Flying-fox (Pteropus spp.) sperm membrane fatty acid composition,its relationship to cold shock injury and implications for cryopreservation success

The very large acrosome of Pteropus species spermatozoa is prone to damage during cooling procedures. Cryogenic succuss has been linked to membrane composition, therefore the lipid composition of five Pteropus species sperm acrosomal and plasma membranes were investigated to provide insight into reasons for cold shock susceptibility. Rapid chilling and re-warming of spermatozoa from three Pteropus species resulted in a decrease (P < 0.05) in acrosomal integrity. Biochemical analysis of lipids revealed that stearic acid (18:0) was the predominant saturated fatty acid and oleic acid (18:1, n-9) the predominant unsaturated fatty acid in both acrosomal and plasma membranes. Linolenic acid (18:3, n-3) was only detected in plasma membranes of Pteropus hypomelanus and was detected in acrosomal membranes of all Pteropus spp. studied (except Pteropus giganteus). Although detected in both plasma and acrosomal membranes of Pteropus vampyrus, docosahexaenoic acid (22:6) was not detected at all in Pteropus poliocephalus, only in trace levels in the acrosomal and plasma membranes of P. giganteus and P. hypomelanus and not in acrosomal membranes of Pteropus rodricensis. No difference was seen in the levels of polyunsaturated fatty acids (PUFAs) within plasma membranes, however PUFAs were lower (P < 0.05) in acrosomal membranes of P. giganteus compared with P. vampyrus. Pteropus spp. spermatozoa have a very low ratio of unsaturated/saturated membrane fatty acids (<0.5). Membranes containing more PUFAs are more fluid, so the use of cryogenic media which improves membrane fluidity should improve Pteropus spp. spermatozoal viability post-thaw.     

The advantages of LDL (Low Density Lipoproteins) in the cryopreservation of canine semen

A medium containing LDL (Low Density Lipoproteins, the cryoprotective component of chicken egg yolk) was compared with egg yolk for the preservation canine spermatozoa during the freeze–thaw process. Twenty sperm samples taken from 10 dogs were frozen in liquid nitrogen at −196 °C in seven different media: one control medium containing 20% egg yolk, and six test media containing 4%, 5%, 6%, 7%, 8%, and 10% LDL, respectively.Following thawing, sperm motility was assessed using a Hamilton-Thorne Sperm Analyser equipped with the CEROS 12 software. The percentage of motile spermatozoa was 55.3% in the 6% LDL medium (optimal concentration) compared with 27.7% in the egg yolk based medium (p < 0.05).In comparison with the egg-yolk medium, the LDL medium also resulted in an improved preservation of spermatozoa during the freezing process (p < 0.05) in terms of acrosomal integrity (FITC-PSA test), flagellar plasma membrane integrity (HOS test), and DNA integrity (Acridine Orange test).In addition, six Beagle bitches were inseminated twice, via the intra-uterine route, at an interval of 24 h; 200 × 10⁶ spermatozoa that had been previously frozen in the 6% LDL medium were used per insemination. All of the bitches became pregnant (gestation rate of 100%).In conclusion, the 6% LDL medium provides improved protection of the spermatozoa during the freeze–thaw process and a marked improvement in the motility parameters of canine spermatozoa in comparison with the control medium containing egg yolk alone.Finally, the use of LDL as a cryoprotectant for canine semen does not interfere with fertility.     

Fatty acid profiles,growth, and immune responses of neonatal lambs fed milk replacer and supplemented with fish oil or safflower oil

