molecular species of X-1,3-, sn-1,2- and sn-2,3-diacylglycerol dinitrophenylurethanes were identified by chiral-phase liquid chromatography/mass spectrometry with electrospray ionization, which demonstrated a preferential association of the paired long-chain acids with the sn-1,2- and sn-2,3-diacylglycerol isomers.

Palmitic acid is absorbed as sn-2 monopalmitin from milk and formula with rearranged triacylglycerols and results in increased plasma triglyceride sn-2 and cholesteryl ester palmitate in piglets.

Milk fatty acids contain 20-30% palmitic acid (16:0), with approximately 70% of the 16:0 esterified to the sn-2 position of the milk triacylglycerol. Formulae containing vegetable and oleo oils contain different amounts of 16:0, but all have unsaturated fatty acids esterified to the triacylglycerol sn-2 position. Intraluminal triacylglycerol hydrolysis by endogenous lipases produces sn-2 monoacylglycerols and free fatty acids, which are absorbed and re-esterified in the enterocyte for secretion to plasma. The extent of absorption and re-esterification of sn-2 monoacylglycerols from milk or formula fats in infants is unknown. This was studied by feeding piglets sow milk or formulae containing similar total saturated fat, 18:1, 18:2(n-6) and 18:3(n-3) with unsaturated fatty acids at the sn-2 position or with rearranged triacylglycerols containing approximately 30% 16:0, with 70% 16:0 in fatty acids at the sn-2 position. Feeding milk or 16:0 on the sn-2 position of formula with rearranged triacylglycerols resulted in higher 16:0 esterified to the plasma triacylglycerol sn-2 position and in cholesteryl esters than feeding formulae with 0.4 to 4.4% 16:0 in the sn-2 position fatty acids. The absorption of 16:0 as monopalmitin from milk and any metabolic importance in human infants has yet to be determined.

Plasma and tissue lipids of piglets fed formula containing saturated fatty acids from medium-chain triglycerides with or without fish oil.

Addition of fish oil (FO) with carbon chain 20 and 22 (long-chain polyunsaturated fatty acids; LCPs) n-3 fatty acids to formula has been considered as a method to provide 22:6n-3 to preterm infants. These studies determined the effects of including 0.73% n-3 LCPs from FO in formula containing saturated fatty acids as medium-chain triglyceride (MCT) oil on plasma and organ triglyceride (TG), phospholipid (PL), and cholesterol ester (CE) contents and compositions in piglets. Reference piglets were fed their own mother's milk. Piglets fed the formula with FO had significantly lower liver and kidney, but not plasma, TG concentrations; lower liver PL 20:4n-6; and higher plasma, liver, and kidney TG, PL, and CEn-3 LCPs than piglets fed the formula without FO. The increase in liver and kidney lipid n-3 LCPs and the decrease in kidney lipid content shows that inclusion of FO in formula with a high content of MCTs and 18:2n-6 does not promote tissue lipid metabolism similar to natural milk feeding. Piglets fed formula with FO had similar brain 20:4n-6 and 20:5n-3 but higher 22:6n-3 than did piglets fed sow milk. Diet-related differences in plasma and red blood cell PL were not reliable predictors of differences in brain LCP.

Dietary saturated, monounsaturated, n-6 and n-3 fatty acids, and cholesterol influence platelet fatty acids in the exclusively formula-fed piglet.

Platelet lipid composition is important to normal platelet morphology and function, and is influenced by dietary fatty acids and cholesterol. The fatty acid composition and cholesterol content of infant formulas differs from those of human milk, but the possible effects on platelet lipids in young infants is not known. This was studied in piglets fed from birth to 18 d of age with one of eight formulas differing in saturated fatty acid chain length, or content of 18:1, 20:5n-3 plus 22:6n-3, or cholesterol. A reference group of piglets fed sow milk was also studied. Sow milk has a fatty acid composition and cholesterol content similar to that of human milk. Piglets fed formulas high in 18:1 (34.9-40.8% wt fatty acids) and low in 16.0 (< or = 6.5% wt fatty acids) had lower platelet counts and greater platelet size than piglets fed sow milk (40.4% 18:1, 30.7% 16:0). Piglets fed formulas high in 16:0 (27-29.6%) and 18:1 (40-40.6%), or low in both 16:0 (5.9-6.1%) and 18:1 (10.8-11.2%), had similar platelet counts and size to piglets fed sow milk. Platelet phospholipid % 20:4n-6 was lower in all the groups of piglets fed formula than in the group fed sow milk. Addition of fish oil with 20:5n-3 plus 22:6n-3 to the formula further decreased platelet phospholipid 20:4n-6. Addition of cholesterol to the formula increased the platelet phospholipid % 20:4n-6 and platelet volume.

