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Research Notes: Essential Fatty AcidsMed Hypotheses. 2007 Jul 24. With the exception of melanoma and non-Hodgkin's lymphoma, the incidence of cancer has peaked in the last several years, but rates and mortality are still high. Moreover, despite 50 years of intensive cancer research increasingly focused on genetic causes, no single unifying cause for cancer has been established. Although it is well-known that tumors are hypoxic, and that there is a correlation between the level of hypoxia and prognosis, with the exception of Warburg's studies, little work has been done to investigate the relationship between hypoxia and cancer. Over 70 years ago, Warburg showed that cells could always be made cancerous by subjecting them to periods of hypoxia. Moreover, he demonstrated that once cells had converted to a cancerous state, reversion could not occur. Modern biochemistry acknowledges that there is a switch from oxidative phosphorylation to glycolysis in tumors that might be concurrent with hypoxia, but does not address the cancer causation. It is our hypothesis that long-term hypoxia of cells in the body, measured in years, is the primary trigger for cancer. We believe that the hypoxia, which has to meet Warburg's findings of a critical 35% reduction in intracellular oxygen levels to initiate cancer, is linked to the incorporation of adulterated, non-oxygenating, or inappropriate polyunsaturated fatty acids (PUFAs) into the phospholipids of cell and mitochondrial membranes. Such incorporation causes changes in membrane properties that impair oxygen transmission into the cell. Trans fats, partially oxidized PUFA entities, and inappropriate omega-6:omega-3 ratios are all potential sources of unsaturated fatty acids that can disrupt the normal membrane structure. In this paper, we explore this hypothesis by examining the evidence, and additionally propose an appropriate PUFA dosage for humans by analyzing requirements and taking into account current PUFA consumption patterns. J Pediatr Psychol. 2006 Mar. Objective. To assess whether natural variations in decosahexaenoic acid (DHA) levels in breast milk would relate to infant neurobehavioral outcomes at the newborn stage following equivocal findings on infant and toddler outcomes of exposure to DHA in formula and breast milk. Methods. Breast milk samples from N = 20 mothers were collected 9 days after delivery, while the Brazelton Neonatal Behavioral Assessment Scale (NBAS) was administered on the infant. Milk samples were later analyzed for fatty acids, including DHA. Results. Pearson correlations revealed a positive association between DHA concentrations in breast milk and infants’ scores on the NBAS Range of State cluster score, suggesting that DHA is related to the infant’s superior ability to maintain optimal arousal. Conclusions. These results suggest that breast milk DHA is beneficial to the neonate’s neurobehavioral functioning and call for investigative attention to DHA’s role in potentiating optimized intellectual functioning at later stages of development. The findings may also be interpreted as supporting formula supplementation with DHA. J Lipid Res. 2006 Feb. This study was conducted to evaluate the chronic effects of eicosapentaenoic acid (EPA) on fatty acid and glucose metabolism in human skeletal muscle cells. Uptake of [14C]oleate was increased >2-fold after preincubation of myotubes with 0.6 mM EPA for 24 h, and incorporation into various lipid classes showed that cellular triacylgycerol (TAG) and phospholipids were increased 2- to 3-fold compared with control cells. After exposure to oleic acid (OA), TAG was increased 2-fold. Insulin (100 nM) further increased the incorporation of [14C]oleate into all lipid classes for EPA-treated myotubes. Fatty acid beta-oxidation was unchanged, and complete oxidation (CO2) decreased in EPA-treated cells. Basal glucose transport and oxidation (CO2) were increased 2-fold after EPA, and insulin (100 nM) stimulated glucose transport and oxidation similarly in control and EPA-treated myotubes, whereas these responses to insulin were abolished after OA treatment. Lower concentrations of EPA (0.1 mM) also increased fatty acid and glucose uptake. CD36/FAT (fatty acid transporter) mRNA expression was increased after EPA and OA treatment compared with control cells. Moreover, GLUT1 expression was increased 2.5-fold by EPA, whereas GLUT4 expression was unchanged, and activities of the mitogen-activated protein kinase p38 and extracellular signal-regulated kinase were decreased after treatment with OA compared with EPA. Together, our data show that chronic exposure of myotubes to EPA promotes increased uptake and oxidation of glucose despite a markedly increased fatty acid uptake and synthesis of complex lipids. J Nutr Biochem. 2005 Mar. Objectives: The aim of the study was to investigate whether fish oil supplementation affected Japanese schoolchildren's behavior, with changes in aggression over time as the primary endpoint. Design and subjects: A placebo-controlled double-blind study with 166 schoolchildren 9-12 years of age was performed. The subjects of the fish oil group (n=83) took fish oil-fortified foods (bread, sausage and spaghetti). These foods were provided in amounts such that each subject in the fish oil group had an intake of 3600 mg of docosahexaenoic acid + 840 mg of eicosapentaenoic acid (EPA)/week for 3 months. The rest (the controls, n=83) took control supplements. At the start and end of the study, psychological tests were performed to assess their aggression. Results: Physical aggression assessed by Hostility-Aggression Questionnaire for Children in girls increased significantly (median: 13 to 15, n=42) in the control group and did not change (13 to 13, n=43) in the fish oil group with a significant intergroup difference (P=.008) with baseline as covariate. The changes in physical aggression scores over time and those of the ratio of EPA/arachidonic acid in RBC (DeltaEPA/AA) were significantly correlated in girls who agreed to blood collection (r=-.53, P=.01, n=23). On the contrary, there were no significant changes in physical aggression in boys. Aggression against others (extraggression) assessed by Picture Frustration Study did not change in the control group (median: 5 to 5) but increased significantly in the fish oil group (4 to 5) with a significant intergroup difference (P=.02) with baseline as covariate. These changes in extraggression might be explained partly by significantly lower baseline values of extraggression in the fish oil group (P=.02) than in the control group. There were no significant correlations between Deltaextraggression and DeltaEPA/AA in blood-sampled children (n=49). Impulsivity of girls assessed by parents/guardians using the diagnostic criteria for attention deficit/hyperactivity disorder of DSM-IV was reduced in the fish oil group (1 to 0) with a significant (P=.008) intergroup difference from the control group (1 to 1). There were no significant correlations between Deltaimpulsivity and DeltaEPA/AA in blood-sampled girls. In males, impulsivity reduced in both groups without any intergroup differences. Conclusions: There is a possibility that changes in fatty acid nutrition might affect physical aggression especially in girls. J Nutr. 2004 Sep. Between 6 and 12 mo of age, blood levels of the (n-3) long-chain PUFA, docosahexaenoic acid (DHA), in breast-fed infants typically decrease due to diminished maternal DHA stores and the introduction of DHA-poor solid foods displacing human milk as the primary source of nutrition. Thus, we utilized a randomized, clinical trial format to evaluate the effect of supplemental DHA in solid foods on visual development of breast-fed infants with the primary outcome, sweep visual-evoked potential (VEP) acuity, as an index for maturation of the retina and visual cortex. At 6 mo of age, breast-fed infants were randomly assigned to receive 1 jar (113 g)/d of baby food containing egg yolk enriched with DHA (115 mg DHA/100 g food; n = 25) or control baby food (0 mg DHA; n = 26). Gravimetric measures were used to estimate the supplemental DHA intake which was 83 mg DHA/d in the supplemented group and 0 mg/d in controls. Although many infants in both groups continued to breast-feed for a mean of 9 mo, RBC DHA levels decreased significantly between 6 and 12 mo (from 3.8 to 3.0 g/100 g total fatty acids) in control infants, whereas RBC DHA levels increased by 34% from 4.1 to 5.5 g/100 g by 12 mo in supplemented infants. VEP acuity at 6 mo was 0.49 logMAR (minimum angle of resolution) and improved to 0.29 logMAR by 12 mo in controls. In DHA-supplemented infants, VEP acuity was 0.48 logMAR at 6 mo and matured to 0.14 logMAR at 12 mo (1.5 lines on the eye chart better than controls). At 12 mo, the difference corresponded to 1.5 lines on the eye chart. RBC DHA levels and VEP acuity at 12 mo were correlated (r = -0.50; P = 0.0002), supporting the need of an adequate dietary supply of DHA throughout 1 y of life for neural development. Selections from the full text article: Long-chain PUFA (LCPUFA) have an important role in visual development during infancy. Compared with infants fed commercial formulas lacking LCPUFA, breast-fed infants have more advanced electroretinographic function as early as 6 wk of age and more mature visual acuity by 4 mo of age. In longitudinal assessment of the effect of maternal and infant dietary factors in infant visual development, Williams et al. reported that the variable most associated with stereoacuity at 3.5 y of age was breast-feeding. Children who had nursed for even short periods during infancy had more mature visual stereoacuity than children who had never received human milk. The (n-3) LCPUFA, docosahexaenoic acid [DHA; 22:6(n-3)], which is present in human milk but absent in unsupplemented infant formulas, may be the major factor responsible for this benefit. Direct evidence for a role of DHA in visual development is that term infants fed formula with adequate amounts of DHA and a balanced amount of the 20-carbon (n-6) LCPUFA, arachidonic acid [ARA; 20:4(n-6)], have improved visual and mental development with no adverse effect on growth. Furthermore, DHA in infant formula was associated with shorter-look duration to novel stimuli on the Fagan test and with improved visual acuity in a multistudy meta-analysis. Critical reviews of this literature were published recently. A woman producing milk for her infant derives a major portion of milk LCPUFA from her endogenous stores. Human milk can vary considerably in its LCPUFA content depending on the diet of the mother and the amount of LCPUFA mobilized from her tissues over the course of the current and any preceding pregnancies and/or lactations. The