Diets supplemented with long chain, n − 3 polyunsaturated fatty acids (PUFA) have improved the health and performance of neonatal and growing animals. This study was conducted with lambs that were orphaned at approximately 1 day of age to determine whether supplementing milk replacer fed lambs with oils rich in long chain n − 3 or n − 6 PUFA would alter plasma lipid profiles and affect growth characteristics and immune functions. From days 1 to 28 of age, lambs had ad libitum access to commercial milk replacer. From days 7 to 28 of age, lambs received twice daily either 1 g of soybean oil, 1 g of fish oil, or 1 g of safflower oil per os in a gelatin capsule (n = 60 pens; 20 pens/treatment; one ewe and one ram with similar initial body weights/pen). On days 7, 14, 21, and 28 of age, lambs were weighed, and jugular blood was collected from ram lambs. Lymphocyte proliferation in vitro, differential white blood cell (WBC) counts, and weight gains were quantified. Plasma from days 7 and 28 was used for fatty acid analyses. Fish oil increased (P < 0.001) plasma total n − 3 fatty acid concentration and total n − 3:total n − 6 fatty acid ratio. Pen body weight (i.e., total lamb weight per pen) increased (P < 0.001) with day (day 7, 11.9 kg; day 14, 15.1 kg; day 21, 18.2 kg; and day 28, 21.2 kg), but oil treatment did not affect pen body weight. Neither oil treatment, day, nor oil treatment × day interaction were significant for pen body weight gains (3.5 kg), pen average daily gains (0.5 kg), pen milk intakes (19.0 kg), or pen gain:feed ratio (0.18) measured during three intervals: days 7–14; days 14–21; and days 21–28. Day, but not oil treatment, affected (P < 0.001) unstimulated, concanavalin A stimulated, and lipopolysaccharides stimulated lymphocyte proliferation: days 14, 21, and 28 proliferation > day 7 proliferation. For neutrophils per 100 WBC, the treatment × day interaction was significant (P < 0.05). Oil treatment and day affected (P < 0.01 and <0.05, respectively) lymphocyte numbers per 100 WBC. For monocytes, eosinophils, and basophils, neither oil treatment, day, nor the oil treatment × day interaction were significant. Fish oil altered plasma fatty acid profiles, but it did not seem to improve measures of the performance or immune function of healthy, milk replacer fed lambs.     

Effects of post-thaw incubation on motility,acrosomal integrity and in vitro fertilizing capacity of boar spermatozoa cryopreserved in 0.5 and 5 ml straws

The effect of post-thaw incubation (0 vs. 5 h at 15 °C) and straw size (5 vs. 0.5 ml) on motility, acrosomal integrity and in vitro fertilizing (IVF) capacity of cryopreserved boar spermatozoa was studied. In samples assessed immediately after thawing, no differences were found between the two straw sizes. After 5 h post-thaw incubation, all parameters, except polyspermy, decreased and, spermatozoa packaged in 5 ml straws showed better functional and IVF parameters than these in 0.5 ml straws.     

Enhancement in liver SREBP-1c/PPAR-α ratio and steatosis in obese patients: Correlations with insulin resistance and n-3 long-chain polyunsaturated fatty acid depletion

Sterol receptor element-binding protein-1c (SREBP-1c) and peroxisome proliferator-activated receptor-α (PPAR-α) mRNA expression was assessed in liver as signaling mechanisms associated with steatosis in obese patients. Liver SREBP-1c and PPAR-α mRNA (RT-PCR), fatty acid synthase (FAS) and carnitine palmitoyltransferase-1a (CPT-1a) mRNA (real-time RT-PCR), and n-3 long-chain polyunsaturated fatty acid (LCPUFA)(GLC) contents, plasma adiponectin levels (RIA), and insulin resistance (IR) evolution (HOMA) were evaluated in 11 obese patients who underwent subtotal gastrectomy with gastro-jejunal anastomosis in Roux-en-Y and 8 non-obese subjects who underwent laparoscopic cholecystectomy (controls). Liver SREBP-1c and FAS mRNA levels were 33% and 70% higher than control values (P < 0.05), respectively, whereas those of PPAR-α and CPT-1a were 16% and 65% lower (P < 0.05), respectively, with a significant 62% enhancement in the SREBP-1c/PPAR-α ratio. Liver n-3 LCPUFA levels were 53% lower in obese patients who also showed IR and hipoadiponectinemia over controls (P < 0.05). IR negatively correlated with both the hepatic content of n-3 LCPUFA (r = ? 0.55; P < 0.01) and the plasma levels of adiponectin (r = ? 0.62; P < 0.005). Liver SREBP-1c/PPAR-α ratio and n-3 LCPUFA showed a negative correlation (r = ? 0.48; P < 0.02) and positive associations with either HOMA (r = 0.75; P < 0.0001) or serum insulin levels (r = 0.69; P < 0.001). In conclusion, liver up-regulation of SREBP-1c and down-regulation of PPAR-α occur in obese patients, with enhancement in the SREBP-1c/PPAR-α ratio associated with n-3 LCPUFA depletion and IR, a condition that may favor lipogenesis over FA oxidation thereby leading to steatosis.     