Saturated fatty acid chain length and positional distribution in infant formula: effects on growth and plasma lipids and ketones in piglets.

Human milk contains a large proportion of palmitic acid (16:0) with > 70% esterified to the center sn-2 position of the milk triglyceride. Infant formulas often use 8:0 + 10:0 [medium-chain triglyceride (MCT)] or 12:0 + 14:0 (coconut oil) as the saturated fat. The effect of formula saturated fatty acid composition; 8:0 + 10:0, 12:0 + 14:0, or 16:0 from palm oil or synthesized triglyceride containing predominantly sn-2 16:0 on plasma lipids and fatty acids was studied in piglets. Although the formulas contained similar 18:1 and 18:2n-6, plasma lipid percentages of 18:1 and 18:2n-6 were higher in piglets fed the formula with MCT or coconut oil rather than the formulas with 16:0, or sow milk. The sn-2 16:0 of the synthesized triglyceride had unique properties. Specifically, piglets fed synthesized triglyceride had significantly higher cholesteryl ester 16:0 identical to that in piglets fed sow milk and higher plasma total and high-density-lipoprotein cholesterol than piglets fed the other formulas.

Nonessential fatty acids in formula fat blends influence essential fatty acid metabolism and composition in plasma and organ lipid classes in piglets.

The n-6 and n-3 fatty acid status of developing organs is the cumulative result of the diet lipid composition and many complex events of lipid metabolism. Little information is available, however, on the potential effects of the saturated fatty acid chain length (8:0-16:0) or oleic acid (18:1) content of the diet on the subsequent metabolism of the essential fatty acids 18:2n-6 and 18:3n-3 and their elongated/desaturated products. The effects of feeding piglets formulas with fat blends containing either coconut oil (12:0 + 14:0) or medium chain triglycerides (MCT, 8:0 + 10:0) but similar levels of 18:1, 18:2n-6 and 18:3n-3, or MCT with high or low 18:1 but constant 18:2n-6 and 18:3n-3 on the fatty acid composition of plasma, liver and kidney triglycerides, phospholipids and cholesteryl esters, and of brain total lipid, were studied. Diet-induced changes in the fatty acid composition of lipid classes were generally similar for plasma, liver and kidney. Dietary 18:1 content was reflected in tissue lipids and was inversely associated with levels of 18:2n-6. Lower percentage of 18:2n-6, however, was not associated with lower levels of its elongated/desaturated product 20:4n-6 but was associated with higher levels of 22:6n-3. Feeding coconut oil vs. MCT resulted in lower 18:1 levels in all lipids, and higher percentages of 20:4n-6 in tissue phospholipid. Increasing the dietary n-6/n-3 ratio from 5 to 8 significantly increased tissue percentage of 18:2n-6 and decreased phospholipid 22:6n-3.(ABSTRACT TRUNCATED AT 250 WORDS)

Dietary saturated fatty acids (12:0, 14:0, 16:0) differ in their impact on plasma cholesterol and lipoproteins in nonhuman primates.

Three species of monkey (rhesus, cebus, and squirrel) were rotated through five purified diets containing 31% energy as various fat blends (P:S between 0.1 and 1.0) for 12-wk periods to compare the impact of specific dietary fatty acids on plasma lipids and lipoproteins. As 12:0 + 14:0 was replaced by 16:0, a significant decrease occurred in total and LDL cholesterol, whereas slight increases in total cholesterol and the LDL-HDL ratio occurred when 16:0 replaced 18:2. Hegsted and Keys regression equations provided a good fit for the observed data, but the predicted total cholesterol response was perfect (r = 0.995) for both equations when 16:0 was considered neutral. Thus, under these conditions 16:0 was less cholesterolemic than 12:0 + 14:0 and only slightly cholesterolemic compared with 18:2.

No effect of long-term DHEA treatment on either hepatocyte and adipocyte pentose pathway activity or adipocyte glycerol release.