concentration of DHA in human milk varies from as little as 0.1% of total fatty acids in women consuming Western diets to as much as 1.4% in Inuit women in North America and 2.78% in Chinese women from a fishing village, both consuming large amounts of marine animal foods. The rationale for this study was that at 6 mo of age, infants are beginning to be fed semisolid foods and thus are likely to have a reduction in dietary DHA as reflected in decreased blood DHA levels. This reduction in the infant's DHA intake may be due to a concomitant reduction in consumption of human milk combined with increasing intake of DHA-poor weaning foods. The objective of this randomized clinical trial was to determine whether DHA-enriched baby food provided as a supplemental source of DHA to breast-fed infants in the second 6 mo of life altered blood lipid fatty acid profiles and modified visual development. In addition, to assess whether the long-chain fatty acids affected infant metabolism, we evaluated total antioxidant capacity, blood chemistry, and hematology. Supplementary DHA was provided in the form of ready-to-feed baby foods made with DHA-enriched egg yolk providing DHA and ARA. Subjects. Healthy term infants receiving human milk born at either Presbyterian Hospital of Dallas or Medical City Dallas Hospital were enrolled in the study. Additional infants were recruited through advertisements. Inclusion criteria were a gestational age at birth >37 wk, a birthweight > 2800 g, exclusive breast-feeding in hospital and for the first 4 mo of life with a maternal intention to continue breast-feeding, a good possibility of long-term follow-up, and informed consent to the protocol. Exclusion criteria were any underlying disease or congenital malformation that was judged likely to interfere with the evaluation of the study material, any abnormal maternal dietary patterns, and any evidence of maternal metabolic disease. [...] Computer-generated randomization codes with variable-length blocks of 8–12 were used to assign 55 infants to 1 of 2 groups at 6 mo of age. Both groups received study baby foods and were directed to feed the baby 1 jar of study food per day. As incentive, all parents received store coupons for purchasing commercial baby foods at their local store. One group received control baby foods, and the other group received baby foods containing DHA-enriched egg yolk. Fifty-one infants completed the study: 26 in the group receiving the control baby food, with 1 drop-out (due to viral infection), and 25 in the group receiving the baby foods made with DHA-enriched egg yolks with 3 drop-outs (2 due to constipation and 1 refusal to eat solid foods at 6 mo of age), yielding a 93% completion rate. Baby foods. DHA-enriched eggs were obtained from hens receiving a diet essentially free of LCPUFA but containing flaxseed and soybean meal as sources of the DHA precursor, -linolenic acid (-LNA). The egg yolks were separated, pasteurized, and spray-dried. The dried egg yolks at 120 g/kg food (12%) were used to prepare semisolid, ready-to-feed DHA-enriched baby foods as described by Theuer et al.; all foods were packaged in hermetically sealed jars containing 113 g food. The control baby foods were devoid of egg yolk but otherwise contained the same ingredients as the DHA-enriched foods. The dried egg yolk and the baby foods were analyzed for fatty acids by Medallion Laboratories (see Table 1). The dried egg yolk contained 2% of fatty acids as DHA; the DHA-enriched baby foods contained 115 mg DHA/100 g food (i.e., 130 mg DHA/113-g baby food jar). The fat content of the different flavors of DHA-enriched baby foods (5.8–8.1 g/100 g) was 5- to 6-fold higher than that of control foods (0.1–2.3 g/100 g), resulting in a higher energy density of the DHA-enriched baby foods (greater by 230 kJ/100 g). [...] Results Despite the expressed intention to continue breast-feeding their infants, mothers in the control and DHA-enriched groups breast-fed for 9.7 mo and 8.8 mo, respectively (see Table 2); 65% of the infants in the control group and 80% of infants in the supplemented group were weaned from human milk to formula before 12 mo of age. The study was largely completed before the commercial availability of infant formula fortified with LCPUFA in the United States. Only 1 infant in the control group and 3 in the DHA-enriched group were weaned from human milk to infant formula containing DHA and ARA during the 6-mo trial interval. Removal of these infants from data analysis did not affect the results of the statistical analyses. Dietary sources of DHA for infants in the study included both human milk and enriched baby foods. The estimated intakes of DHA from human milk during the trial were 37 mg/d in control infants and 28 mg/d in the supplemented group. Consumption of baby food by the control group was 84 ± 23 g food/d (0.75 jar/d; Table 2). Based on gravimetric measures, control infants consumed 0 and 0.3 mg supplemental DHA and ARA/d, respectively, from baby food during the 6-mo study. Infants in the DHA-enriched group consumed 72 ± 31 g baby food/d (about 0.66 jar/d); this was not different from controls (P = 0.12). Infants fed DHA-enriched baby foods were estimated to have consumed 83 mg supplementary DHA/d and 56 mg supplementary ARA/d during the 6-mo trial. The fatty acid content of RBC lipids did not differ between groups at the start of the trial (6 mo; Table 3); however, by 12 mo, the groups differed in RBC lipid DHA, docosapentaenoic acid [DPA; 22:5(n-6)], and total (n-3) LCPUFA (P < 0.002). RBC DHA levels decreased in the control group from 3.8% at 6 mo to 3.0% at 12 mo (P = 0.012). In contrast, RBC DHA levels increased (P < 0.002) in the DHA-enriched group from 4.1% at 6 mo to 5.5% at 12 mo. RBC DHA levels expressed as mass concentration showed similar changes (Table 3, P < 0.002). The sufficiency indices for (n-3) fatty acids ([(n-6)/(n-3) LCPUFA ratio], DHA [DHA/DPA(n-6)], and essential fatty acids (Mead acid [20:3(n-9)]/ARA) improved in the DHA-supplemented group. The unsaturation index was significantly elevated in RBC of supplemented infants; this summation of double bonds is reflective of an increase in the fluidity of RBC membranes. Treen et al. reported that an increase in the unsaturation index of 10 units (based on the percentage of total fatty acids) in cultured retinoblastoma cells increased the lateral mobility in the membrane bilayer (i.e., an 30% increase in pyrene eximer formation) and increased transport of choline across the cell membrane by 12%. In control infants, VEP acuity at 6 mo was 0.49 ± 0.13 logMAR; it improved to 0.45 ± 0.14 logMAR at 9 mo and to 0.29 ± 0.11 logMAR at 12 mo (Fig. 1). In the DHA-supplemented group, VEP acuity was 0.48 ± 0.10 logMAR at 6 mo and improved to 0.31 ± 0.13 log MAR at 9 mo and to 0.13 ± 0.1 logMAR at 12 mo. Compared with controls, infants in the DHA-supplemented group had improved visual acuity by 0.14 and 0.16 logMAR at 9 and 12 mo, respectively, (P < 0.002), equivalent to 1.5 lines on an eye chart. VEP acuity at 12 mo was correlated with RBC DHA levels at 12 mo (r = –0.50; P = 0.0002) (Fig. 2) such that infants with high DHA levels had lower logMAR values, i.e., more mature acuity. In addition, the estimated dietary intake of DHA from human milk and baby food on an individual basis was correlated with VEP acuity (r = –0.49, P < 0.0002) as well as with RBC DHA levels (r = 0.57, P < 0.0002). Stereoacuity at 6 mo was 2.54 ± 0.54 log arc s and improved to 2.25 ± 0.47 log arc s at 12 mo in the control infants. In the supplemented infants, stereoacuity was 2.37 ± 0.34 log arc s at 6 mo and improved to 2.22 ± 0.55 log arc s at 12 mo. However, stereoacuity at 12 mo did not differ between the 2 diet groups (P = 0.8). Despite differences in energy and fat content of the study baby foods, the groups did not differ in weight, length, head circumference, or skin-fold thicknesses at 6, 9, and 12 mo (P > 0.3 for all measures; data not shown). Total plasma antioxidant capacity did not differ between the 2 diet groups at the onset of the trial (345 ± 78 µmol/L for controls and 328 ± 107 µmol/L Trolox equivalents for the supplemented group; P = 0.53) or at 12 mo (335 ± 63 vs. 321 ± 109 µmol/L Trolox equivalents; P = 0.58, respectively). Protocol compliance was excellent and the DHA-enriched foods were well tolerated. There were 3 adverse events recorded for controls: 2 were not diet-related (neuroblastoma and occluded tear duct, both requiring surgery), and 1 infant had a 3-fold elevation in aspartate aminotransferase at 12 mo. In the supplemented group, there were 3 adverse events: 2 were not diet-related (genetically associated elevation in alkaline phosphatase and eczema since birth) and 1 infant had a 5-fold elevation in alkaline phosphatase at 12 mo. All events were reported to the Institutional Review Boards and to patients' pediatricians. The groups did not differ in hematological measures at either 6 or 12 mo of age (P > 0.1). Similarly, their blood chemistries did not differ at 12 mo (P > 0.15). Upon termination of the study, neither group had mean hematological results that were outside of the normal range, although both groups had levels of creatinine (for control and supplemented groups, 32.2 and 30.4 µmol/L, respectively) and carbon dioxide (18.2 and 18.5 mEq/L) that were marginally below normal and albumin levels (43.6 and 42.8 g/L) that were slightly higher than normal; none were of clinical importance. Discussion In the current randomized clinical trial, breast-fed infants receiving LCPUFA-enriched baby foods during mo 6–12 of life had an 83% elevation in RBC DHA levels (Table 3) resulting from an approximately 2-fold greater intake of DHA compared with unsupplemented infants (Table 2). In addition, DHA-supplemented infants had more mature VEP acuity than control infants at 9 and 12 mo of age (by 0.14 and 0.16 logMAR, i.e., 1.5 lines on an eye chart; Fig. 1). Furthermore, the blood lipid level of DHA was significantly correlated with VEP acuity such that infants with higher levels of RBC DHA had better visual acuity (Fig. 2). Metabolic measures were equivalent in both groups with no major diet-related adverse events. No benefit to stereoacuity attributable to DHA-enriched baby food was evident in the current trial; this is consistent with a previous trial using LCPUFA-enriched infant formula. In both of these studies, infants received human milk for the first 4–6 mo of life, which may have provided sufficient nutriture for optimal development of stereoacuity. However, in a separate trial in which infants were randomized to receive control formula or LCPUFA supplemented formula beginning at 6 wk of life, this environmental influence was evident at a 4-mo time point but not later. Thus, a "critical