Ethanol withdrawal increases glutathione adducts of 4-hydroxy-2-hexenal but not 4-hydroxyl-2-nonenal in the rat cerebral cortex

Ethanol withdrawal increases lipid peroxidation of the polyunsaturated fatty acid (PUFA) docosahexaenoate (22:6; n-3) in the CNS. To further define the role of oxidative damage of PUFAs during ethanol withdrawal, we measured the levels of glutathione adducts of 4-hydroxy-2-hexenal (GSHHE) and 4-hydroxy-2-nonenal (GSHNE) as biomarkers of brain lipid peroxidation of n-3 and n-6 PUFAs, respectively. In this study rats received an ethanol-containing diet for 6 weeks followed by withdrawal ranging from 0 to 7 days. GSHHE content was elevated (> 350%) in the cerebral cortex after 2 days of withdrawal with no change in GSHNE. The levels of GSHHE were significantly greater (2- to 20-fold) than those of GSHNE in multiple brain regions. Experiments demonstrated that intoxication and withdrawal did not alter the enzymatic rate of formation of GSHHE or GSHNE, but the rate of formation of GSHHE was higher (～ 50%) than that of GSHNE. These results indicate that selective oxidative damage to n-3 PUFAs occurs in the cerebral cortex as a result of ethanol withdrawal and that 4-hydroxy-2-hexenal is metabolized to the GSH adduct more efficiently than HNE.     

Antioxidant activity of blueberry fruit is impaired by association with milk

The antioxidant properties of dietary phenolics are believed to be reduced in vivo because of their affinity for proteins. In this study we assessed the bioavailability of phenolics and the in vivo plasma antioxidant capacity after the consumption of blueberries (Vaccinium corymbosum L.) with and without milk. In a crossover design, 11 healthy human volunteers consumed either (a) 200 g of blueberries plus 200 ml of water or (b) 200 g of blueberries plus 200 ml of whole milk. Venous samples were collected at baseline and at 1, 2, and 5 h postconsumption. Ingestion of blueberries increased plasma levels of reducing and chain-breaking potential (+ 6.1%, p < 0.001; + 11.1%, p < 0.05) and enhanced plasma concentrations of caffeic and ferulic acid. When blueberries and milk were ingested there was no increase in plasma antioxidant capacity. There was a reduction in the peak plasma concentrations of caffeic and ferulic acid (? 49.7%, p < 0.001, and ? 19.8%, p < 0.05, respectively) as well as the overall absorption (AUC) of caffeic acid (p < 0.001). The ingestion of blueberries in association with milk, thus, impairs the in vivo antioxidant properties of blueberries and reduces the absorption of caffeic acid.     

Total phenolics,flavonoids and antioxidant properties of three Ceropegia species from Western Ghats of India

The aim of present work was to assess the total phenolic content (TPC), total flavonoid content (TFC), phenolic compounds and antioxidant properties of various extracts of three Ceropegia spp.: Ceropegia spiralis, Ceropegia panchganiensis and Ceropegia evansii from Western Ghats of India. TPC of the samples varied from 0.3 ± 0.2 to 28.5 ± 0.3 mg TAE/g FW, whereas, TFC of the samples ranged between 0.1 ± 0.1 and 15.3 ± 0.3 mg RE/g FW. The major phenolic compounds identified were gallic acid, vanillin, cathechol and ferulic acid. All the extracts possess 1,1-diphenyl-2-picryl hydrazyl (DPPH) radical scavenging activity, ferric reducing antioxidant power (FRAP) as well as metal chelating ability and this was also supported by significant correlation with TPC and TFC. To the best of our knowledge, this is the first paper presenting comprehensive data on TPC, TFC, phenolic profile and antioxidant properties of the Ceropegia spp.     