The effects of long-term dehydroepiandrosterone treatment on the pentose shunt in adipocytes and hepatocytes from female lean Zucker rats were determined. No significant effects could be attributed to this treatment. There was also no effect on basal or epinephrine-stimulated glycerol release from adipocytes.

Plasma, tissue and fecal cholesterol of young pigs fed restricted or liberal amounts of beef, soy or conventional diets.

Cholesterol disposition (tissue deposition + fecal excretion) was determined in young pigs fed restricted (trial 1) or liberal (trial 2) amounts of beef-based, soy-based or conventional swine diets. Cholesterol content of diet is greater than average United States consumption per kilogram body weight but is very similar on a per-kilocalorie intake basis. Both beef and soy diets contained more cholesterol (0.09% by weight vs. 0%) and fat (40-50% of calories vs. 8-9%) than did conventional diets. Dried egg yolk was the source of cholesterol in the soy diets. Beef- and soy-fed pigs had greater plasma, HDL and LDL cholesterol concentrations than did conventionally fed pigs; beef-fed pigs had greater plasma, HDL and LDL cholesterol concentrations than did soy-fed pigs in trial 2 only. Final HDL-to-LDL cholesterol ratios did not differ. Neutral steroid and bile acid excretion was twofold greater in soy-fed than in conventionally or beef-fed pigs in trial 2 only. Differences in cholesterol concentrations were found in liver (soy, beef greater than conventional) and heart (soy greater than beef, conventional) in trial 1 and in liver (soy, beef greater than conventional), adipose tissue (soy greater than conventional greater than beef) and aorta and other viscera (conventional greater than soy, beef) in trial 2. In both trials, soy-fed pigs had greatest whole-body cholesterol concentrations. Thus, in both trials, disposition of cholesterol was similar in conventionally fed and beef-fed pigs, despite greater cholesterol intake in the latter, but was greater soy-fed pigs.

Effects of beef, soy and conventional diets on body composition and plasma lipids of young pigs fed restricted or liberal amounts of diet.

Effects were determined of restricted and liberal feeding of beef-based, soy-based and conventional diets on growth rate, nutrient absorption, body composition and plasma lipid and urea concentrations of young pigs. Beef and soy diets contained more fat (40-50% of calories vs. 8-9%) and cholesterol (0.09 vs. 0%) than did conventional diets; calorie and protein intakes were equal across diets. Beef-fed pigs had greater average daily gain and absorption of fat and gross energy than did conventionally fed pigs; soy-fed pigs had intermediate growth rates but the greatest absorption of fat and gross energy. With restricted intakes, percentage of body fat was greater in soy-fed pigs than in pigs fed a conventional diet or beef and with liberal feeding, greater in soy- and beef-fed pigs than in pigs provided with a conventional diet. Plasma free fatty acid, triglyceride and cholesterol concentrations were greater in soy- and beef-fed pigs than in conventionally fed pigs. These results indicate that absorption and subsequent partitioning of nutrients toward body fat are influenced by composition as well as the amount of diet consumed. Our results also demonstrate that the pig can successfully be fed diets resembling human diets (i.e., high fat) and because of its physiological similarities to humans our understanding of nutrition and lipid metabolism of humans may be advanced.

Effects of dietary calcium and fat on cholesterol in tissues and feces of young goats.

To examine the effects of amount of calcium and type of fat in the diet on distribution of cholesterol in plasma, several tissues and feces, 4-week-old goats (6 females and 18 males) were fed isocaloric diets consisting of goat milk supplemented with cholesterol and 1) soybean oil, 2) tallow, 3) soybean oil and CaCO3 or 4) tallow and CaCO3 for 20 weeks. Plasma cholesterol increased initially in all animals; plasma and lipoprotein cholesterol concentrations, however, were not affected by dietary fat or calcium. Goats fed CaCO3 excreted more calcium and ash in their feces and had greater cholesterol and fat concentrations in their livers. Neutral steroid, but not bile acid, excretion was greater in soybean oil-fed goats than in goats fed tallow. Soybean oil-fed goats had greater cholesterol concentrations in the viscera (minus liver) and tended to have greater cholesterol concentrations in the carcass and whole body. Aortas of soybean oil-fed goats exhibited greater fat deposition. In summary, dietary fat, but not calcium, significantly altered cholesterol disposition in young goats by affecting cholesterol deposition in tissues and excretion in feces, without a concomitant change in plasma cholesterol.