Determination of protein concentration in raw milk by mid-infrared fourier transform infrared/attenuated total reflectance spectroscopy.

This study investigates the potential use of attenuated total reflectance spectroscopy in the mid-infrared range for determining protein concentration in raw cow milk. The determination of protein concentration is based on the characteristic absorbance of milk proteins, which includes 2 absorbance bands in the 1500 to 1700 cm(-1) range, known as the amide I and amide II bands, and absorbance in the 1060 to 1100 cm(-1) range, which is associated with phosphate groups covalently bound to casein proteins. To minimize the influence of the strong water band (centered around 1640 cm(-1)) that overlaps with the amide I and amide II bands, an optimized automatic procedure for accurate water subtraction was applied. Following water subtraction, the spectra were analyzed by 3 methods, namely simple band integration, partial least squares (PLS) and neural networks. For the neural network models, the spectra were first decomposed by principal component analysis (PCA), and the neural network inputs were the spectra principal components scores. In addition, the concentrations of 2 constituents expected to interact with the protein (i.e., fat and lactose) were also used as inputs. These approaches were tested with 235 spectra of standardized raw milk samples, corresponding to 26 protein concentrations in the 2.47 to 3.90% (weight per volume) range. The simple integration method led to very poor results, whereas PLS resulted in prediction errors of about 0.22% protein. The neural network approach led to prediction errors of 0.20% protein when based on PCA scores only, and 0.08% protein when lactose and fat concentrations were also included in the model. These results indicate the potential usefulness of Fourier transform infrared/attenuated total reflectance spectroscopy for rapid, possibly online, determination of protein concentration in raw milk.

Allergy to sesame in humans is associated primarily with IgE antibody to a 14 kDa 2S albumin precursor.

The goal of this study was to identify and characterise the major allergen(s) of sesame seed. Detection of specific IgE to sesame proteins was performed with Pharmacia CAP System and Western blotting, after separation of sesame proteins by SDS-PAGE, using sera from 28 subjects diagnosed as allergic to sesame. The major allergen was separated by gel filtration chromatography and identified by selective proteolysis followed by peptide sequence analyses, employing electrospray-ionization mass spectrometer. Twenty-four of the 28 subjects had sesame-specific IgE. A 14 kDa protein belonging to the 2S albumin family was recognised by 22 of the 24 sera used. Subjecting the 14 kDa after HPLC separation to proteolysis with Lys C yielded 3 peptides, but only one reacted positively in the dot blot test. This peptide, corresponds in the whole protein chain to residues 24-94. The reactivity of the 14 kDa protein with most of the sera indicates that this is the major sesame allergen, later identified as 2S albumin precursor; and its peptide which reacted positively in the dot blot test evidently contains an epitope(s). Some minor sesame allergens, of higher molecular weight, were also revealed.

A two-step enzymatic resolution process for large-scale production of (S)- and (R)-ethyl-3-hydroxybutyrate.

An efficient two-step enzymatic process for production of (R)- and (S)-ethyl-3-hydroxybutyrate (HEB), two important chiral intermediates for the pharmaceutical market, was developed and scaled-up to a multikilogram scale. Both enantiomers were obtained at 99% chemical purity and over 96% enantiomeric excess, with a total process yield of 73%. The first reaction involved a solvent-free acetylation of racemic HEB with vinylacetate for the production of (S)-HEB. In the second reaction, (R)-enriched ethyl-3-acetoxybutyrate (AEB) was subjected to alcoholysis with ethanol to derive optically pure (R)-HEB. Immobilized Candida antarctica lipase B (CALB) was employed in both stages, with high productivity and selectivity. The type of butyric acid ester influenced the enantioselectivity of the enzyme. Thus, extending the ester alkyl chain from ethyl to octyl resulted in a decrease in enantiomeric excess, whereas using bulky groups such as benzyl or t-butyl, improved the enantioselectivity of the enzyme. A stirred reactor was found unsuitable for large-scale production due to attrition of the enzyme particles and, therefore, a batchwise loop reactor system was used for bench-scale production. The immobilized enzyme was confined to a column and the reactants were circulated through the enzyme bed until the targeted conversion was reached. The desired products were separated from the reaction mixture in each of the two stages by fractional distillation. The main features of the process are the exclusion of solvent (thus ensuring high process throughput), and the use of the same enzyme for both the acetylation and the alcoholysis steps. Kilogram quantities of (S)-HEB and (R)-HEB were effectively prepared using this unit, which can be easily scaled-up to produce industrial quantities.