Mammalian ALOX15 orthologs exhibit pronounced dual positional specificity with docosahexaenoic acid

Mammalian lipoxygenases (LOX) have been implicated in cell differentiation and in the pathogenesis of inflammatory, hyperproliferative and neurological diseases. Although the reaction specificity of mammalian LOX with n − 6 fatty acids (linoleic acid, arachidonic acid) has been explored in detail little information is currently available on the product patterns formed from n − 3 polyenoic fatty acids, which are of particular nutritional importance and serve as substrate for the biosynthesis of pro-resolving inflammatory mediators such as resolvins and maresins. Here we expressed the ALOX15 orthologs of eight different mammalian species as well as human ALOX12 and ALOX15B as recombinant his-tag fusion proteins and characterized their reaction specificity with the most abundantly occurring polyunsaturated fatty acids (PUFAs) including 5,8,11,14,17-eicosapentaenoic acid (EPA) and 4,7,10,13,16,19-docosahexaenoic acid (DHA). We found that the LOX isoforms tested accept these fatty acids as suitable substrates and oxygenate them with variable positional specificity to the corresponding n − 6 and n − 9 hydroperoxy derivatives. Surprisingly, human ALOX15 as well as the corresponding orthologs of chimpanzee and orangutan, which oxygenates arachidonic acid mainly to 15S-H(p)ETE, exhibit a pronounced dual reaction specificity with DHA forming similar amounts of 14- and 17-H(p)DHA. Moreover, ALOX15 orthologs prefer DHA and EPA over AA when equimolar concentrations of n − 3 and n − 6 PUFA were supplied simultaneously. Taken together, these data indicate that the reaction specificity of mammalian LOX isoforms is variable and strongly depends on the chemistry of fatty acid substrates. Most mammalian ALOX15 orthologs exhibit dual positional specificity with highly unsaturated n − 3 polyunsaturated fatty acids.     

Chemical and acidic composition of Longissimus dorsi muscle of Comisana lambs fed with Trifolium subterraneum and Lolium multiflorum

Aim of this study was to evaluate the effects of grazing on Trifolium subterraneum and Lolium multiflorum, as pure or associated crops, on the chemical composition and on the fatty acid profile of the intramuscular lipids of the meat of lambs. Forty Comisana male lambs, on average weighing 13.75 ± 1.90 kg, were divided into four homogenous groups of ten and called, in relation to the diet: group T those grazing on T. subterraneum; Group L on L. multiflorum; Group TL on adjacent monocultures of T. subterraneum and L. multiflorum (66.6 and 33.3% of surface, respectively); Group LT on adjacent monocultures of T. subterraneum and L. multiflorum (33.3 and 66.6% of surface, respectively). Every 10 days, samples of forage species ingested by grazing lambs were collected and analysed. At 90 days of age, with an average live weight of 25.44, 23.44, 24.69 and 24.75 kg for T, L, TL and LT group, respectively, all lambs were slaughtered and a sample of Longissimus dorsi muscle for each animal was collected to study the chemical and acidic composition. No significant differences among the groups were observed for the growth performance and for the chemical composition of the meat. As regards the fatty acid classes, significant differences (P < 0.05) were observed for the monounsaturated fatty acids, which were lower in the group T (35.46%) than those of the groups L (38.24%), TL (38.63%) and LT (38.59%), whereas, significant higher values for the group T were observed for the polyunsaturated fatty acids of the n-3 (4.49%) and n-6 (8.26%) series than those of the n-6 series for group L (6.79%; P < 0.05) and than those of both series for group LT (n-3 = 3.64%; P < 0.05 and n-6 = 6.43%; P < 0.05). The fatty acids that have significantly determined the modifications of the acidic classes were: oleic acid, which showed significant (P < 0.05) lower values in the group T (26.70%) than the levels observed in the groups L (30.33%), TL (30.39%) and LT (30.63%) and the linoleic, linolenic and rumenic acids which were significantly (P < 0.05) higher in the groups T (linoleic = 5.13%; linolenic = 1.97%; rumenic = 0.46%) and TL (linoleic = 4.75%; linolenic = 1.82%; rumenic = 0.41%) than those of the groups L (linoleic = 4.10%; linolenic = 1.52%; rumenic = 0.26%) and LT (linoleic = 3.95%; linolenic = 1.42%; rumenic = 0.33%). These differences could be due to the different dynamic activity of the cellulolytic bacteria in the rumen, related to the different levels of fibrous fractions of the diets. No significant difference was observed for saturated fatty acid, unsaturated/saturated fatty acids ratio and Atherogenic and Thrombogenic indices among the groups, whereas, PUFA/SFA ratio showed significant (P < 0.05) higher value in group T than that in the group LT.T. subterraneum monoculture grazed as monoculture (T) and in mixture with L. multiflorum (66/33, TL) increased the linoleic, linolenic and rumenic acids improving the dietetic-nutritional characteristics of the lamb meat.     