period of sensitivity" appears to occur up to 6 mo of age in the maturational susceptibility of stereoacuity to environmental influences (e.g., dietary factors). The biochemical and functional results from this study are consistent with an earlier randomized clinical trial of breast-fed infants weaned between 4 and 6 mo of age to receive either DHA + ARA-enriched or nonenriched infant formula. At 12 mo, infants fed a nonenriched diet had a 50% reduction in RBC-DHA concentration compared with weaning levels. In contrast, infants fed the LCPUFA-enriched formula had a 24% higher RBC DHA content compared with weaning levels and at 12 mo had a 1.5-fold higher DHA level than that in the nonenriched infant group. In this formula trial, we estimated that the supply of DHA was 0.2–0.4 g DHA/6 mo in the control group (primarily due to endogenous DHA synthesis from -LNA) compared with a dietary intake of 22 g DHA/6 mo in LCPUFA-supplemented infants. The 1-y-old supplemented infants had improved VEP acuity by 0.103 log MAR (1 line on the eye chart) compared with the nonsupplemented group. The average amount of human milk consumed each day between 6 and 9 mo of age decreases from 750 mL to about 625 mL (17). Because the average fat content of human milk is 37 g/L (17), the daily intake of human milk fat over this period would be 25 g. With an average DHA content of human milk fat in the United States of 0.2 g/100 g total milk fatty acids (27,32), the DHA intake of exclusively breast-fed older infants in the United States would be 50 mg/d. Between 6 and 9 mo, the average dietary DHA intake of infants fed the baby foods made with DHA-enriched egg yolks was estimated to be 133 mg/d from both human milk (50 mg/d) and solid food sources (83 mg/d), whereas between 9 and 12 mo, the majority of infants were weaned and solid foods alone contributed DHA at an average of 83 mg/d. Thus, for the entire 6-mo trial period, the supplemented infants received an average of 108 mg DHA/d compared with 38 mg DHA/d in control infants who received only human milk until 9.7 mo of age. This corresponded to an approximate 2-fold increase in DHA intake by the supplemented group (7 vs. 20 g/6 mo; Table 2). Body weight over the 6- to 12-mo period averaged 8.4 kg; thus, the mean intake of DHA for the DHA-supplemented infants was 13 mg/(kg · d). However, intake for these infants from 6 to 9 mo while still breast-feeding was 17 mg/(kg · d) but dropped to 9 mg DHA/(kg · d) from 9 to 12 mo when the only source of DHA was from enriched baby foods. These amounts are 15 and 55% lower than the 20 mg DHA/(kg · d) recommended by the FAO/WHO Joint Expert Consultation. By comparison, the mean DHA intake for the control group during the 6-mo study was only 4.5 mg/(kg · d). Human milk and supplemented infant formula are among the few foods available to infants in the United States that contain a nutritionally relevant amount of DHA + ARA. Infant formula is the logical choice as a vehicle for providing DHA and ARA to younger infants who are not breast-fed. During weaning to solid foods, the North American infant receives very little DHA from the diversified mixture of ordinary foods customarily included in the weaning diet. This assumption was validated in Australia and in Finland. Only 3 foods common in the U.S. diet contain significant amounts of DHA, i.e., egg yolks, chicken, and oily fish. Both regular egg yolks and those from chickens fed special diets to increase the (n-3) fatty acid content contain measurable quantities of DHA. A large egg yolk contains between 25 and 140 mg DHA, depending on the diet of the hen. Egg yolks have long been recognized as a safe food for babies and were used in various ancient cultures as a first solid food. Egg yolk was recommended >40 y ago to be started between 4 and 6 mo of age unless there was allergy in the family. More recent suggested guidelines for infants during the first 6 mo of life include the introduction of egg yolk at 5–6 mo. Most recently, Gibson et al. and Makrides et al. reported the effects of feeding normal and DHA-enriched egg yolks to formula-fed and breast-fed infants in the second 6 mo of life. Consuming 4 DHA-enriched egg yolks weekly significantly increased RBC DHA levels at 12 mo in breast-fed infants. Blood cholesterol levels were no higher than those of breast-fed infants. Gibson et al. also found that infants fed egg yolk had improved iron status, as measured by higher serum iron levels and higher transferrin saturation. Based on estimates of food intake (Table 1) and the content of yolk in baby foods (12%), the consumption of yolk (67 g/wk), and thus, cholesterol and iron, in the current study was nearly equivalent to that in the Gibson study. Egg yolk is also a rich source of choline-rich lecithin; choline is a vital constituent of membrane phospholipids and was shown to be an essential nutrient for brain development. Chicken meat contains only a small amount of DHA. Pureed chicken with broth intended for infant use contains 7 mg of DHA (and 43 mg of ARA) in the 55-g Recommended Amount Customarily Consumed Per Eating Occasion. Although chicken is a important source of DHA for adults, the low concentration of DHA in chicken makes this a poor source of DHA for infants. In the United States, no commercial foods intended for infants contain fish. Fish is perceived to be highly allergenic by U.S. pediatricians and parents, even though oily fish such as salmon and tuna have reduced allergenicity if canned commercially. A more difficult issue nutritionally is that oily fish contain substantial amounts of DHA and eicosapentaenoic acid (EPA) but very little ARA. Pureed baby foods available in Europe made with trout and nasello (hake; whiting) supply, per 100 g, 100 to 200 mg of DHA and 25 to 70 mg of EPA but only 3 to 6 mg of ARA. Human milk contains some EPA if the maternal diet contains an EPA source (e.g., fish); thus, normal infant growth and development can occur in the presence of small amounts of EPA. However, providing supplemental EPA to infants without sufficient ARA is problematic. EPA inhibits the elongation of LA to ARA. A DHA-enriched (0.31% DHA) infant formula made with a low-EPA fish oil and containing relatively little EPA (0.08% of total fatty acids) but even less ARA (0.03%) significantly depressed RBC phospholipid ARA levels at 4 mo of age. Thus, the level of EPA in the infant diet should be limited. Infant formulas containing fish oils with a substantial EPA content were shown not to support and to support normal growth in preterm infants. This trial demonstrates that the visual maturation of healthy infants is improved by continued supplies of DHA from both human milk and DHA-enriched baby foods well into 1 y of life. Modifications later in childhood to visual function and other neural processes by this DHA supplementation in baby foods are currently under investigation. Lipids. 2003 Oct. This pilot study evaluated the effects of supplementation with PUFA on blood FA composition and behavior in children with Attention-Deficit/Hyperactivity Disorder (AD/HD)-like symptoms also reporting thirst and skin problems. Fifty children were randomized to treatment groups receiving either a PUFA supplement providing a daily dose of 480 mg DHA, 80 mg EPA, 40 mg arachidonic acid (AA), 96 mg GLA, and 24 mg alpha-tocopheryl acetate, or an olive oil placebo for 4 mon of double-blind parallel treatment. Supplementation with the PUFA led to a substantial increase in the proportions of EPA, DHA, and alpha-tocopherol in the plasma phospholipids and red blood cell (RBC) total lipids, but an increase was noted in the plasma phospholipid proportions of 18:3n-3 with olive oil as well. Significant improvements in multiple outcomes (as rated by parents) were noted in both groups, but a clear benefit from PUFA supplementation for all behaviors characteristic of AD/HD was not observed. For most outcomes, improvement of the PUFA group was consistently nominally better than that of the olive oil group; but the treatment difference was significant, by secondary intent-to-treat analysis, on only 2 out of 16 outcome measures: conduct problems rated by parents (-42.7 vs. -9.9%, n = 47, P = 0.05), and attention symptoms rated by teachers (-14.8 vs. +3.4%, n = 47, P = 0.03). PUFA supplementation led to a greater number of participants showing improvement in oppositional defiant behavior from a clinical to a nonclinical range compared with olive oil supplementation (8 out of 12 vs. 3 out of 11, n = 33, P = 0.02). Also, significant correlations were observed when comparing the magnitude of change between increasing proportions of EPA in the RBC and decreasing disruptive behavior as assessed by the Abbreviated Symptom Questionnaire (ASQ) for parents (r = -0.38, n = 31, P < 0.05), and for EPA and DHA in the RBC and the teachers' Disruptive Behavior Disorders (DBD) Rating Scale for Attention (r = -0.49, n = 24, P < 0.05). Interestingly, significant correlations were observed between the magnitude of increase in alpha-tocopherol concentrations in the RBC and a decrease in scores for all four subscales of the teachers' DBD (Hyperactivity, r = -0.45; Attention, r= -0.60; Conduct, r = -0.41; Oppositional/Defiant Disorder, r = -0.54; n = 24, P < 0.05) as well as the ASQ for teachers (r = -0.51, n = 24, P < 0.05). Thus, the results of this pilot study suggest the need for further research with both n-3 FA and vitamin E in children with behavioral disorders. Encephale. 