Short-term supplementation of EPA-enriched ethanolamine plasmalogen increases the level of DHA in the brain and liver of n-3 PUFA deficient mice in early life after weaning.

A lack of n-3 polyunsaturated fatty acids (PUFAs) in mothers' diet significantly reduced the amount of docosahexaenoic acid (DHA) in the brains of offspring, which might affect their brain function. Our previous research has proven multiple benefits of eicosapentaenoic acid (EPA)-enriched ethanolamine plasmalogen (pPE) in enhancing the learning and memory ability. However, the effect of dietary supplementation with EPA-pPE on the DHA content in the brain and liver of offspring lacking n-3 PUFAs in early life is still unclear. Female ICR mice were fed with n-3 PUFA-deficient diets throughout the gestation and lactation periods to get n-3 PUFA-deficient offspring. The lipid profiles in the cerebral cortex and liver of offspring were analyzed using lipidomics after dietary supplementation with EPA-pPE (0.05%, w/w) and EPA-phosphatidylcholine (PC) (0.05%, w/w) for 2 weeks after weaning. Dietary supplementation with EPA could significantly change fatty acid composition in a variety of phospholipid molecular species compared with the n-3 deficient group. EPA-pPE and EPA-PC remarkably increased the DHA content in the brain PC, ether-linked phosphatidylcholine (ePC), and phosphatidylethanolamine plasmalogen (pPE) and liver triglyceride (TG), lyso-phosphatidylcholine (LPC), ePC, phosphatidylethanolamine (PE), and pPE molecular species, in which EPA-pPE showed more significant effects on the increase of DHA in cerebral cortex PC, ePC and liver PC compared with EPA-PC. Both EPA-phospholipids could effectively increase the DHA levels, and the pPE form was superior to PC in the contribution of DHA content in the cerebral cortex PC, ePC and liver PC molecular species. EPA-enriched ethanolamine plasmalogen might be a good nutritional supplement to increase DHA levels in the brains of n-3 PUFA-deficient offspring.

Absorption Kinetics of Ethanolamine Plasmalogen and Its Hydrolysate in Mice.

Ethanolamine plasmalogen (PlsEtn), a subclass of ethanolamine glycerophospholipid (EtnGpl), has been reported to have many biological and dietary functions. In terms of PlsEtn absorption, some studies have reported that PlsEtn is re-esterized at the sn-2 position using lymph cannulation and the everted jejunal sac model. In this study, we aimed to better understand the uptake kinetics of PlsEtn and increase its absorption. We thus compared the uptake kinetics of PlsEtn with that of the lyso-form, in which the fatty acid at the sn-2 position was hydrolyzed enzymatically. Upon administration of EtnGpl (extracted from oysters or ascidians, 75.4 mol% and 88.4 mol% of PlsEtn ratio, respectively), the plasma PlsEtn species in mice showed the highest levels at 4 or 8 hours after administration. In the contrast, administration of the EtnGpl hydrolysate, which contained lysoEtnGpl and free fatty acids, markedly increased the plasma levels of PlsEtn species at 2 h after administration. The area under the plasma concentration-time curve (AUC), especially the AUC0-4 h of PlsEtn species, was higher with hydrolysate administration than that with EtnGpl administration. These results indicate that EtnGpl hydrolysis accelerated the absorption and metabolism of PlsEtn. Consequently, using a different experimental approach from that used in previous studies, we reconfirmed that PlsEtn species were absorbed via hydrolysis at the sn-2 position, suggesting that hydrolysis in advance could increase PlsEtn uptake.

Dietary PlsEtn Ameliorates Colon Mucosa Inflammatory Stress and ACF in DMH-Induced Colon Carcinogenesis Mice: Protective Role of Vinyl Ether Linkage.

Ethanolamine plasmalogen (PlsEtn), a sub-class of ethanolamine glycerophospholipids (EtnGpl), is a universal phospholipid in mammalian membranes. Several researchers are interested in the relationship between colon carcinogenesis and colon PlsEtn levels. Here, we evaluated the functional role of dietary purified EtnGpl from the ascidian muscle (87.3 mol% PlsEtn in EtnGpl) and porcine liver (7.2 mol% PlsEtn in EtnGpl) in 1,2-dimethylhydrazine (DMH)-induced aberrant crypt foci (ACF) in vivo, and elucidated the possible underlying mechanisms behind it. Dietary EtnGpl-suppressed DMH-induced aberrant crypt with one foci (AC1) and total ACF formation (P < 0.05). ACF suppression by dietary ascidian muscle EtnGpl was higher compared with dietary porcine liver EtnGpl. Additionally, dietary EtnGpl decreased DMH-induced oxidative damage, overproduction of TNF-α, and expression of apoptosis-related proteins in the colon mucosa. The effect of dietary ascidian muscle EtnGpl showed superiority compared with dietary porcine liver EtnGpl. Our results demonstrate the mechanisms by which dietary PlsEtn suppress ACF formation and apoptosis. Dietary PlsEtn attained this suppression by reducing colon inflammation and oxidative stress hence a reduction in DMH-induced intestinal impairment. These findings provide new insights about the functional role of dietary PlsEtn during colon carcinogenesis.