DNA integrity of fresh and frozen canine epididymal spermatozoa

The aims of this study were to evaluate: (1) the effect of cryopreservation on DNA fragmentation of canine epididymal spermatozoa, and (2) the potential protective effect of melatonin on post-thaw sperm quality (motility, morphology, acrosomal and DNA integrity). Epididymal spermatozoa were collected after orchiectomy of ten dogs. Sperm samples were frozen in the presence or absence of melatonin (1 mM). DNA fragmentation index (percentage of spermatozoa with fragmented DNA) was similar in fresh samples (3.3 ± 3.6) and samples frozen with (4.2 ± 3.8) or without (3.6 ± 3.7) melatonin. Sperm motility was significantly (p < 0.0001) higher in fresh compared to frozen samples. The presence of melatonin in the freezing extender did not affect the sperm motility. Proportions of spermatozoa with normal morphology were similar in fresh and frozen samples, irrespective of the presence of melatonin in the extender. Acrosome integrity was significantly decreased (p < 0.01) by cryopreservation, and melatonin did not exert any beneficial effects. In conclusion, DNA fragmentation of canine epididymal spermatozoa was not affected by the freezing procedure, and the presence of melatonin did not preserve motility and acrosome integrity which were adversely affected by cryopreservation. The evaluation of DNA status of thawed gametes is particularly relevant for epididymal spermatozoa since these spermatozoa are usually stored and used in assisted reproductive techniques.     

The cytochrome b5 reductase HPO-19 is required for biosynthesis of polyunsaturated fatty acids in Caenorhabditis elegans

Polyunsaturated fatty acids (PUFAs) are fatty acids with backbones containing more than one double bond, which are introduced by a series of desaturases that insert double bonds at specific carbon atoms in the fatty acid chain. It has been established that desaturases need flavoprotein-NADH-dependent cytochrome b5 reductase (simplified as cytochrome b5 reductase) and cytochrome b5 to pass through electrons for activation. However, it has remained unclear how this multi-enzyme system works for distinct desaturases. The model organism Caenorhabditis elegans contains seven desaturases (FAT-1, -2, -3, -4, -5, -6, -7) for the biosynthesis of PUFAS, providing an excellent model in which to characterize different desaturation reactions. Here, we show that RNAi inactivation of predicted cytochrome b5 reductases hpo-19 and T05H4.4 led to increased levels of C18:1n − 9 but decreased levels of PUFAs, small lipid droplets, decreased fat accumulation, reduced brood size and impaired development. Dietary supplementation with different fatty acids showed that HPO-19 and T05H4.4 likely affect the activity of FAT-1, FAT-2, FAT-3, and FAT-4 desaturases, suggesting that these four desaturases use the same cytochrome b5 reductase to function. Collectively, these findings indicate that cytochrome b5 reductase HPO-19/T05H4.4 is required for desaturation to biosynthesize PUFAs in C. elegans.     