The effects of bile acids on beta-adrenoceptors, fluidity, and the extent of lipid peroxidation in rat cardiac membranes.

Bile acids have been proposed as a causative factor for the cardiomyopathy of cholestatic liver disease, since they cause negative inotropism and chronotropism and attenuate cardiac responsiveness to sympathetic stimulation. Bile acids can also modify membrane fluidity and generate reactive oxygen species (ROS). The effects of 10(-6)-10(-3) M deoxycholic acid (DCA) and chenodeoxycholic acid (CDCA) and their taurine conjugates, TDCA and TCDCA, on (1) the binding characteristics of beta-adrenoceptors, (2) membrane fluidity, and (3) the extent of lipid peroxidation in rat cardiac membranes were assessed. The results were compared to the effects of the oxidant, 10(-4)-10(-3) M hydrogen peroxide (H(2)O(2)), and the membrane-fluidizing compound, 5 x 10(-5) M 2-(2-methoxyethoxy)ethyl 8-(cis-2-n-octylcyclopropyl)octanoate (A(2)C). Cardiac beta-adrenoceptor density alone was reduced at 10(-4) M bile acid concentration while, at 10(-3) M bile acids, reductions in both receptor density and affinity were seen. At 10(-4) M H(2)O(2), receptor number and affinity were reduced, whereas A(2)C increased receptor affinity without affecting receptor density. Bile acids (10(-3) M) and 10(-4) M H(2)O(2) reduced membrane fluidity. H(2)O(2) caused a concentration-dependent increase in the extent of lipid peroxidation, whereas the bile acids and A(2)C had no effect. Bile acids (10(-4) M) reduced beta-adrenoceptor density in the absence of variations in membrane fluidity and in the extent of membrane lipid peroxidation. This result suggests that bile acids, at concentrations equivalent to the plasma/serum total or estimated free bile acid concentration, may have a possible role in the etiology of cardiomyopathy of cholestatic liver disease. At 10(-3) M bile acid concentration, beta-adrenoceptor number and affinity were adversely affected, accompanied by a decrease in membrane fluidity but without any significant increase in the extent of membrane lipid peroxidation. Although cardiac beta-adrenoceptor density and affinity and membrane fluidity were adversely affected by bile acids, the relevance of these findings to our understanding of the etiological basis of hepatic cardiomyopathy is questionable, since such concentrations exceeded the highest concentrations seen in the plasma and/or tissues of patients with cholestatic liver disease.

Dietary oxidized linoleic acid enhances liver cholesterol biosynthesis and secretion in rats.

Based on studies showing that excretion of cholesterol is elevated in rats fed oxidized linoleic acid, we hypothesized that cholesterol metabolism is enhanced under such oxidative stress. Liver cholesterol biosynthesis and secretion and fecal cholesterol excretion were studied in rats fed for 4 weeks diets containing 10% oxidized linoleic acid. Incubation of liver slices with 1-(14)C acetate and intraperitoneal injection of 5-(3)H-mevalonate showed the occurrence of enhanced hepatic cholesterol biosynthesis and elevated liver cholesterol secretion in animals subjected to oxidative stress. In addition, impaired liver cholesterol uptake was suggested. Higher levels of excreted cholesterol observed in the experimental animals were accompanied by augmented levels of liver phospholipids, primarily phosphatidylcholine, which most likely increased to enable the excessive cholesterol excretion. This study thus demonstrates that ingestion of oxidized lipids causes profound alterations in cholesterol metabolism.

Enhanced peroxidation of proteins of the erythrocyte membrane and of muscle tissue by dietary oxidized oil.

Male weanling rats were fed on diets containing 10% of either oxidized or fresh (control) soybean oil for periods of 4 and 7 weeks. The ingestion of oxidized oil was found to reduce the content of free thiols in the erythrocyte membrane and to increase the amount of membrane-extractable spectrin. The level of reactive carbonyl groups was higher in the proteins of the ghosts and of the muscle tissue derived from the experimental animals. These alterations which are characteristic of peroxidized proteins, suggest that oxidative stress caused by oxidized dietary lipids may damage tissue proteins.

Dietary oxidized linoleic acid modifies lipid composition of rat liver microsomes and increases their fluidity.