2003 Jan-Feb. Polyunsatured fatty acids are made out of a hydrocarbonated chain of variable length with several double bonds. The position of the first double bond (omega) differentiates polyunsatured omega 3 fatty acids (for example: alpha-linolenic acid or alpha-LNA) and polyunsatured omega 6 fatty acids (for example: linoleic acid or LA). These two classes of fatty acids are said to be essential because they cannot be synthetised by the organism and have to be taken from alimentation. The omega 3 are present in linseed oil, nuts, soya beans, wheat and cold water fish whereas omega 6 are present in maize, sunflower and sesame oil. Fatty acids are part of phospholipids and, consequently, of all biological membranes. The membrane fluidity, of crucial importance for its functioning, depends on its lipidic components. Phospholipids composed of chains of polyunsatured fatty acids increase the membrane fluidity because, by bending some chains, double bonds prevent them from compacting themselves perfectly. Membrane fluidity is also determined by the phospholipids/free cholesterol ratio, as cholesterol increases membrane viscosity. A diet based on a high proportion of essential polyunsatured fatty acids (fluid) would allow a higher incorporation of cholesterol (rigid) in the membranes to balance their fluidity, which would contribute to lower blood cholesterol levels. Brain membranes have a very high content in essential polyunsatured fatty acids for which they depend on alimentation. Any dietary lack of essential polyunsatured fatty acids has consequences on cerebral development, modifying the activity of enzymes of the cerebral membranes and decreasing efficiency in learning tasks. Epidemiological data: The prevalence of depression seems to increase continuously since the beginning of the century. Though different factors most probably contribute to this evolution, it has been suggested that it could be related to an evolution of alimentary patterns in the Western world, in which polyunsatured omega 3 fatty acids contained in fish, game and vegetables have been largely replaced by polyunsatured omega 6 fatty acids of cereal oils. Some epidemiological data support the hypothesis of a relation between lower depression and/or suicide rates and a higher consumption of fish. These data do not however prove a relation of causality. Cholesterol and depression: Several cohort studies (on nondepressed subjects) have assessed the relationship between plasma cholesterol and depressive symptoms with contradictory results. Though some results found a significant relationship between a decrease of total cholesterol and high scores of depression, some other did not. Studies among patients suffering from major depression signalled more constantly an association between low cholesterol and major depression. Besides, some trials showed that clinical recovery may be associated with a significant increase of total cholesterol. Cholesterol and suicidal behavior: The hypothesis that a low cholesterol level may represent a suicidal risk factor was discovered accidentally following a series of epidemiological studies which revealed an increase of the suicidal risk among subjects with a low cholesterol level. Though some contradictory studies do exist, this relationship has been confirmed by several subsequent cohort studies. These findings have challenged the vast public health programs aimed at promoting the decrease of cholesterol, and even suggested to suspend the administration of lipid lowering drugs. Recent clinical studies on populations treated with lipid lowering drugs showed nevertheless a lack of significant increase of mortality, either by suicide or accident. In addition, several controlled studies among psychiatric patients revealed a decrease of the concentrations of plasma cholesterol among patients who had attempted suicide in comparison with other patients. Polyunsaturated fatty acids and depression: In major depression, all studies revealed a significant decrease of the polyunsaturated omega 3 fatty acids and/or an increase of the omega 6/omega 3 ratio in plasma and/or in the membranes of the red cells. In addition, two studies found a higher severity of depression when the level of polyunsaturated omega 3 fatty acids or the ratio omega 3/omega 6 was low. Parallel to these modifications, other biochemical perturbations have been reported in major depression, particularly an activation of the inflammatory response system, resulting in an increase of the pro-inflammatory cytokines (interleukins: IL-1b, IL-6 and interferon g) and eicosanoids (among others, prostaglandin E2) in the blood and the CSF of depressed patients. These substances cause a peroxidation and, consequently a catabolism of membrane phospholipids, among others those containing polyunsaturated fatty acids. The cytokines and eicosanoids derive from polyunsaturated fatty acids and have opposite physiological functions according to their omega 3 or omega 6 precursor. Arachidonic acid (omega 6) is, among others, precursor of pro-inflammatory prostaglandin E2 (PGE2), whereas polyunsaturated omega 3 fatty acids inhibit the formation of PGE2. It has been shown that a dietary increase of polyunsaturated omega 3 fatty acids reduced strongly the production of IL-1 beta, IL-2, IL-6 and TNF-alpha (tumor necrosis factor-alpha). In contrast, diets with a higher supply of linoleic acid (omega 6) increased significantly the production of pro-inflammatory cytokines, like TNF-alpha. Therefore, polyunsaturated omega 3 fatty acids could be associated at different levels in the pathophysiology of major depression, on the one hand through their role in the membrane fluidity which influences diverse steps of neurotransmission and, on the other hand, through their function as precursor of pro-inflammatory cytokines and eicosanoids disturbing neurotransmission. In addition, antidepressants could exhibit an immunoregulating effect by reducing the release of pro-inflammatory cytokines, by increasing the release of endogenous antagonists of pro-inflammatory cytokines like IL-10 and, finally, by acting like inhibitors of cyclo-oxygenase. Therapeutic use of fatty acids: Data available concerning the administration of supplements of DHA (docosahexanoic acid) or other polyunsaturated fatty acids omega 3 are limited. In a double blind placebo-controlled study on 30 patients with bipolar disorder, the addition of polyunsaturated omega 3 fatty acids was associated with a longer period of remission. Moreover, nearly all the other prognosis measures were better in the omega 3 group. Very recently, a controlled trial showed the benefits of adding an omega 3 fatty acid, eicosopentanoic acid, among depressed patients. After 4 weeks, six of the 10 patients receiving the fatty acid were considered as responders in comparison with only one of the ten patients receiving placebo. Conclusions: Some epidemiological, experimental and clinical data favour the hypothesis that polyunsaturated fatty acids could play a role in the pathogenesis and/or the treatment of depression. More studies however are needed in order to better precise the actual implication of those biochemical factors among the various aspects of depressive illness. Am J Clin Nutr. 2002 Jun. Background: Egg yolks can be a source of docosahexaenoic acid (DHA) and iron but are often associated with adverse consequences on plasma cholesterol. Objective: Our goal was to investigate the effect of consumption of 4 egg yolks/wk on infant DHA status and hemoglobin, ferritin, and plasma cholesterol concentrations. Secondary outcomes included plasma iron, transferrin, and transferrin saturation. Design: This was a randomized controlled trial comparing no dietary intervention, consumption of 4 regular egg yolks/wk, and consumption of 4 n-3 fatty acid-enriched egg yolks/wk in breast-fed and formula-fed infants from 6 to 12 mo of age. Erythrocyte DHA concentrations, cholesterol, and iron status were assessed at 6 and 12 mo of age. Results: Of the 82 breast-fed infants recruited, 23 of 28 (no intervention), 23 of 27 (regular eggs), and 24 of 27 (n-3 eggs) completed the trial. Of the 79 formula-fed infants enrolled, 23 of 27 (no intervention), 24 of 26 (regular eggs), and 20 of 26 (n-3 eggs) completed the trial. Erythrocyte DHA concentrations were 30-40% higher after the n-3 egg intervention than after treatment with regular eggs or no eggs in both breast-fed and formula-fed infants. Egg treatment had no significant effect on plasma cholesterol, hemoglobin, ferritin, and transferrin but did result in improvements in plasma iron and transferrin saturation compared with no egg treatment. Conclusions: n-3 Fatty acid-enriched eggs may provide a means of increasing dietary DHA during the second 6 mo of life. Egg yolks may also be a useful source of iron during the weaning period and can be safely included in the weaning diet with no perturbations in plasma cholesterol. Excerpts from the full text article: Weaning of both breast-fed and formula-fed infants is usually recommended from 4 to 6 mo of age. Although it is common to introduce iron-fortified cereals as a first food, this contrasts with more traditional weaning practices in which egg yolks and brains were used as first foods. Iron stores in breast-fed infants become depleted by 6 mo. Because breast milk is not a good source of iron, iron-rich weaning foods are considered important to avoid iron deficiency. Like meat, egg yolks contain both heme and nonheme iron. Heme iron refers to the iron in hemoglobin, myoglobin, and heme-containing enzymes; nonheme iron includes all other forms of iron. Heme iron is absorbed more efficiently than is nonheme iron and its absorption is not significantly influenced by iron status or other constituents of the diet. Conversely, the absorption of nonheme iron, mainly from iron salts, can be modified by different dietary factors, and studies investigating the addition of whole eggs to meals containing nonheme iron show a reduction in iron absorption. These observations are supported by other studies showing that egg white inhibits nonheme iron absorption. However, iron absorption from egg yolk alone has not been investigated. Given that egg yolks contain some heme iron, are small in volume, and have a soft texture suitable for weaning infants, an investigation of whether egg yolk in the weaning diet can influence iron status is warranted. The n-3 long-chain polyunsaturated fatty acid (LCPUFA) docosahexaenoic acid (DHA; 22:6n-3) is an integral component of breast milk and until recently was not added to infant formulas. Randomized controlled trials of infant formula supplemented with DHA compared with formula containing only precursor fatty acids have consistently shown short-term improvements in visual and neural development of preterm infants. Trials involving term infants have reported neutral or positive outcomes. Although the potential long-term benefits of DHA are still being explored, biochemical data indicate that breast-fed infants accumulate DHA in the brain until 12 mo of age and at a greater rate than do infants fed formula without DHA. To our knowledge, no trials have addressed whether weaning foods high in n-3 fatty acids, such as n-3 fatty acid–enriched egg yolks, can improve infant DHA status during late infancy. The most commonly cited reason for delaying the introduction of whole eggs to infants is to avoid sensitizing infants to egg white proteins and hence the development of egg-related allergies. Furthermore, although dietary recommendations for adults to reduce their intake of eggs to limit the cholesterol content of the diet have little relevance to young children, such guidelines may influence a parent's food choices for his or her child. The purpose of our trial was to investigate the nutritional role of egg yolks in the weaning diets of breast-fed and formula-fed infants. Primary outcome measures included erythrocyte DHA concentrations, infant iron status, and plasma cholesterol concentrations. Secondary outcomes included growth and plasma indexes of allergy. Protocol