Production of human milk fat substitutes enriched in omega-3 polyunsaturated fatty acids using immobilized commercial lipases and Candida parapsilosis lipase/acyltransferase

In human milk fat (HMF), palmitic acid (20–30%), the major saturated fatty acid, is mostly esterified at the sn-2 position of triacylglycerols, while unsaturated fatty acids are at the sn-1,3 positions, conversely to that occurring in vegetable oils.This study aims at the production of HMF substitutes by enzyme-catalyzed interesterification of tripalmitin with (i) oleic acid (system I) or (ii) omega-3 polyunsaturated fatty acids (omega-3 PUFA) (system II) in solvent-free media. Interesterification activity and batch operational stability of commercial immobilized lipases from Rhizomucor miehei (Lipozyme RM IM), Thermomyces lanuginosa (Lipozyme TL IM) and Candida antarctica (Novozym 435) from Novozymes, DK, and Candida parapsilosis lipase/acyltransferase immobilized on Accurel MP 1000 were evaluated. After 24-h reaction at 60 °C, molar incorporation of oleic acid was about 27% for all the commercial lipases tested and 9% with C. parapsilosis enzyme. Concerning omega-3 PUFA, the highest incorporations were observed with Novozym 435 (21.6%) and Lipozyme RM IM (20%), in contrast with C. parapsilosis enzyme (8.5%) and Lipozyme TL IM (8.2%). In system I, Lipozyme RM IM maintained its activity for 10 repeated 23-h batches while for Lipozyme TL IM, Novozym 435 and C. parapsilosis enzyme, linear (half-life time, t_1/2 = 154 h), series-type (t_1/2 = 253 h) and first-order (t_1/2 = 34.5 h) deactivations were respectively observed. In system II, Lipozyme RM IM showed linear deactivation (t_1/2 = 276 h), while Novozym 435 (t_1/2 = 322 h) and C. parapsilosis enzyme (t_1/2 = 127 h), presented series-type deactivation. Both activity and stability of the biocatalysts depended on the acyl donor used.     

Chemical and immunochemical identification of propanoyllysine derived from oxidized n-3 polyunsaturated fatty acid

It is known that n-3 polyunsaturated fatty acids (PUFAs), such as docosahexaenoic acid and eicosapentaenoic acid, are rapidly oxidized in vitro. N?-(propanoyl)lysine (propionyllysine, or PRL) is formed from the reaction of the oxidized products of n-3 PUFAs and lysine. To evaluate the oxidized n-3 PUFA-derived protein modifications in vivo, we have developed detection methods using a novel monoclonal antibody against PRL as well as liquid chromatography–mass spectrometry (LC/MS/MS). The antibody obtained specifically recognized PRL. A strong positive staining in atherosclerotic lesions of hypercholesterolemic rabbits was observed. We have also simultaneously identified and quantified both urinary PRL and urinary N?-(hexanoyl)lysine, using LC/MS/MS using isotope dilution methods. The level of urinary PRL (21.6 ± 10.6 μmol/mol of creatinine) significantly correlated with the other oxidative stress markers, 8-oxo-deoxyguanosine, dityrosine, and isoprostanes. The increase in the excretion of amide adducts into the urine of diabetic patients was also confirmed compared to healthy subjects. These results suggest that PRL may be good marker for n-3 PUFA-derived oxidative stress in vivo.     

