Landscape of Sequence Variations in Homologous Copies of FAD2 and FAD3 in Rapeseed(Brassica napus L.)Germplasm with High/Low Linolenic Acid Trait

Genetic manipulation(either restraint or enhancement)of the biosynthesis pathway ofα-linolenic acid(ALA)in seed oil is an important goal in Brassica napus breeding.B.napus is a tetraploid plant whose genome often har-bors four and six homologous copies,respectively,of the two fatty acid desaturases FAD2 and FAD3,which con-trol the last two steps of ALA biosynthesis during seed oil accumulation.In this study,we compared their promoters,coding sequences,and expression levels in three high-ALA inbred lines 2006L,R8Q10,and YH25005,a low-ALA line A28,a low-ALA/high-oleic-acid accession SW,and the wildtype ZS11.The expression levels of most FAD2 and FAD3 homologs in the three high-ALA accessions were higher than those in ZS11 and much higher than those in A28 and SW.The three high-ALA accessions shared similar sequences with the pro-moters and CDSs of BnFAD3.C4 and BnFAD3.A3.In A28 and SW,substitution of three amino acid residues in BnFAD2.A5 and BnFAD2.C5,an absence of BnFAD2.C1 locus,and a 549 bp long deletion on the BnFAD3.A3 promoter were detected.The profile of BnFAD2 mutation in the two low-ALA accessions A28 and SW is different from that reported in previous studies.The mutations in BnFAD3 in the high-ALA accessions are reported for thefirst time.In identifying the sites of these mutations,we provide detailed information to aid the design of mole-cular markers for accelerated breeding schemes.

Preparation of High-Purity Linolenic Acid from Oil of Lithospermum Erythrorhizon by Urea Inclusion and Column Chromatography

Aim To separate high purity linolenic acid from the oil of Lithospermumerythrorhizon growing in the Northeast of China. Methods Urea inclusion and column chromatographywere used. Results Unsaturated fatty acid was separated, with a purity of 99.30 wt% of linolenicacid. Conclusion The experiment shows excellent reproducibility and high feasibility for industrialproduction.

Preparation and Charaterization of Self-assembled Nanoparticles Based on Linolenic-acid Modified Chitosan

Chitosan was modified by conjugating coupling with linolenic acid through the 1-ethyl-3-（3-dimethylami- nopropyyl） carbodiimide （EDC）-mediated reaction. The degree of substitution 1.8% （ i.e. 1.8 linolenic acid group per 100 anhydroglucose units） was measured by ^1H NMR. The critical aggregation concentration （CAC） of the self-aggregate of hydrophobically modified chitosan was determined by measuring the fluorescence intensity of the pyrene as a fluorescent probe. The CAC value in phosphate-buffered saline （PBS） solution （pH 7.4） was 5 × 10^-2 mg mL^-1. The average particle size of selfaggregates of hydrophobically modified chitosan in PBS solution （pH7.4） was 210.8 nm with a unimodal size distribution ranging from 100 to 500 nm. Transmission electron microscopy （TEM） study showed that the formation of near spherical shape nanoparticles has enough structural integrity. The loading ability of hydrophibically modified chitosan （LA-chitosan） was investigated by using bovine serum albumin （BSA） as the model. The loading capacity of self-aggregated nanoparticles increases （ 19.85 % ± 0.04 % to 37.57 % ± 0.25 % ） with the concentration of BSA （0.1-0.5 mg mL^-1 ）.

Effects of Ce^(3+) on improvement of spectral characteristics and function of chloroplasts damaged by linolenic acid in spinach

Linolenic acid has great effects on the structure and function of chloroplast. We studied the effects of Ce3＋ on the improvement of chloroplast spectral characteristics and oxygen evolution damaged by linolenic acid in spinach. Results showed that Ce3＋ could decrease the light absorption increased by linolenic acid and promote the distribution of excitation energy to PS II and alleviate the decrease of PS Ⅱ fluo- rescence yield caused by linolenic acid. The linolenic acid treatments in various concentrations reduced the oxygen-evolving rate of chloroplasts, but the rate was accelerated since adding Ce3＋.

Effect of Altering Linoleic Acid and Linolenic Acid Dietary Ratios on the Performance and Tissue Fatty Acid Profiles of Prawn, Macrobrachium rosenbergii （de Man 1879） Post Larvae