The Impact of Ascidian (Halocynthia roretzi)-derived Plasmalogen on Cognitive Function in Healthy Humans: A Randomized, Double-blind, Placebo-controlled Trial.

OBJECTIVES: Plasmalogen, phospholipids with previously shown associations with dementia, has attracted attention as a substance found in some studies to improve cognitive function. The effects of ascidian-derived plasmalogens on cognitive performance improvement were assessed in a randomized, double-blind, placebo-controlled study including Japanese adult volunteers with mild forgetfulness. METHODS: Participants consumed either the active food containing ascidian-derived plasmalogen (1 mg as plasmalogen) or the placebo food for 12 weeks, and their cognitive performance was assessed by Cognitrax. Participants were randomly allocated into the intervention (ascidian-derived plasmalogen; 8 males, and 17 females; 45.6 ± 11.1 years) or the placebo (9 males, and 15 females; mean age, 46.4 ± 10.8 years) group. RESULTS: Compared to the placebo group, the intervention group showed a significant increase score in composite memory (eight weeks: 3.0 ± 16.3 points, 12 weeks: 6.7 ± 17.5 points), which was defined as the sum of verbal and visual memory scores. CONCLUSIONS: These results indicate the consumption of ascidian-derived plasmalogen maintains and enhances memory function. This study was registered at the University Hospital Medical Information Network Clinical Trial Registry (UMIN-CTR, registry no. UMIN000026297). This study did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

Eicosapentaenoic Acid-Enriched Phosphoethanolamine Plasmalogens Alleviated Atherosclerosis by Remodeling Gut Microbiota to Regulate Bile Acid Metabolism in LDLR-/- Mice.

Eicosapentaenoic acid (EPA)-enriched phosphoethanolamine plasmalogens (EPA-PlsEtns) might be retained in the intestine rich in gut microbiota for a long time after treatment. It reminded us that EPA-PlsEtns might affect intestinal microbiota composition and its metabolites, which have been identified as a contributing factor in the development of cardiovascular diseases. In the present study, EPA-PlsEtn administration for 8 weeks significantly reduced the atherosclerotic lesion area in low-density lipoprotein receptor deficient (LDLR-/-) mice. Notably, the serum total cholesterol and low-density lipoprotein cholesterol levels were significantly reduced by 33.6 and 38.2%, respectively, by EPA-PlsEtns instead of EPA in the form of ethyl ester (EPA-EE) treatment compared with the model group. EPA-PlsEtn administration also increased total neutral sterol and bile acids in feces by 92 and 39%, respectively, rather than EPA-EE. Mechanistically, EPA-PlsEtns might affect the abundance of gut microbiota contributing to the alteration of bile acid profiles, which might further accelerate bile acid synthesis via increasing cholesterol 7 α-hydroxylase expression induced by the inhibition of farnesoid X receptor activation.

Biological Functions of Plasmalogens.

Plasmalogens (Pls) are one kind of phospholipids enriched in the brain and other organs. These lipids were thought to be involved in the membrane bilayer formation and anti-oxidant function. However, extensive studies revealed that Pls exhibit various beneficial biological activities including prevention of neuroinflammation, improvement of cognitive function, and inhibition of neuronal cell death. The biological activities of Pls were associated with the changes in cellular signaling and gene expression. Membrane-bound GPCRs were identified as possible receptors of Pls, suggesting that Pls might function as ligands or hormones. Aging, stress, and inflammatory stimuli reduced the Pls contents in cells, and addition of Pls inhibited inflammatory processes, which could suggest that reduction of Pls might be one of the risk factors for the diseases associated with inflammation. Oral ingestion of Pls showed promising health benefits among Alzheimer's disease (AD) patients, suggesting that Pls might have therapeutic potential in other neurodegenerative diseases.