Healthy 6-mo-old infants born at term (>37 wk gestation) with birth weights >2500 g were eligible for the trial. Infants were excluded if they had known protein intolerances or allergies. Infants were eligible for entry into the breast-fed cohort if they received <120 mL formula (or cow milk)/wk and were eligible for entry into the formula-fed cohort if they were receiving all their nutrition as formula feeds by 4 wk after birth and were subsequently formula fed. All formulas consumed by infants did not contain LCPUFAs and were fortified with iron. [...] Dietary intervention The aim of the dietary intervention was to include 4 egg yolks/wk in the diet of weaning infants between 6 and 12 mo of age without significantly altering intake from other foods. The dietary intervention with n-3 eggs was designed to match the amount of DHA a breast-fed infant would normally receive, ie, 100 mg/d. The fatty acid and nutrient compositions of the eggs are shown in Tables 1and 2. The eggs were enriched with DHA by feeding hens diets rich in n-3 fatty acids, including DHA (11). Eggs (weighing 60–65 g each) were purchased from South Coast Eggs (Myponga, South Australia) and were supplied in plain cartons coded A or B. The regular and n-3 eggs were similar in appearance, taste, and smell. Both the study participants and the research personnel were unaware of the type of eggs provided. Compliance was encouraged by providing the family with 2 dozen eggs per fortnight, delivered by courier to the home of each infant allocated to an egg group. All mothers received advice regarding preparation, received instruction about separating the egg yolk from the egg white, and were supplied with plastic egg separators and a recipe booklet compiled by a dietitian. [...] The normal reference ranges used in this study for the iron status of infants (6 mo to 2 y) are as follows: hemoglobin, 105–135 g/L; ferritin, 10–250 µg/L; iron, 8–30 µmol/L; transferrin, 2.0–3.6 g/L; and transferrin saturation, 10–60%. Infants whose hemoglobin and plasma iron markers fell within these limits were considered iron sufficient. All infants with a hemoglobin concentration <105 g/L were classified as having anemia. Infants who had plasma ferritin concentrations 10 mg/L were classified as having iron deficiency. Infants with iron deficiency who also had hemoglobin concentrations < 105 g/L were classified as having iron deficiency anemia. Study sample A total of 251 eligible mother-infant pairs were approached to enter the trial, and 161 gave written, informed consent to participate (Figure 1). Of these, 82 were breast-fed and 79 were formula-fed. Of the 82 breast-fed infants recruited, 23 of 28 (control group), 23 of 27 (regular egg group), and 24 of 27 (n-3 egg group) completed the trial. Twelve breast-fed infants were withdrawn for the following reasons: anemia at trial entry (n = 8), failure to attend the initial appointment (n = 2), ceased breast-feeding before 9 mo (n = 1), and perceived adverse reaction to eggs by the parents (n = 1). Of the 79 formula-fed infants recruited, 23 of 27 (control group), 24 of 26 (regular egg group), and 20 of 26 (n-3 egg group) completed the trial. Twelve formula-fed infants were withdrawn for the following reasons: failure to attend the initial appointment (n = 9), loss to follow-up (n = 1), perceived adverse reaction to eggs by the parents (n = 1), and unknown reason (n = 1). The dietary intervention resulted in mothers reporting increased use of egg yolks in both the breast-fed and formula-fed cohorts (Table 4). Reported consumption of 4 egg yolks/wk did not affect the frequency of consumption of other foods, such as meats and cereals, that are likely to contribute iron or LCPUFAs to infant diets. The formula-fed infants consumed more adult cereal, meat, chicken, and fish than did the breast-fed infants, whereas the breast-fed infants consumed more baby cereal. Compliance with the dietary intervention is highlighted by the fact that erythrocyte DHA concentrations were 30% higher after treatment with n-3 eggs than after treatment with regular eggs in both the breast-fed and formula-fed infants at 12 mo of age (Table 5). Note that adding 4 n-3 egg yolks/wk to the diets of infants fed formula without LCPUFAs resulted in erythrocyte DHA concentrations that were not significantly different from those in the breast-fed infants in the control group at 12 mo of age. Erythrocyte AA concentrations were <10% lower after treatment with n-3 eggs than after the control or treatment with regular eggs in both the breast-fed and formula-fed cohorts at 12 mo of age. Increasing egg yolk consumption from 1 to 4/wk did not alter mean plasma cholesterol concentrations in either the breast-fed or formula-fed infants (Table 5). The breast-fed infants did, however, have higher cholesterol concentrations than the formula-fed infants at 6 mo of age [4.3 ± 0.8 (n = 70) compared with 3.8 ± 0.7 (n = 67) mmol/L; P < 0.001]. This difference disappeared by 12 mo of age [4.3 ± 0.7 (n = 70) compared with 4.1 ± 0.7 (n = 67) mmol/L], when all infants were consuming a greater variety of foods. Hemoglobin, ferritin, and transferrin values did not differ significantly between the combined egg treatment groups and the control group within both the breast-fed and formula-fed cohorts (Table 6). At 12 mo, however, plasma iron and transferrin saturation were higher in the egg-treated breast-fed and formula-fed infants than in the respective control groups. Only one breast-fed infant receiving regular eggs had iron deficiency anemia at 12 mo of age. Iron status did not differ significantly between the breast-fed and formula-fed infants, except that plasma ferritin concentrations tended to be lower in the breast-fed infants (P = 0.06). At 12 mo of age, more breast-fed infants (17.1%, 12 of 70) than formula-fed infants (7.5%, 5 of 67) were iron deficient (NS). The frequency of positive values for antibodies specific to egg yolk and egg white did not differ significantly between the combined egg treatment group and the control group within both cohorts or between the breast-fed and formula-fed cohorts (Table 7). Weight, length, and head circumferences were not significantly different between the dietary groups nor were there any significant interactions between diet and time (data not shown). However, the breast-fed infants were consistently lighter [at 6 mo, 7575 ± 892 (breast-fed, n = 70) compared with 7942 ± 943 (formula-fed, n = 67) g; at 9 mo, 8584 ± 999 compared with 9081 ± 1043 g; and at 12 mo, 9626 ± 1060 compared with 10121 ± 1130 g; P < 0.01] and shorter [at 6 mo, 67.1 ± 2.6 (breast-fed, n = 70) compared with 67.6 ± 2.3 (formula-fed, n = 67) cm; at 9 mo, 70.9 ± 2.4 compared with 72.1 ± 2.5 cm; and at 12 mo, 75.1 ± 2.5 compared with 75.9 ± 3.5 cm; P < 0.05] than the formula-fed infants. There was no significant difference in head circumference between the breast-fed and formula-fed infants. Discussion Our trial was a systematic study of the nutritional value of including egg yolk in the weaning diet of breast-fed and formula-fed infants. With the separate random assignment of the breast-fed and formula-fed infants, the trial was structured to answer current nutritional issues pertinent to all infants. For example, although formula-fed infants are generally iron sufficient, the issue of improving their LCPUFA status remains topical. Conversely, breast-fed infants have an ample supply of dietary LCPUFAs but have been reported to be at risk of depleted iron stores during late infancy. Our trial addressed whether egg yolk, which can be a good source of both LCPUFAs and iron, is beneficial for all infants regardless of whether they are breast-fed or fed formula, without adverse consequences on plasma cholesterol concentrations or plasma indexes of allergy. A major finding of the current trial was that infants fed formulas without DHA but who consumed 4 n-3 egg yolks/wk had erythrocyte DHA percentages that were not significantly different from those of the breast-fed infants. We showed in an earlier study that weaning diets are very low in LCPUFAs, such that formula-fed infants have little chance in the normal course of events to achieve LCPUFA status similar to that seen in breast-fed infants. In that earlier study, we calculated that it could take 14 regular egg yolks/wk to provide an infant with the same amount of DHA that breast-fed infants receive. The results of the present trial, which showed that 4 regular egg yolks/wk in the weaning diet had no significant effect on erythrocyte DHA status, add credibility to this calculation. However, the addition of 4 n-3 egg yolks/wk, which provided 100 mg additional DHA/d in the weaning diets of the formula-fed infants, resulted in erythrocyte DHA amounts not significantly different from those of the breast-fed infants, who also obtained 100 mg DHA/d. It is also interesting to note that erythrocyte DHA concentrations decreased with age in the breast-fed infants in the control group, whereas there was no significant change in DHA status with time in the formula-fed infants in the control group. Although breast milk remains the main dietary source of DHA in 6–12-mo-old infants, the increasing consumption of weaning foods low in DHA and the reduction in breast milk volume consumed during this age bracket probably explain the reduction in DHA status of the breast-fed infants in the control group. On the other hand, DHA status remains low in infants consuming formulas without DHA and DHA-poor weaning foods and does not alter with age. Our data indicate that n-3 egg yolks can be a useful dietary source of DHA for both breast-fed and formula-fed infants, and show for the first time that egg yolks can be incorporated into the weaning diet for a sustained period of time. The implications of these findings remain to be determined because no trials have specifically addressed changes in physiologic functions or clinical outcomes of infants supplemented with DHA during the second 6 mo of life. Our trial showed no significant effect of egg yolk intervention on hemoglobin and ferritin, the major determinants of iron status, although there was a positive effect of egg treatment on transferrin saturation and plasma iron. This is perhaps not surprising because the infants studied were largely iron sufficient and had adequate hemoglobin and iron stores (as measured by ferritin). The improvements in plasma iron and transferrin saturation with egg treatment reflect increases in iron transportation. Given that the additional amount of iron supplied by the egg yolk intervention was 0.5 mg/d, it is possible that either a greater dose or a longer treatment period would be necessary before improvements in iron stores could be observed. Other comparable trials involving