The effect of dietary oxidized oil on the lipid composition, fluidity and function of rat liver microsomes was studied. Male growing rats were fed diets containing 10 g/100 g of a fresh (control) or oxidized (experimental) linoleic acid-rich preparation for 4 wk. High levels of fluorescent compounds and of thiobarbituric acid reactive substances indicated the occurrence of substantial lipid peroxidation in the microsomes of the experimental rats. The fluidity of the liver microsomes derived from rats fed the experimental diet was significantly higher than that of the membranes of the controls. This was due to profound differences in lipid composition of the liver microsomes, namely, a lower cholesterol to phospholipid molar ratio and a greater arachidonic acid content in the phospholipids of the rats fed the experimental diet. The fluidity differences were accompanied by greater activity of the microsomal enzymes, aldehyde dehydrogenase and NADPH cytochrome C reductase. The study demonstrated that ingestion of oxidized lipids caused profound alterations in membrane composition, fluidity and function. These alterations are likely to be associated with an enhanced cholesterol turnover, as indicated by the greater cholesterol excretion observed for the experimental rats.

Lovastatin decreases plasma and platelet cholesterol levels and normalizes elevated platelet fluidity and aggregation in hypercholesterolemic patients.

The lipid composition of whole platelets and the fluidity of platelet membranes, as well as the sensitivity of the cell to aggregation, were studied in type IIA hypercholesterolemic human subjects before and after treatment with lovastatin. Fourteen patients with primary hypercholesterolemia having initial cholesterol levels of 383 +/- 52 mg/dL (mean +/- standard deviation) were studied and compared with 21 control subjects having cholesterol levels of 187 +/- 32 mg/dL. Lovastatin was administered orally at a starting dose of 40 mg daily. The dose was increased to 80 mg daily for eight patients who did not achieve the target cholesterol level of 200 mg/dL at 6 weeks. Serum cholesterol level was decreased by 37% following 20 weeks' administration of the drug. The fluidity of platelet membranes expressed in terms of the fluorescence anisotropy parameter was determined using the probe 1,6-diphenyl-1,3,5-hexatriene (DPH). When compared with platelets obtained from normocholesterolemic controls, platelets from hypercholesterolemic patients had a higher molar ratio of cholesterol to phospholipids ([C/PL] 0.86 +/- 0.15 v 0.57 +/- 0.06 for controls) and of phosphatidylcholine to sphingomyelin ([PC/SM] 2.64 +/- 0.87 v 2.00 +/- 0.15 for controls), enhanced fluidity (anisotropy parameter at 37 degrees C of 0.892 +/- 0.066 v 0.977 +/- 0.065 for controls), and a greater tendency to aggregate (aggregation of 84.2% +/- 6.3% v 78.5% +/- 7.6% for controls).(ABSTRACT TRUNCATED AT 250 WORDS)

An abnormal platelet membrane lipid composition in patients with low and high blood cholesterol.

Platelet lipid composition was determined in subjects with hypercholesterolemia (HC) (9.17 ± 2.15 mmol/L), hypocholesterolemia (HYPOC) (3.29 ± 1.01 mmol/L), and normocholesterolemic controls (NC) (5.29 ± 0.82 mmol/L). Lipid composition was quantitated in washed platelets by measuring platelet cholesterol (C) and phospholipid (PL) content, C/PL molar ratio and platelet PL phosphatidylcholine (PC) to sphingmyelin (SM) ratio. There was a significant increase and a significant decrease in C/PL molar ratio in subjects with HC and HYPOC compared to normals (0.74 ± 0.06 and 0.53 ± 0.06 vs 0.59 ± 0.04, p<0.01, respectively). These alterations were due to a significant change in platelet C content, whereas, platelet PL content was stable. PC/SM ratio was markedly decreased in HYPOC compared to NC (0.80 ± 0.18 vs 2.03 ± 0.18, p<0.01, respectively). These results indicate that blood cholesterol is a major determinant of platelet membrane lipid composition. This effect may be of concern in relation to a possible excess morbidity in patients not only with HC, but also with a low blood cholesterol.

Abnormal erythrocyte rheology in patients with morbid obesity.