Therapeutic Efficacy of Plasmalogens for Alzheimer's Disease, Mild Cognitive Impairment, and Parkinson's Disease in Conjunction with a New Hypothesis for the Etiology of Alzheimer's Disease.

It has been reported in recent years that blood levels of plasmalogens (Pls) are decreased in various diseases. None of those reports, however, conducted any clinical trials to examine the effect of Pls on those diseases. This article describes our recent report on a therapeutic efficacy of orally administered Pls in mild cognitive impairment (MCI), mild to severe Alzheimer's disease (AD), and Parkinson's disease (PD). A 24-week, multicenter, randomized, double-blind, placebo-controlled trial was performed in patients with MCI (n = 178) and mild AD (n = 98). The study design for moderate AD (n = 57) and severe AD (n = 18) was 12-week open-labeled, and the design for patients with PD (n = 10) was 24-week open-labeled. They showed a significant improvement in cognitive function and other clinical symptoms with elevation of the blood Pls levels. No adverse events were reported. The baseline levels of plasma ethanolamine plasmalogen and erythrocyte ethanolamine plasmalogen in MCI, AD, and PD were significantly lower than those of normal aged. The degree of reduction in the blood Pls levels was in the order of MCI â‰º mild AD ≺ moderate AD ≺ severe AD ≺ PD. The findings suggest that the blood levels of Pls may be a beneficial biomarker for assessing AD severity. Based on these results, we have proposed a new hypothesis for the etiology of AD and other neuropsychiatric disorders.

Preparation and effects on neuronal nutrition of plasmenylethonoamine and plasmanylcholine from the mussel Mytilus edulis.

Plasmenylethonoamine (pPE) and plasmanylcholine (aPC) are important phospholipid subclasses. Herein we explored optimum conditions for enzymatic purification and preparation of pPE and aPC from the mussel Mytilus edulis and bovine brain. Among them, pPE in Mytilus edulis PE was mainly p18:0-20:5 and p18:0-22:6, and its purity was 92.7%; aPC in PC was primarily a16:0-22:6 and a16:0-20:5, and aPC accounted for 90.2% of PC. We thereafter evaluated neurotrophic effects of Mytilus edulis pPE, aPC, and bovine brain pPE in a NGF-induced PC12 cell model. Morphologically, pPE and aPC could both promote differentiation, manifested in a significant increase in neurite length and number, due to increased expression of synaptophysin and growth protein GAP-43 in a dose-independent and structure-selective manner. Importantly, the effect on neuronal nutrition of pPE was better than aPC, and marine pPE was better than terrestrial pPE, which might be ascribed to vinyl-ether bond and differences in fatty acid composition.Abbreviations: AA: arachidonic acid; DHA: docosahexaenoic acid; EIC: extracted ion chromatogram; EPA: eicosapentanoic acid; GAP: growth-associated protein; HPLC: high-performance liquid chromatography; LC-MS/MS: liquid chromatography-tandem mass spectrometry; LPC: lyso-PC; LPE: lyso-PE; MS: mass spectrometry; NGF: nerve growth factor; PC: phosphatidylcholine; aPC: plasmanylcholine; PE: phosphatidylethanolamine; pPE: plasmenylethonoamine; PG: phosphoglycerols; PLs: phospholipids; PS: phosphoserines; TIC: total ion chromatogram.

Neuroprotection and immunomodulation in the gut of parkinsonian mice with a plasmalogen precursor.

Parkinson's disease (PD) is the second most common neurodegenerative disease worldwide. It is typically associated with motor symptoms originating from the degeneration of nigrostriatal dopamine (DA) neurons. Early stages of PD have been associated with an alteration in DA production in intestinal DAergic neurons along with inflammation. Interestingly, decreased serum concentrations of ethanolamine plasmalogens (PlsEtn) have been reported in PD patients. Ethanolamine plasmalogens play a role in vesicular fusion and release during neurotransmission, and store neuroprotective polyunsaturated fatty acids, such as docosahexaenoic acid (DHA) and are strong anti-oxidants, highlighting areas of potential therapeutic interest. Docosahexaenoic acid is known to play important roles in both the central nervous and peripheral systems, in addition to acting as a precursor of several molecules that regulate the resolution of inflammation. The present study investigated the neuroprotective and anti-inflammatory properties of the DHA-containing PlsEtn precursor, PPI-1011, in the intestine of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mice. Treatment with PPI-1011 prevented the MPTP-induced decrease in PlsEtn levels. In addition it prevented the loss of tyrosine hydroxylase (TH) expression and reduced the infiltration of macrophages in the myenteric plexus of MPTP-treated mice. The protective effects of PPI-1011 were observed regardless of whether it was administered pre- or post- MPTP treatment. These results suggest that PPI-1011 has neuroprotective and anti-inflammatory properties in the gut and indicate its potential utility as a treatment for both early and more advanced stages of PD.