largely iron-sufficient infants that investigated the effect of different iron-containing weaning foods on infant iron status had different results. Engelmann et al showed that partially breast-fed infants randomly allocated to a high-meat (27 g/d) diet had improved iron status compared with infants allocated to a low-meat (10 g/d) weaning diet. Note that the iron contribution from meat in the high-meat group is similar to the amount of iron received from the egg yolk treatment in the current trial. Conversely, one of our earlier trials showed that infants fed a high-iron weaning diet (8.2 ± 2.9 mg/d), in which the iron was derived largely from iron-fortified cereal, had iron status that was not different from that of a control group (5.2 ± 3.4 mg/d) at 12 mo of age. The infants in the intervention group of this earlier trial received an extra 3 mg Fe/d compared with an extra 0.5 mg Fe from egg yolk in the current trial. Collectively, these data suggest that foods such as egg yolks and meat may make a useful contribution to the weaning diet and that interventions aimed at increasing the intake of such foods in iron-deplete infants deserve to be tested in a large, community-based trial. Although an egg yolk typically contains 200 mg cholesterol and 6 g fat (2 g of which is saturated fat), introducing 4 eggs/wk to the diets of weaning infants did not significantly alter plasma cholesterol concentrations in either breast-fed or formula-fed infants. This finding parallels observations from a trial involving adults in which the intake of 4 regular or 4 n-3 eggs/wk resulted in no increase in blood lipids. One of the largest randomized intervention trials that investigated early diet and lifestyle counseling to reduce atherosclerosis risk factors showed that intervention to reduce fat intake and replace saturated fat with monounsaturated and polyunsaturated fat in the weaning diet of infants resulted in no effect on blood lipids when infants were 13 mo of age. On the other hand, breast-fed infants are consistently reported to have higher plasma cholesterol concentrations than formula-fed infants. Various randomized trials of cholesterol-supplemented formula have been undertaken with the aim of investigating the effect of cholesterol supplementation on blood lipids and lipid metabolism. Although both trials reported increases in plasma cholesterol with cholesterol supplementation of infant formula, cholesterol synthesis remained lower in breast-fed infants than in cholesterol-supplemented, formula-fed infants. Few data exists regarding the implications of these findings for physiologic functions and clinical outcomes, and these remain controversial. It has been suggested that cholesterol consumption in human milk may promote the delivery of adequate substrate for brain lipids, but other work suggests that the rat brain synthesizes its cholesterol de novo. Egg allergy and intolerance are among the most common food allergies and intolerances. In our group of healthy infants with no known protein allergies or intolerances, only 1 of 82 breast-fed and 1 of 79 formula-fed infants were withdrawn from the trial because the parents perceived that the infants had an adverse reaction to the egg yolk intervention. Both of these infants were ingesting egg protein as part of their usual weaning diet, however, and we did not have the opportunity to confirm egg intolerance by using a double-blind, placebo-controlled challenge. The presence of antigen-specific antibodies to egg yolk and egg white were used as indexes of egg intolerance, although IgE antibody responses to food proteins can appear in healthy infants with no clinical manifestations of allergy or intolerance. The results of our trial confirm this because none of the infants had overt signs of egg intolerance, but 4% and 8% of the egg-treated infants had specific antibody responses to egg yolk and egg white, respectively, compared with 2% and 15% of the control infants, respectively. Within the limits of our trial there were no significant differences in specific antibody responses between the treated and control infants. However, given the relatively low rates of positive responses, hundreds of infants per treatment group would have been required to detect differences. Further studies should be conducted to better define the clinical and plasma markers of allergy in response to egg intervention. In summary, the results of our trial indicate that it is possible and practical for weaning infants to consume 4 egg yolks/wk without affects on the intake of other foods such as cereals and meats. The egg yolk intervention resulted in modest improvements in iron status that may be most beneficial to infants who are iron deplete, and this deserves further investigation. Eggs fortified with n-3 fatty acids provide a means of increasing dietary DHA during the second 6 mo of life without altering plasma cholesterol. The emerging work in the field of dietary DHA and infant outcomes will determine whether there are any physiologic or clinical benefits to improved DHA status during the second half of infancy. J Mol Neurosci. 2001 Apr-Jun. Patients with the Zellweger syndrome and its variants have very low levels of docosahexaenoic acid (DHA) in the brain, retina, and other tissues. Such a marked DHA deficiency could be related to the pathogenesis of peroxisomal disorders. Therefore, restoring the DHA levels in these patients can probably improve the clinical course of the disease. With this rationale, 20 patients with generalized peroxisomal disorders have been treated to date with DHA ethyl ester, at daily doses of 100-500 mg, for variable periods of time. Treatment has been always accompanied by a nutritious diet, normal for the age, in order to provide all the necessary nutrients and avoid a polyunsaturated fatty acid (PUFA) imbalance. The most constant improvement has been normalization of the DHA levels and liver function. Vision has improved in about half the patients and muscle tone has generally increased. Magnetic resonance imaging (MRI) examination revealed improvement of myelination in 9 patients. Significantly, the clinical improvement has been most marked in those patients who started the treatment before 6 mo of age. Biochemically, the plasma very long-chain fatty acids (VLCFA) 26:0 and 26:1n-9 decreased markedly despite the complete diet provided. In erythrocytes, the plasmalogen ratio 18: ODMA/18:0 increased in most cases, and sometimes even normalized. All these beneficial effects suggest that DHA deficiency plays a fundamental role in the pathogenesis of peroxisomal disease. Because DHA accretion is maximal during early brain development, it is essential to initiate the treatment as soon as possible. Otherwise, restoration of brain DHA levels and prevention of further damage will not be possible. J Child Adolesc Psychopharmacol. 2000 Summer. Zinc is an important co-factor for metabolism relevant to neurotransmitters, fatty acids, prostaglandins, and melatonin, and indirectly affects dopamine metabolism, believed intimately involved in attention-deficit/hyperactivity disorder (ADHD). To explore the relationship of zinc nutrition to essential fatty acid supplement and stimulant effects in treatment of ADHD, we re-analyzed data from an 18-subject double-blind, placebo-controlled crossover treatment comparison of d-amphetamine and Efamol (evening primrose oil, rich in gamma-linolenic acid). Subjects were categorized as zinc-adequate (n = 5), borderline zinc (n = 5), and zinc-deficient (n = 8) by hair, red cell, and urine zinc levels; for each category, placebo-active difference means were calculated on teachers' ratings. Placebo-controlled d-amphetamine response appeared linear with zinc nutrition, but the relationship of Efamol response to zinc appeared U-shaped; Efamol benefit was evident only with borderline zinc. Placebo-controlled effect size (Cohen's d) for both treatments ranged up to 1.5 for borderline zinc and dropped to 0.3-0.7 with mild zinc deficiency. If upheld by prospective research, this post-hoc exploration suggests that zinc nutrition may be important for treatment of ADHD even by pharmacotherapy, and if Efamol benefits ADHD, it likely does so by improving or compensating for borderline zinc nutrition. Dev Med Child Neurol. 2000 Mar. The effects of dietary docosahexaenoic acid (DHA) supply during infancy on later cognitive development of healthy term infants were evaluated in a randomized clinical trial of infant formula milk supplemented with 0.35% DHA or with 0.36% DHA and 0.72% arachidonic acid (AA), or control formula which provided no DHA or AA. Fifty-six 18-month-old children (26 male, 30 female) who were enrolled in the trial within the first 5 days of life and fed the assigned diet to 17 weeks of age were tested using the Bayley Scales of Infant Development, 2nd edition (BSID-II) (Bayley 1993) at the Retina Foundation of the Southwest, Dallas, TX. These children had also been assessed at 4 months and 12 months of age for blood fatty-acid composition, sweep visual evoked potential (VEP) acuity, and forced-choice preferential looking (FPL) acuity (Birch et al. 1998). Supplementation of infant formula with DHA+AA was associated with a mean increase of 7 points on the Mental Development Index (MDI) of the BSID-II. Both the cognitive and motor subscales of the MDI showed a significant developmental age advantage for DHA- and DHA+AA-supplemented groups over the control group. While a similar trend was found for the language subscale, it did not reach statistical significance. Neither the Psychomotor Development Index nor the Behavior Rating Scale of the BSID-II showed significant differences among diet groups, consistent with a specific advantage of DHA supplementation on mental development. Significant correlations between plasma and RBC-DHA at 4 months of age but not at 12 months of age and MDI at 18 months of age suggest that early dietary supply of DHA was a major dietary determinant of improved performance on the MDI. Am J Clin Nutr. 2000 Jan. Generalized peroxisomal disorders are severe congenital diseases that involve the central nervous system, leading to severe psychomotor retardation, retinopathy, liver disease, and early death. In these disorders, peroxisomes are not normally formed and their enzymes are deficient. Characteristically, plasmalogen synthesis and beta-oxidation of very-long-chain fatty acids (VLCFAs) are affected. We found that patients with generalized peroxisomal disorders have a profound brain deficiency of docosahexaenoic acid (DHA; 22:6n-3) and low DHA concentrations in all tissues and the blood. Given the fundamental role of DHA in neuronal and retinal membranes, a DHA deficiency of this magnitude might be pathogenic. Thus, we studied the possible therapeutic effect of normalizing DHA concentrations in patients with peroxisomal disorders. We chose the DHA ethyl ester (DHA-EE) because of its high degree of purity at daily oral doses of 100-500 mg. This article summarizes the results of treatment of 13 patients with DHA-EE, with some follow-up evidence of clinical improvement. Supplementation with DHA-EE normalized blood DHA values within a few weeks. Plasmalogen concentrations increased in erythrocytes in most patients and after DHA concentrations were normalized, amounts of VLCFAs decreased in plasma. Liver enzymes returned almost to normal in most cases. From a clinical viewpoint, most patients showed improvement in vision, liver function, muscle tone, and social contact. In 3 patients, normalization of brain myelin was detected by magnetic resonance imaging. In 3 others, myelination improved. In a seventh patient, myelination is progressing at a normal rate. These results suggest a fundamental role of DHA in the pathogenesis of Zellweger syndrome. DHA therapy is thus strongly recommended, not only to alleviate symptoms in patients with life-threatening diseases, but also to clarify remaining questions regarding the role of DHA in health and disease. Biofactors. 