Hyperuricemia influences tryptophan metabolism via inhibition of multidrug resistance protein 4 (MRP4) and breast cancer resistance protein (BCRP)

Hyperuricemia is related to a variety of pathologies, including chronic kidney disease (CKD). However, the pathophysiological mechanisms underlying disease development are not yet fully elucidated. Here, we studied the effect of hyperuricemia on tryptophan metabolism and the potential role herein of two important uric acid efflux transporters, multidrug resistance protein 4 (MRP4) and breast cancer resistance protein (BCRP). Hyperuricemia was induced in mice by treatment with the uricase inhibitor oxonic acid, confirmed by the presence of urate crystals in the urine of treated animals. A transport assay, using membrane vesicles of cells overexpressing the transporters, revealed that uric acid inhibited substrate-specific transport by BCRP at clinically relevant concentrations (calculated IC₅₀ value: 365 ± 13 μM), as was previously reported for MRP4. Moreover, we identified kynurenic acid as a novel substrate for MRP4 and BCRP. This finding was corroborated by increased plasma levels of kynurenic acid observed in Mrp4^?/? (107 ± 19 nM; P = 0.145) and Bcrp^?/? mice (133 ± 10 nM; P = 0.0007) compared to wild type animals (71 ± 11 nM). Hyperuricemia was associated with > 1.5 fold increase in plasma kynurenine levels in all strains. Moreover, hyperuricemia led to elevated plasma kynurenic acid levels (128 ± 13 nM, P = 0.005) in wild type mice but did not further increase kynurenic acid levels in knockout mice. Based on our results, we postulate that elevated uric acid levels hamper MRP4 and BCRP functioning, thereby promoting the retention of other potentially toxic substrates, including kynurenic acid, which could contribute to the development of CKD.     

Allometric scaling of fatty acyl chains in fowl liver,lung and kidney,but not in brain phospholipids

The phospholipid (PL) fatty acyl chain (FA) composition (mol%) was determined in the kidney, liver, lung and brain of 8 avian species ranging in body mass from 150 g (Japanese quail, Coturnix coturnix japonica) to 19 kg (turkey, Meleagris gallopavo). In all organs except the brain, docosahexaenoic acid (C22:6 n3, DHA) was found to show a negative allometric scaling (allometric exponent: B = ? 0.18; ? 0.20 and ? 0.24, for kidney, liver and lung, respectively). With minor inter-organ differences, smaller birds had more n3 FAs and longer FA chains in the renal, hepatic and pulmonary PLs. Comparing our results with literature data on avian skeletal muscle, liver mitochondria and kidney microsomes and divergent mammalian tissues, the present findings in the kidney, liver and lung PLs seem to be a part of a general relationship termed “membranes as metabolic pacemakers”. Marked negative allometric scaling was found furthermore for the tissue malondialdehyde concentrations in all organs except the brain (B = ? 0.17; ? 0.13 and ? 0.05, respectively). In the liver and kidney a strong correlation was found between the tissue MDA and DHA levels, expressing the role of DHA in shaping the allometric properties of membrane lipids.     

Arachidonic acid is important for efficient use of light by the microalga Lobosphaera incisa under chilling stress

The oleaginous microalga Lobosphaera incisa (Trebouxiophyceae, Chlorophyta) contains arachidonic acid (ARA, 20:4 n − 6) in all membrane glycerolipids and in the storage lipid triacylglycerol. The optimal growth temperature of the wild-type (WT) strain is 25 °C; chilling temperatures (≤ 15 °C) slow its growth. This effect is more pronounced in the delta-5-desaturase ARA-deficient mutant P127, in which ARA is replaced with dihomo-γ-linolenic acid (DGLA, 20:3 n − 6). In nutrient-replete cells grown at 25 °C, the major chloroplast lipid monogalactosylglycerol (MGDG) was dominated by C18/C16 species in both strains. Yet ARA constituted over 10% of the total fatty acids in the WT MGDG as a component of C20/C18 and C20/C20 species, whereas DGLA was only a minor component of MGDG in P127. Both strains increased the percentage of 18:3 n − 3 in membrane lipids under chilling temperatures. The temperature downshift led to a dramatic increase in triacylglycerol at the expense of chloroplast lipids. WT and P127 showed a similarly high photochemical quantum yield of photosystem II, whereas non-photochemical quenching (NPQ) and violaxanthin de-epoxidation were drastically higher in P127, especially at 15 °C. Fluorescence anisotropy measurements indicated that ARA-containing MGDG might contribute to sustaining chloroplast membrane fluidity upon dropping to the chilling temperature. We hypothesize that conformational changes in chloroplast membranes and increased rigidity of the ARA-deficient MGDG of P127 at chilling temperatures are not compensated by trienoic fatty acids. This might ‘lock’ violaxanthin de-epoxidase in the activated state causing high constitutive NPQ and alleviate the risk of photodamage under chilling conditions in the mutant.     