Scallop-derived plasmalogens attenuate the activation of PKCδ associated with the brain inflammation.

Activation of protein kinase C delta (PKCδ) has been linked to the neuroinflammation but the relationship with the various neurodegenerative diseases including the Alzheimer's disease (AD) was mostly elusive. In the AD brains, the special phospholipids, ethanolamine plasmalogens (Pls), were found to be reduced and our previous study showed that these lipids possess neuroprotective and anti-inflammatory functions. In the present study, we could find that these lipids can significantly attenuate the microglial expression of PKCδ in the neuroinflammation model and in the AD model mice brains. We also show an increase of PKCδ in the human postmortem AD brains. In addition, we also report that scallop derived Pls (sPls) inhibited the p38MAPK and JNK protein expression which are involved in the expressional regulation of PKCδ in the microglial cells. In addition, the lentiviral shRNA-mediated knockdown of PKCδ attenuated the LPS-induced p65 (NF-kB) activation and inflammatory cytokine expression, suggesting that the PKCδ can induce the inflammatory response which can be inhibited by the sPls. Taken together, our recent findings suggest that the sPls can attenuate the increased expression of PKCδ associated with the neuro-inflammation in the murine brain.

A comparative study of EPA-enriched ethanolamine plasmalogen and EPA-enriched phosphatidylethanolamine on Aß42 induced cognitive deficiency in a rat model of Alzheimer's disease.

Ethanolamine plasmalogen (pPE), a major phospholipid in neuronal membranes, is specifically reduced in postmortem brains from patients with Alzheimer's disease (AD). The purpose of the present study was to compare the effects of EPA-enriched ethanolamine plasmalogen (EPA-pPE) and EPA-enriched phosphatidylethanolamine (EPA-PE) on cognitive deficiency and illustrate the possible underlying mechanisms. SD rats were divided into four groups including the sham group injected with 0.9% saline and three amyloid-ß (Aß) infusion groups, Aß42 group, EPA-pPE group and EPA-PE group. EPA-pPE and EPA-PE were administered by gavage (150 mg kg-1 day-1), respectively, once a day for 26 days. Administration of EPA-pPE exerted better effects than EPA-PE in improving Aß-induced cognitive deficiency in a rat model of Alzheimer's disease. Further mechanical research indicated that EPA-pPE was superior to EPA-PE in regulating oxidative stress via increasing SOD activity and decreasing MDA level, as well as reducing GSK-3ß and tau phosphorylation. Moreover, EPA-PE was more effective than EPA-pPE at inhibiting the protein expressions of Bax and caspase 9. The results of neuro-inflammation and inflammasome activation showed that EPA-pPE exerted more significant effects than EPA-PE in inhibiting the expressions of TNF-α and IL-1ß, and decreasing NLRP3, pro-caspase 1 and caspase 1 levels. EPA-pPE alleviated Aß-induced neurotoxicity by inhibiting oxidative stress, neuronal injury, apoptosis and neuro-inflammation, which might depend on the vinyl ether linkage at the sn-1 position.

Oral ingestion of plasmalogens can attenuate the LPS-induced memory loss and microglial activation.

Plasmalogens (Pls) are the special phospholipids which were reported to be reduced in brain and blood samples of Alzheimer's disease (AD) patients, suggested a possibility that an oral ingestion of Pls may prevent the disease progression. Interestingly, the clinical study showed that the daily oral ingestion of Pls among the mild AD patients improved cognition. However, it is unknown of whether the oral ingestion of Pls inhibits the AD like changes in brain e.g., glial activation and accumulation of amyloid beta (Aß) proteins. To elucidate the beneficial effects of the Pls oral ingestion, we have used the chronic lipopolysaccharide (LPS) injection model mice where the glial activation and Aß accumulation were well reported. In the present study, we have found that the Pls drinking at the doses of 0.1 µg/ml and 10 µg/ml for 3 months attenuated the glial activation and accumulation of amyloid beta (Aß) proteins in the murine brain. Interestingly, the LPS injection reduced the hippocampal dependent memory in the control mice but the groups of Pls drinking mice showed a better performance in the memory test, suggesting that oral intake of Pls can inhibit LPS-mediated memory loss associated with a reduction of glial activation and Aß accumulation in the brain. We, therefore, suggest that the oral ingestion of Pls among the AD patients may also inhibit the glial activation resulting in the improvement of cognition.