2000. DHA is abundant in the brain. Deficiency of DHA changes behavior in animals. The purpose of the present studies was to clarify the effect of DHA intake on hostility and plasma catecholamines. In study 1, forty-one students took either DHA-rich oil capsules containing 1.5-1.8 g DHA/d (17 females and 5 males) or control oil capsules containing 97% soybean oil plus 3% fish oil (12 females and 7 males) for 3 mon in a double blind fashion. They took a psychological test (P-F Study) at the start and end of the study. Study I started at the end of summer vacation and ended in the middle of mental stress of final exams. In the control group, hostility measured by P-F Study was significantly increased at the end of the study as compared with that measured at the start (+58%), whereas it was not significantly changed in the DHA group (-14%). In a similar double blind two-mon study (study 2), we measured plasma catecholamines and cortisol of students (3 females and 4 males for the DHA group and the same numbers for the control) at the start and end of the study. In study 2 the students were under a continuous stress of final exams that lasted for two mon throughout the whole study period. The plasma cortisol did not change in either group, but the norepinephrine concentration was significantly decreased in the DHA group (-31%), whereas it stayed at the same level in the control group. These effects of DHA intake may be applied to people under psychological stress. J Nutr Sci Vitaminol (Tokyo). 1999 Oct. We previously found that docosahexaenoic acid (DHA) intake prevented aggression from increasing at times of mental stress. In the present study, we investigated whether DHA intake modified the plasma catecholamines and cortisol of medical students during a 9-wk period of final exams. We also investigated the effects of DHA intake on a 75 g oral glucose tolerance test (oGTT). Fourteen medical students participated in the present study. They were randomly allocated to either control or DHA group in a double-blind manner. Subjects in the control group (4 males and 3 females) took 10 control capsules/d, each capsule containing 280 mg of mixed plant oil, and those in the DHA group (4 males and 3 females) took 10 DHA capsules/d containing 1.5 g DHA for 9 wk, during which subjects underwent more than 20 stressful final exams. At the start and end of the study, plasma catecholamines (epinephrine, norepinephrine (NE) and dopamine) and cortisol were measured; a 75 g OGTT was also performed. There were no intra- or intergroup differences in plasma glucose concentrations. However, NE concentrations were significantly reduced after DHA administration (-31%, p < 0.03). The other catecholamines and cortisol did not change significantly. The plasma ratio of epinephrine to NE increased in every DHA subject (+78%, p < 0.02), and intergroup differences were significant (p < 0.03). We conclude that these effects of DHA may be applied to people under long-lasting psychological stress to prevent stress-related diseases. Rev Neurol. 1999 Jan. INTRODUCTION: We found that patients with the Zellweger syndrome and other generalized peroxisomal disorders have a dramatic decrease of docosahexaenoic acid (DHA, 22:6n-3) in the blood, brain, retina and other tissues. DHA is believed to play an important role in the brain and retina. DEVELOPMENT: Patients with the Zellweger syndrome and its variants have severe cerebral and retinal defects that could be related to their DHA deficiency. With this rationale, we have been treating peroxisomal-disorder patients with a DHA derivative of a high degree of purity (DHA ethyl ester, > 90% pure) since 1991. So far, we have treated 13 DHA-deficient peroxisomal patients, one with the classic Zellweger syndrome and 12 with milder variants of the disease. This paper presents the follow-up of these DHA-treated patients. In summary, we have found important improvements in liver function, in the plasmalogen levels and in the two ratios 26:0/22:0 y 26:1/22:0, diagnostic of the disease. We have also found clear clinical improvements in most cases. Most significantly, magnetic resonance imaging has shown advances in brain myelination, so far in 6 of the treated patients. CONCLUSION: We strongly recommend treatment with DHA ethyl ester in all DHA-deficient patients with generalized peroxisomal disorders. Logically, treatment should be started as soon as possible, in the hope of preventing cerebral and visual damage. Lipids. 1999. The purpose of the present research was to clarify the effect of docosahexaenoic acid (DHA) intake on behavior and plasma catecholamines (CA). In Study 1, 42 students took either DHA-rich oil capsules containing 1.5-1.8 g DHA/d or control oil capsules containing 97% soybean oil plus 3% of another fish oil for 3 mon in a double-blind fashion. They took a psychological test (PF Study) at the start and end of the study. This study started at the end of summer vacation and ended just before the final exams. In the control group, external aggression (aggression against others) in PF Study was significantly increased at the end of the study as compared with that measured at the start (+8.9%), whereas it was not significantly changed in the DHA group (-1.0%). In a similar double-blind study (Study 2), we measured external aggression under nonstressful conditions. External aggression slightly decreased in the control group, whereas there were no significant changes in the DHA group. In Study 3 with 14 students, plasma CA were measured at the start and end of capsule administration period of 2 mon. Subjects were under continuous stress of the final exams that lasted throughout the whole study period. The ratio of plasma epinephrine to norepinephrine concentrations was significantly increased in the DHA group (78%), whereas it stayed at the same level in the control group. In Study 4, mice were fed either DHA-deficient diet or -sufficient diet for 4 wk, and their rearing frequency (an anxiety index) was measured. In the DHA-sufficient group, the rearing frequency was significantly less than in the other group. These effects of DHA intake may be applied to people in an attempt to ameliorate stress-related diseases. Lipids. 1998 Oct. Long-chain polyunsaturated fatty acids (LCPUFA) are important for normal visual and cortical development. In a previous study of the effects of LCPUFA on cognitive function of term infants at the age of 3 mon, we indicated that infants with evidence of reduced growth parameters at birth and impaired attention control as manifested by a late peak fixation during infant habituation assessment may benefit from LCPUFA supplementation. The aim of this prospective study was to determine whether LCPUFA supplementation and late peak fixation are related to means-end problem-solving ability in these same infants at the age of 9 mon. Term infants (58) were randomized to one of two formulas containing either LCPUFA or no LCPUFA and completed 4 mon of feeding with their formula. Cognitive function was assessed at 3 mon of age by measures of infant habituation. Infants (20 LCPUFA and 20 no-LCPUFA) completed the problem-solving assessment at 9 mon. The no-LCPUFA group had lower scores on both measures of intention and number of solutions, but neither of these differences was significant. Analysis of covariance for the effects of group and peak fixation, covaried with gestation and birth weight, showed that the number of solutions was significantly reduced in the late peak-fixation infants receiving no LCPUFA (P<0.02). Intention scores tended to be reduced in this group (P<0.06). The late peak-fixation infants who received LCPUFA had solution and intention scores similar to early peak-fixation infants receiving LCPUFA or no LCPUFA. These findings suggest that in term infants who have reduced growth parameters at birth and who show evidence of impaired attention control, information processing and problem-solving ability in infancy may be enhanced by LCPUFA supplementation. Lancet. 1998 Aug 29. Background: Long-chain polyunsaturated fatty acids (LCPUFA) are important for normal visual and brain development. Although present in human milk, LCPUFA have until recently been absent from artificial formulas, and infants may have limited ability to synthesise LCPUFA. To determine the clinical significance of this relative deficiency of LCPUFA, we undertook a randomised trial of the relation between LCPUFA supplementation and infant cognitive behaviour. Methods: 44 term infants had been randomised to a formula supplemented with LCPUFA (21) or not supplemented with LCPUFA (23), which they had taken from birth to age 4 months. Infant cognitive behaviour was assessed at 10 months of age by a means-end problem-solving test--the intentional execution of a sequence of steps to achieve a goal. The problem required three intermediate steps to achieve the final goal, uncovering and retrieving a hidden toy. Findings: Infants who received LCPUFA-supplemented formula had significantly more intentional solutions than infants who received the no-LCPUFA formula (median 2.0 vs 0, p=0.021). Intention scores (median 14.0 vs 11.5 [maximum 18]) were also increased in this group (p=0.035). Interpretation: These findings suggest that term infants may benefit from LCPUFA supplementation, and that the effects persist beyond the period of supplementation. Since higher problem-solving scores in infancy are related to higher childhood IQ scores, supplementation with LCPUFA may be important for the development of childhood intelligence. Neurology. 