Seasonal dynamics and spatial distribution of epiphytic dinoflagellates in Peter the Great Bay (Sea of Japan) with special emphasis on Ostreopsis species

Studies of epiphytic dinoflagellates in Peter the Great Bay, Sea of Japan in 2008–2011 revealed the presence of 13 species. Five of the species are known as potentially toxic: Amphidinium carterae, A. operculatum, Ostreopsis cf. ovata, O. cf. siamensis and Prorocentrum lima. The maximum species richness and abundance of epiphytic dinoflagellates were observed in autumn (from September to October). Ostreopsis spp. were most widely distributed and predominated, amounting to 99% of the total density of dinoflagellates. Multi-year seasonal dynamics of Ostreopsis spp. in Peter the Great Bay showed that these cells appear as epiphyton in August after maximum warming of surface waters (22–24 °С) and disappear in early November, when the water temperature decreases below 7 °С. Ostreopsis spp. proliferation occurred in September, when the water temperature was 17.2–21.0 °C. The highest densities of Ostreopsis spp. were recorded on September 9, 2010 on the rhodophyte Neorhodomela aculeata – 230 × 10³ cells g⁻¹ DW or 52 × 10³ cells g⁻¹ FW. The spatial distribution of epiphytic dinoflagellates was investigated in the near-shore areas of Peter the Great Bay during the second half of September 2010 to evaluate the role of hydrodynamic conditions. Epiphytic dinoflagellates were not found in sheltered sites having weak mixing hydrodynamics. However, the abundances of Ostreopsis spp. were significantly higher at sites having moderate turbulence compared to biotopes experiencing strong wave action. Densities of Ostreopsis spp. were not significantly different on macrophytes with branched thallus of all taxonomic divisions. However, the average cell densities of Ostreopsis spp. on green algae with branched thallus were significantly higher than on green algae having laminar thallus.     

Glutathione-supplemented tris-citric acid extender improves the post-thaw quality and in vivo fertility of buffalo (Bubalus bubalis) bull spermatozoa

In this study we evaluated the effects of semen extender supplementation with different concentrations of glutathione (GSH) on buffalo (Bubalus bubalis) bull sperm motility, plasma membrane integrity, viability and DNA integrity as well as in vivo fertility. Semen from three Nili-Ravi buffalo bulls was collected, and qualified semen ejaculates (n = 18) were split into five aliquots for dilution (37 °C; 50 × 10⁶ spermatozoa ml^?1) with experimental tris-citric acid extender containing 0, 0.5, 1.0, 1.5 or 2.0 mM GSH. Extended semen was cooled to 4 °C, equilibrated and filled in French straws. The straws were kept on liquid nitrogen vapors (5 cm above the LN₂ level) for 10 min and plunged in liquid nitrogen for storage. Sperm motility (%), plasma membrane integrity (%), viability (%) and DNA integrity (%) were assessed at 0, 2 and 4 h post-thawing (37 °C). Extender supplementation with GSH (0.5, 1.0, 1.5 and 2.0 mM) increased sperm motility, plasma membrane integrity and viability in a dose dependent manner. Sperm DNA integrity was higher (p < 0.05) in all experimental extenders containing GSH when compared to the control extender (0 mM GSH). The in vivo fertility rate of cryopreserved buffalo bull (n = 2) spermatozoa was higher (p < 0.05) in extender containing 2.0 mM GSH compared to that of control. In summary, tris-citric acid extender supplemented with glutathione improved the freezability of buffalo bull spermatozoa in a dose dependant manner. Moreover, the addition of 2.0 mM GSH to the extender enhanced the in vivo fertility of buffalo (Bubalus bubalis) bull spermatozoa.