Efficacy and Blood Plasmalogen Changes by Oral Administration of Plasmalogen in Patients with Mild Alzheimer's Disease and Mild Cognitive Impairment: A Multicenter, Randomized, Double-blind, Placebo-controlled Trial.

BACKGROUND: Plasmalogens (Pls) reportedly decreased in postmortem brain and in the blood of patients with Alzheimer's disease (AD). Recently we showed that intraperitoneal administration of Pls improved cognitive function in experimental animals. In the present trial, we tested the efficacy of oral administration of scallop-derived purified Pls with respect to cognitive function and blood Pls changes in patients with mild AD and mild cognitive impairment (MCI). METHODS: The study was a multicenter, randomized, double-blind, placebo-controlled trial of 24weeks. Participants were 328 patients aged 60 to 85years who had 20 to 27 points in Mini Mental State Examination-Japanese (MMSE-J) score and five or less points in Geriatric Depression Scale-Short Version-Japanese (GDS-S-J). They were randomized to receive either 1mg/day of Pls purified from scallop or placebo. The patients and study physicians were masked to the assignment. The primary outcome was MMSE-J. The secondary outcomes included Wechsler Memory Scale-Revised (WMS-R), GDS-S-J and concentration of phosphatidyl ethanolamine plasmalogens (PlsPE) in erythrocyte membrane and plasma. This trial is registered with the University Hospital Medical Information Network, number UMIN000014945. FINDINGS: Of 328 patients enrolled, 276 patients completed the trial (140 in the treatment group and 136 in the placebo group). In an intention-to-treat analysis including both mild AD (20≤MMSE-J≤23) and MCI (24≤MMSE-J≤27), no significant difference was shown between the treatment and placebo groups in the primary and secondary outcomes, with no severe adverse events in either group. In mild AD patients, WMS-R improved significantly in the treatment group, and the between group difference was nearly significant (P=0.067). In a subgroup analysis of mild AD patients, WMS-R significantly improved among females and those aged below 77years in the treatment group, and the between-group differences were statistically significant in females (P=0.017) and in those aged below 77years (P=0.029). Patients with mild AD showed a significantly greater decrease in plasma PlsPE in the placebo group than in the treatment group. INTERPRETATION: Oral administration of scallop-derived purified Pls may improve cognitive functions of mild AD. FUNDING: The Japanese Plasmalogen Society.

Analysis of Plasmalogen Species in Foodstuffs.

Ethanolamine plasmalogen (PlsEtn), which is present at high levels in brains, is believed to be involved in neuronal protection. The present study was performed to search for PlsEtn resources in foodstuffs. The foodstuffs examined showed a wide range of PlsEtn contents from 5 to 549 µmol/100 g wet wt. The marine invertebrates, blue mussel, and ascidian had high PlsEtn contents (over 200 µmol/100 g wet wt). Profiling of the molecular species showed that the predominant fatty acids of PlsEtn species were 20:5 (EPA) and 22:6 (DHA) at the sn-2 position of the glycerol moiety in marine foodstuffs, whereas major PlsEtn species in land foodstuffs were 20:4. Following quantitative analysis by multiple reaction monitoring, the ascidian viscera were shown to contain the highest levels of 18:0/20:5-PlsEtn and 18:0/22:6-PlsEtn (86 and 68 µmol/100 g wet wt, respectively). In order to evaluate a neuronal antiapoptotic effect of these PlsEtn species, human neuroblastoma SH-SY5Y cells were treated with ethanolamine glycerophospholipid (EtnGpl), purified from the ascidian viscera, under serum starvation conditions. Extrinsic EtnGpl from ascidian viscera showed stronger suppression of cell death induced by serum starvation than with bovine brain EtnGpl. The EtnGpl from ascidian viscera strongly suppressed the activation of caspase 3. These results suggest that PlsEtn, especially that containing EPA and DHA, from marine foodstuffs is potentially useful for a therapeutic dietary supplement preventing neurodegenerative diseases, such as Alzheimer's disease (AD).

Dietary plasmalogen increases erythrocyte membrane plasmalogen in rats.