1998 Jul. OBJECTIVE: We have found that docosahexaenoic acid (DHA; 22:6n-3), an important constituent of the CNS and retina, is dramatically decreased in patients with generalized peroxisomal disorders. Such a DHA deficiency could be pathogenic. Our aim was to test the possible beneficial effects of normalizing the DHA levels in these patients. The current report focuses on MRI changes during the treatment and reports follow-up evidence of myelin improvement in five peroxisomal disorder patients treated with DHA. METHODS: DHA ethyl ester (DHA-EE), about 90% pure, was used at daily oral doses of 100 to 600 mg. The treatment was monitored both clinically and biochemically, with periodic neurophysiologic and MRI examinations and repeated controls of liver tests and blood fatty acids by capillary column gas chromatography. RESULTS: DHA-EE normalized the blood levels of DHA in a few weeks. The levels of very-long-chain fatty acids decreased in plasma and those of plasmalogens increased in erythrocytes. Visual and liver function improved. On MRI, a virtual normalization of the brain myelin images was found in the three youngest patients. A clear improvement was noticed in the other two. CONCLUSIONS: The beneficial effects obtained indicate that DHA deficiency plays an important role in the pathogenesis of peroxisomal disease, probably in relation to myelinogenesis. Early initiation of DHA therapy is thus strongly recommended in patients with generalized peroxisomal disorders. Lipids. 1996 Mar. There is a need to determine whether there is a dietary requirement for docosahexaenoic acid (DHA, 22:6n-3) by term infants to achieve their full developmental potential. Studies of brain fatty acid composition demonstrated that infants who were breast fed had greater levels of cerebral cortex DHA than did infants who were formula fed, suggesting that DHA in the cerebrum is dependent on a supply in the diet. Some physiological studies reported that electrophysiological and behavioral assessments of visual function were improved in breast-fed infants relative to those fed formula and that this was related to the length of breast feeding. While some randomized studies of DHA supplementation of infant formula to term infants demonstrated that the visual function of formula-fed infants could be improved to breast-fed levels by adding DHA to formula, others failed to demonstrate an effect. Variations in dietary treatments and methods of assessment make comparison of the studies difficult. Further work is necessary to rigorously establish if there are long-term benefits of dietary DHA to the term infant. Lipids. 1996 Mar. A patient with classic Zellweger syndrome was treated with docosahexaenoic acid ethyl ester (DHA-EE) for three months. Five other patients with Zellweger variants (four of them less than one-year-old and a five-year-old) were treated with DHA-EE until normalization of the DHA levels in erythrocytes. When arachidonic acid (AA) concentration decreased, AA was added to the diet. Thereafter, a combined treatment with DHA plus AA followed, in a variable proportion that allowed the high levels of DHA in erythrocytes to be maintained. In the patient with Zellweger syndrome, DHA therapy produced an increase in plasmalogen and a decrease in 26:0 and 26:1. No clear clinical improvement could be detected in this patient during the short period of treatment with DHA-EE. The most consistent clinical effect produced by DHA therapy in the other patients with disorders of peroxisomal biogenesis was visual improvement, even in those patients that were virtually blind before the treatment. In general, the developmental curve began to accelerate. The infants became more alert, acquired better visual and social contact and muscular tone improved, with the beginning of good head control. The liver tests tended to normalize and some patients showed a reduction of hepatomegaly. All these favorable changes occurred when the patients were taking the DHA-EE alone. In some of the patients, muscular tone seemed to improve further after introducing AA supplements. From the biochemical point of view, the plasmalogen levels increased in most cases in erythrocytes, and the two ratios 26:0/22:0 and 26:1/22:0 decreased in plasma. In some patients there was a tendency for 26:1 to increase in plasma and for 18:0 plasmalogen to decrease in erythrocytes when AA was introduced in the diet. The significance of these findings remains to be elucidated, but they stress the importance of strict monitoring and control of the polyunsaturated fatty acids status during DHA therapy. J Clin Invest. 1996 Feb 15. 41 students took either docosahexaenoic acid (DHA)-rich oil capsules containing 1.5-1.8 grams DHA/day (17 females and 5 males) or control oil capsules containing 97% soybean oil plus 3% fish oil (12 females and 7 males) for 3 mo in a double-blind fashion. They took a psychological test (P-F Study) and Stroop and dementia-detecting tests at the start and end of the study. The present study started at the end of summer vacation and ended in the middle of mental stress such as final exams. In the control group extraggression (aggression against others) in P-F Study was significantly increased at the end of the study as compared with that measured at the start (delta = +8.9%, P = 0.0022), whereas it was not significantly changed in the DHA group (delta = -1.0%). The 95% CI of differences between the DHA and control groups were -16.8 to -3.0%. DHA supplementation did not affect the Stroop and dementia-detecting tests. Thus, DHA intake prevented extraggression from increasing at times of mental stress. This finding might help understand how fish oils prevent disease like coronary heart disease. Neurology. 1993 Jul. Docosahexaenoic acid (DHA, 22:6 omega 3) is a major constituent of brain membrane phospholipids and photoreceptor cells. Patients with generalized peroxisomal disorders have extremely low levels of DHA in the brain and other tissues. Since a DHA deficiency could explain some basic symptoms in peroxisomal-disorder patients, we tested the possible beneficial effects of DHA in two patients with neonatal adrenoleukodystrophy (NALD). Before the treatment, both patients had very low DHA levels in plasma and erythrocytes. We first gave DHA in the form of fish oil and, in both patients, the rapid increase in red-cell DHA levels indicated that this fatty acid was being absorbed and incorporated into membrane phospholipids very fast. However, a low ratio 22:6 omega 3/22:5 omega 3 was still present in erythrocyte membranes, and the content of 20:5 omega 3 (eicosapentaenoic acid) was too high with the fish oil diet. We then began treatment with pure DHA ethyl ester and, after a few weeks, erythrocyte omega 3 polyunsaturated fatty acids were normal. There was an increase in the 18:0 molecular species of plasmalogens in both patients, most significantly in the child with affected plasmalogen biosynthesis in cultured fibroblasts. In the less severely affected NALD patient, treatment with DHA produced a very significant decrease in the ratios 24:1/22:0 and 26:1/22:0, and this child improved neurologically. The present data suggest that DHA deficiency may be the cause for some of the most characteristic abnormalities in peroxisomal-disorder patients and open new therapeutic possibilities for these patients. J Lipid Res. 1993 Jan. The effect of docosahexaenoic acid (DHA) on mitochondrial and peroxisomal fatty acid oxidation and on key enzymes in triglyceride metabolism was investigated in the liver of rats fed a standard diet, a cholesterol diet, and a pelleted chow diet. Unexpectedly, in all three rat models repeated administration of highly purified DHA (92% pure) at different doses and times, at a dose of 1000 mg/day per kg body weight, resulted in no significant decrease of hepatic and plasma concentration of triglycerides. The serum concentrations of cholesterol and phospholipids showed an increase in a time-dependent manner in rats fed the pelleted chow diet. The hepatic concentration of cholesterol was increased in rats fed the cholesterol diet and pelleted chow diet after administration of DHA compared to palmitic acid. In all rat models, treatment with DHA tended to increase the peroxisomal beta-oxidation. This was accompanied with a significant increase (1.5-fold) of fatty acyl-CoA oxidase activity. The mitochondrial fatty acid oxidation system and carnitine palmitoyl-transferase activity, however, were almost unchanged. Moreover, palmitoyl-CoA synthetase activity was increased, whereas the palmitoyl-CoA hydrolase activity was decreased. Neither microsomal phosphatidate phosphohydrolase activity nor cytosolic phosphatidate phosphohydrolase activity was affected by DHA feeding in the three rat models. Acyl-CoA:1,2-diacylglycerol acyltransferase activity was also unaffected. In contrast to docosahexanoic acid feeding, eicosapentaenoic acid (EPA) administration possessed a hypotriglyceridemic effect and resulted in an increase of mitochondrial and peroxisomal oxidation of fatty acids. Carnitine palmitoyltransferase activity was also stimulated. Phosphatidate phosphohydrolase activity was unaffected whereas diacylglycerol acyltransferase activity was increased by EPA treatment compared with palmitic acid feeding. The results indicate that docosahexaenoic acid, in contrast to eicosapentaenoic acid, does not inhibit the synthesis and secretion of triglycerides in the liver. In addition, the results emphasize the importance that stimulation of peroxisomal beta-oxidation by these n-3 fatty acids is not sufficient to decrease the serum levels of triglycerides. In addition, increased mitochondrial beta-oxidation of fatty acids and thereby decreased availability of nonesterified fatty acids may be a mechanism by which EPA inhibits triglyceride, and subsequently very low density lipoprotein-triglyceride, production. Whether DHA and EPA possess different metabolic properties should be considered. Biol Psychiatry. 1989 Jan 15. In a Latin-square double-crossover with random assignment to sequence, 18 boys, aged 6-12 years, with attention-deficit hyperactivity disorder received 1 month each of placebo, D-amphetamine, and Efamol (evening primrose oil containing gamma-linolenic acid, with vitamin E as preservative). Parents' ratings were noncontributory. Teachers' ratings showed a trend of Efamol effect between placebo and D-amphetamine. The trend reached significance (p less than 0.05) only on Conners Hyperactivity Factor. Dosage may be crucial; 8 Efamol capsules per day were used in this study. Heuristic data scrutiny suggested possible interaction (sequence effect). Further study with a different design and dose is suggested. This study does not establish Efamol as an effective treatment. |