The objectives of this study were to investigate the rheological properties of the erythrocyte in patients with morbid obesity and to follow them up after a short-term weight loss. A fluorescent polarization method was used to assess erythrocyte membrane biorheology and to measure its fluidity. Eighteen subjects participated in the study: 8 healthy controls and 10 patients with morbid obesity. The erythrocyte membrane fluidity was obtained in the healthy subjects and in the patients with morbid obesity prior to and after a ten-day zero-calorie diet. Fluidity was determined by steady-state fluorescence polarization after incorporation of the lipophilic probe 1,6-diphenyl-1,3,5 hexatriene (DPH). With this method, the anisotropy parameter at 37 degrees C, which is inversely related to membrane fluidity, was obtained. The patients with morbid obesity revealed an abnormal erythrocyte rheology. The exhibited an abnormally low erythrocyte membrane fluidity as compared with normal subjects. The anisotropy parameter at 37 degrees C was 1.417 +/- 0.093 in these obese patients compared with 1.279 +/- 0.043 in normal-weight controls (p < 0.01). Upon a short-term significant weight loss from a body mass index (BMI) (weight/height2) of 39 +/- 5 to 36 +/- 5 kg/m2 (p < 0.05), the anisotropy parameter did not change (1.401 +/- 0.190). Thus, fluidity measurements permit recognition of an abnormal erythrocyte rheology in patients with morbid obesity. This abnormality may partially explain the excess cardiovascular and thromboembolic morbidity in morbid obesity.

Reduction of plasma cholesterol by lovastatin normalizes erythrocyte membrane fluidity in patients with severe hypercholesterolaemia.

The effect of lovastatin on erythrocyte membrane composition and fluidity was investigated in eight patients with severe hypercholesterolaemia (mean LDL-cholesterol of 7.2 mmol l-1). Lovastatin was administered at a dosage of 40-80 mg for 20 weeks and was discontinued for 5 weeks thereafter. Parallel to a 47% fall in plasma LDL cholesterol, there was a significant reduction (P < 0.01) in erythrocyte membrane cholesterol:phospholipid molar ratio, while erythrocyte membrane fluidity assessed by diphenylhexatriene (DPH) fluorescence polarization increased significantly (P < 0.01). Discontinuation of lovastatin resulted in the reversal of erythrocyte membrane composition and fluidity to pre-treatment values.

Effect of lovastatin on lipoprotein fluidity in patients with hypercholesterolaemia.

Lovastatin was administered to six hypercholesterolaemic patients (mean plasma cholesterol 450 mg dl-1). Plasma lipoproteins (VLDL, LDL, and HDL) were separated before and following 7 and 12 weeks treatment with lovastatin. Fluidity was quantified by fluorescence polarization measurements using 1,6-diphenyl 1,3,5 hexatriene (DPH) as the fluorescent probe. Lovastatin treatment resulted in a significant reduction of total plasma cholesterol, LDL cholesterol and VLDL cholesterol (-41%, -44%, -68%, respectively). Fluidity measurements showed significant (p < 0.01) increase in LDL fluidity by 11% and 21% after 7 and 12 weeks of lovastatin treatment, whereas, VLDL fluidity was increased by 27% after 12 weeks of therapy. HDL fluidity was not altered. These alterations in the fluidity of the atherogenic lipoproteins (LDL and VLDL) in hypercholesterolaemic patients may prove to be of significance in reducing the risk of atherosclerosis.

Dietary fish oil modulates the alkaline phosphatase activity and not the fluidity of rat intestinal microvillus membrane.

The effect of dietary fish (mackerel) oil on the function and fluidity of the intestinal microvillus membrane in 1-y-old rats was compared with that of coconut and soybean oils. The animals were fed diets containing 10% protein (derived from casein) and 15% oil. Intestinal microvillus membranes and RBC ghosts were isolated after a 6-wk feeding period and examined for fluidity by fluorescence polarization with 1,6-diphenyl-1,3,5-hexatriene. The functionality of the microvillus membrane was assessed by the activity of the intrinsic enzyme alkaline phosphatase. No differences in the fluidity were observed between the microvillus membranes or between the RBC ghosts derived from the various dietary groups. The alkaline phosphatase activity of the microvillus membrane was found to be the lowest in the membranes isolated from the fish oil-fed rats. In these membranes the contents of 20:5(n-3) and 22:6(n-3) fatty acids were higher than in the membranes derived from the coconut and soybean oil-fed animals. The cholesterol to phospholipid molar ratio was lower in the coconut oil-fed group than in the other two groups. It is suggested that the compensatory effect of the elevated cholesterol to phospholipid molar ratio on the membrane fluidity was such that no differences in the overall fluidity were detected. It is likely that the incorporation of these long chain fatty acids might have caused compositional changes in the lipid microenvironment of the enzyme. Such changes could alter the fluidity of these microdomains, thereby affecting the activity of the integral enzyme alkaline phosphatase.