BACKGROUND: Many disorders with plasmalogen deficiency have been reported. Replenishment or replacement of tissue plasmalogens of these disorders would be beneficial to the patients with these disorders, but effects of dietary plasmalogen on mammals have not been reported. METHODS: Plasmalogens were purified from chicken skin. The purified plasmalogens consisted of 96.4% ethanolamine plasmalogen (PlsEtn), 2.4% choline plasmalogen (PlsCho) and 0.5% sphingomyelin (SM). A diet containing 0.1% the purified plasmalogens (PlsEtn diet) was given to rats. Relative composition of phospholipids was measured by a high performance liquid chromatography (HPLC) method that can separate intact plasmalogens and all other phospholipid classes by a single chromatographic run. RESULTS: The PlsEtn diet given to Zucker diabetic fatty (ZDF) rats for 4 weeks caused decreases of plasma cholesterol and plasma phospholipid as compared to control diet. The other routine laboratory tests of plasma including triacylglycerol, glucose, liver and renal functions, albumin, and body weight were not different. Relative compositions of erythrocyte PlsEtn and phosphatidylethanolamine (PE) increased, and that of phosphatidylcholine (PC) decreased in PlsEtn diet group. The PlsEtn diet given to normal rats for 9 weeks again caused decrease of plasma cholesterol and phospholipid, and it induced increase of relative composition of PlsEtn of the erythrocyte membrane. The other routine laboratory tests of plasma and body weight were not different. CONCLUSIONS: Dietary PlsEtn increases relative composition of PlsEtn of erythrocyte membranes in normal and ZDF rats, and it causes decreases of plasma cholesterol and plasma phospholipids. Dietary PlsEtn for 9 weeks seemingly causes no adverse effect to health of normal rats.

Lymphatic absorption of plasmalogen in rats.

Plasmalogen is a subclass of phospholipids that is widely distributed in man and animals. Many physiological roles have been proposed for this lipid; however, there have been no reports on the intestinal absorption of plasmalogen. In the present study, we examined lymphatic absorption of plasmalogen after the duodenal infusion of emulsified brain phospholipids (BPL) containing plasmalogen (22 mol % of total phospholipids) and soyabean lecithin (SPL) (100 g emulsified phospholipid/l). Male Wistar rats with implanted cannulas in the mesenteric lymph duct and the duodenum were kept in a Bollman-type restraining cage, and were infused the emulsion after 1 d recovery with duodenal infusion of a glucose-NaCl solution. Lymphatic plasmalogen output was increased at 2-4 h after the switch to BPL emulsion, and peaked at 4-6 h. However, no increases were observed after SPL infusion. Lymphatic recovery of plasmalogen for 8 h was 198 nmol, which was 0.22 mol % of the total plasmalogen disappeared from the intestine. We did not detect any increases in long-chain fatty aldehydes, which are the degradation product of plasmalogen, either in the blood or the small intestine. We conclude that a small percentage but a significant amount of the plasmalogen was absorbed into the lymph.

Ingestion of plasmalogen markedly increased plasmalogen levels of blood plasma in rats.

Plasmalogens, a subclass of phospholipids, are widely distributed in human and animals, and are taken into the body as food. However, no data exist on the intestinal absorption or fate of ingested plasmalogen. Here, we determined whether dietary plasmalogen is absorbed and whether blood and tissue concentrations increased in normal male Wistar rats by using four separate experiments. Phospholipids containing more than 20 wt% of plasmalogen extracted from the bovine brain were incorporated into test diets (10-15 wt%). In experiment 1, we estimated the absorption rate by measuring the plasmalogen vinyl ether bonds remaining in the alimentary tract of rats after the ingestion of 2 g of test diet containing 91 micromol plasmalogen. The absorption rate of plasmalogen was nearly 80 mol% after 4 h, comparable to the total phospholipid content in the test diet. In experiment 2, we observed no degradation of the plasmalogen vinyl ether bonds under in vitro conditions simulating those of the stomach and small intestinal lumen. In experiment 3 we confirmed a comparable absorption (36 mol%) by using a closed loop of the upper small intestine in anesthetized rats 90 min after injecting a 10 wt% brain phospholipid emulsion. Feeding a test diet containing 10 wt% brain phospholipids for 7 d increased plasmalogen concentration threefold in blood plasma and by 25% in the liver; however, no increases were seen in blood cells, skeletal muscle, brain, lungs, kidneys, or adipose tissue (experiment 4). We concluded that dietary plasmalogen is absorbed from the intestine and contributes to a large increase in plasmalogen levels in blood plasma.