Lovastatin inhibits low-density lipoprotein oxidation and alters its fluidity and uptake by macrophages: in vitro and in vivo studies.

Under experimental conditions, oxidized low-density lipoprotein (Ox-LDL) may possess atherogenic properties, as is evidenced by its contribution to cholesterol accumulation in macrophages. LDL was oxidized in a cell-free system by predialysis of the lipoprotein against EDTA-free buffer and the addition of copper ions. Oxidation of LDL in the presence of lovastatin (10 to 1,000 mumol/L) resulted in a time- and dose-dependent reduction in thiobarbituric-acid-reactive substances (TBARS) concentration that was accompanied by increased LDL lysine-amino-group reactivity, in comparison with Ox-LDL produced in the absence of lovastatin. At 100 mumol/L, the drug reduced malondialdehyde concentration and increased amino-group reactivity by 24% and 42%, respectively. However, lovastatin's antioxidant effect was limited relative to other antioxidants, such as probucol and vitamin E. The fluidity of Ox-LDL was substantially reduced in comparison with native LDL. However, lovastatin inhibited this reduction in fluidity by 20%. Upon incubation of J-774 macrophage-like cell line with Ox-LDL, the lovastatin-treated Ox-LDL induced a reduction in the cellular cholesterol esterification rate in comparison with the effect of Ox-LDL that was produced in the absence of the drug. In four patients with hypercholesterolemia, the effect of lovastatin therapy (20 mg/d) on the sensitivity of their LDL to in vitro oxidation was studied. In all patients, Ox-LDL prepared from LDL obtained during lovastatin treatment demonstrated a reduced TBARS content, an increased trinitrobenzenesulfonic acid (TNBS) reactivity, an increased fluidity, and an impaired uptake by macrophages. These results were similar to those obtained by adding lovastatin in vitro.

Food restriction and membrane fluidity.

The fluidity of the erythrocyte membrane derived from growing rats raised under restricted food intake was found to be higher than the fluidity of the respective membranes from ad libitum fed animals. Considering the apparent relationship between the decrease in membrane fluidity and aging, the results point to the possible beneficial effects of food restriction at a young age.

A Sex-dependent Effect of Aspirin on Platelet Membrane Fluidity.

Aspirin, 250 mg/day, was administered to 14 normolipidemic healthy subjects for 7 days. Platelet lipid composition was determined in washed platelets by quantitating cholesterol to phospholipid molar ratio. Platelet membrane fluidity was measured by steady state fluorescence polarization using the probe diphenyl hexatriene. Upon 7 days aspirin ingestion platelet lipid composition was not altered. There was a sex-dependent effect of aspirin administration on platelet membrane fluidity. Whereas overall platelet membrane fluidity did not change at 37°C, there was a significant decrease in female subjects; the anisotropy parameter which is inversely related to membrane fluidity increased from 0.937±0.043-1.048 ±0.027 (p<0.01). In male subjects there was an increase in platelet membrane fluidity, which was significant at 25° C; the anisotropy parameter decreased from 1.350 ± 0.039-1.283±0.023 (p < 0.05). These results indicate that aspirin alters the membrane dynamics of platelets. This effect results from mechanisms other than alterations in platelet cholesterol or phospholipid content and operates in an opposite direction in men and women.

Excessive dietary tryptophan enhances plasma lipid peroxidation in rats.

High plasma cholesterol levels and plasma lipid peroxidation are associated with atherosclerosis. The effect of excessive dietary tryptophan on plasma lipid peroxidation was studied in rats fed a diet containing soybean oil (control), as well as an atherogenic diet, containing coconut oil and cholesterol. Feeding the atherogenic diet resulted in a 5-fold increment in plasma cholesterol concentration with no significant effect of the tryptophan supplementation. The plasma obtained from the hypercholesterolemic rats exhibited a 67% increased lipid oxidation (measured as thiobarbituric acid reactive substances) in comparison to normocholesterolemic plasma. Dietary tryptophan supplementation increased plasma lipid peroxidation by 9 and 21% in the control and in the hypercholesterolemic animals, respectively. Similarly, the excessive dietary tryptophan enhanced macrophage cholesterol esterification rate by 40 and 38% following cell incubation with the plasma obtained from the control and from the hypercholesterolemic animals, respectively. Since tryptophan is the precursor of serotonin we have measured urine concentration of 5-hydroxyindoleacetic acid (5HIAA), the metabolite of serotonin, and found 22 and 118% elevation in 5HIAA in the tryptophan fed control and hypercholesterolemic rats, respectively. The direct effect of tryptophan and serotonin on in vitro lipid peroxidation was also studied. Low density lipoprotein (LDL) was peroxidized by incubation with copper ions in the presence of tryptophan or serotonin. Serotonin was shown to enhance LDL peroxidation whereas tryptophan had no effect on LDL peroxidation. We conclude that excessive dietary tryptophan may be atherogenic since it enhanced plasma lipid peroxidation in hypercholesterolemic rats and increased macrophage uptake of plasma cholesterol. These effects are probably associated with increased plasma concentration of serotonin following the consumption of a tryptophan supplemented diet.

Decreased very low density lipoprotein fluidity in familial hypercholesterolemia.

The fluidity of lipoproteins from normolipidemic subjects and from familial hypercholesterolemic patients was investigated by fluorescence polarization. The fluorescent probe DPH (1,6-diphenyl-1,3,5-hexatriene) was incorporated into the lipoprotein fractions, and assessment of the fluidity pattern of each particle was determined by fluorescence anisotropy measurements over a temperature range of 10 to 40 degrees C. The very low density lipoprotein (VLDL) of the hypercholesterolemic patients was found to be considerably more rigid than the respective VLDL of normolipidemic subjects. Analysis of the constituents of the various lipoproteins suggested that the large difference in the fluidity between hypercholesterolemic and normal VLDL patients might be due to increased VLDL cholesterol/triglyceride, cholesterol/protein and cholesterol/phospholipid ratios, which were 10, 2 and 1.4 fold higher, respectively, in the hypercholesterolemic VLDL patients. Decreased VLDL fluidity in familial hypercholesterolemic patients may be of importance in the pathogenesis of their accelerated atherosclerosis.

Dietary tryptophan enhances platelet aggregation in rats.

The effect of diets enriched with tryptophan (4 and 10 g/kg) on ADP induced platelet aggregation and on plasma lipids was studied with growing rats, following feeding periods of up to 3 weeks. Dietary tryptophan was found to enhance markedly ADP induced platelet aggregation. It appears that this effect was not related to plasma lipid levels. In vitro studies aimed to clarify the mechanism by which tryptophan exerts its action showed that the latter, at concentrations up to 100 micrograms/ml, had no effect on ADP induced platelet aggregation. On the other hand, serotonin, the metabolic product of tryptophan, increased the ADP induced platelet aggregation in a dose dependent pattern. The possible involvement of serotonin in the observed enhancement of platelet aggregation was further substantiated by the observed high levels of 5-hydroxyindole acetic acid, which is a catabolite of serotonin, in the urine of the experimental animals. It is conceivable that tryptophan enhances platelet aggregation in rats via its metabolite, serotonin, and this may in turn contribute to increased atheroslerotic risk.

Riboflavin deficiency and the function and fluidity of rat erythrocyte membranes.

The effect of riboflavin deficiency on the fluidity and function of the red blood cell (RBC) membrane and on the activity of some enzymes involved in antioxidant defense mechanisms was studied. Growing male rats were fed an experimental (riboflavin-deficient) or a control (riboflavin-supplemented) diet. Following 7 wk of feeding, RBC from riboflavin-deficient rats contained higher levels of peroxidation products, most likely due to decreased glutathione reductase activity. An elevation in glutathione peroxidase activity was also observed whereas the activity of catalase and superoxide dismutase was not affected. Membrane fluidity was studied by fluorescence polarization, using 1,6-diphenyl-1,3,5 hexatriene (DPH) as a probe. The fluidity of RBC membranes isolated from riboflavin-deficient rats was significantly lower than that of the controls. This decreased fluidity was accompanied by an increase in the activity of the membrane-bound enzyme acetylcholinesterase. This study demonstrated that a decrease in cells' ability to cope with peroxidative damage as a result of riboflavin deficiency may lead to changes in the fluidity and function of membranes.