A 90-day experiment was conducted to investigate the effects of different dietary linoleic acid （LA; 18：2n-6） and linolenic acid ratios （LNA; 18：3n-3） on growth induces, feed utilization and tissue fatty acid profile of freshwater prawn, Macrobrachium rosenbergii post-larvae （PL）. The experiment was conducted in cubic indoor fiberglass tanks, each holding 700 L in triplicate. Post-larvae with an average weight of 20.8 ± 0.20 mg were stocked at 80 PL m2. Five experimental isocaloric （15.06 MJ kgl digestible energy）, and isonitrogenous （30.45% digestible protein） diets were formulated by blending of soybean oil and linseed oil to containing five dietary LA/LNA ratios （7.80, 2.75, 1.28, 0.65 and 0.30）. The highest survival values were recorded for prawn PL fed diet containing 0.65 LA/LAN ratios. Growth indices of PL significantly increased （P 〈 0.05） with decreased dietary LA/LAN ratios to 0.65. The same trend was observed for the highest （P ≤ 0.05） protein efficiency ratio, protein productive value, fat retention, energy retention and best feed conversion ratio. The total whole tissue polyunsaturated fatty acid composition of M. rosenbergii PL was dominated by LA followed by LAN. Post larvae fed the diets containing higher LA/LNA ratios showed a higher tissue LA/LNA ratio. The obtained findings revealed that fatty acid patterns ofM. rosenbergii PL were influenced by fatty acid profiles of diets. The diet containing 0.65 LA/LNA ratio is recommended to obtaining optimum growth performance and feed utilization for M. rosenbergii PL.

Alleviation effects of Ce^(3+) on inhibition of photochemical activity caused by linolenic acid in spinach chloroplast

Linolenic acid has great effects on the structure and function of chloroplast. The function of Ce^3＋ on the improvement of chloroplast photoreduction activity and oxygen evolution damaged by linolenic acid in spinach by in vitro investigation was studied. Results showed that adding Ce^3＋ to the linolenic acid treated chloroplast could greatly decrease the reduction linolenic acid exerted on the whole chain electron transport rate and the photoreduction activity of photosystem Ⅱ （PSII） and photosystem Ⅰ （PSI） as well as the oxygen evolution rate of chloroplast. It indicated that Ce^3＋ had the ability to relieve the inhibition of the photochemical reaction of chloroplast caused by linolenic acid to some extent.

Evaluation of anti-tubercular activity of linolenic acid and conjugatedlinoleic acid as effective inhibitors against Mycobacterium tuberculosis

Objective: To evaluate a new pharmacological activity/effect of linolenic acid(α- and γ-form) and conjugated-linoleic acid(CLA) causing antibacterial activity against Mycobacterium tuberculosis(Mtb). Methods: The anti-Mtb activity/effect of linolenic acid and CLA were determined using different anti-Mtb indicator methods such as resazurin microtiter assay(REMA) and MGIT 960 system assay. The Mtb was incubated with various concentrations(12.5–200) μg/m L of the compounds and anti-Mtb first-line drugs for 5 d in the REMA, and for 3 wk in MGIT 960 system assay. Results: Linolenic acid and CLA obviously indicated their anti-Mtb activity/effect by strongly inhibiting the growth/proliferation of Mtb in a dosedependent manner in the REMA and the MGIT 960 system assay. Interestingly, linolenic acid and CLA consistently induced anti-Mtb activity/effect by effectively inhibiting the growth/proliferation of Mtb in MGIT 960 system for 21 d with a single-treatment, and their minimum inhibitory concentrations were measured as 200 μg/m L respectively. Conclusions: These results demonstrate that linolenic acid and CLA not only have effective anti-Mtb activity/properties, but also induce the selective-anti-Mtb effects by strongly inhibiting and blocking the growth/proliferation of Mtb through a new pharmacological activity/action. Therefore, this study provides novel perspectives for the effective use of them and the potential that can be used as potent anti-Mtb candidate drugs, as well as suggests the advantage of reducing the cost and/or time for developing a new/substantive drug by effectively repurposing the existing drugs or compounds as one of new strategies for the global challenge of tuberculosis.

Formulation ofα-linolenic acid microemulsion free of co-surfactant

A novel class ofα-linolenic acid-in-water microemulsion free of co-surfactant was investigated as potential food delivery systems.Rough demarcation within the transparent region was deduced from the results of conductivity and polarizing optical microscopy.The microemulsion mean hydrodynamic diameter and characterization were determined by dynamic light scattering and negative-staining TEM.The location of ALA molecules in the microemulsion formulations was determined by ~1H NMR spectroscopy.

Cotton BLH1 and KNOX6 antagonistically modulate fiber elongation via regulation of linolenic acid biosynthesis

BEL1-LIKE HOMEODOMAIN(BLH)proteins are known to function in various plant developmental processes.However,the role of BLHs in regulating plant cell elongation is still unknown.Here,we identify a BLH gene,GhBLH1,that positively regulates fiber cell elongation.Combined transcriptomic and biochemical analyses reveal that GhBLH1 enhances linolenic acid accumulation to promote cotton fiber cell elongation by activating the transcription of GhFAD7A-1 via binding of the POX domain of GhBLH1 to the TGGA cis-element in the GhFAD7A-1 promoter.Knockout of GhFAD7A-1 in cotton significantly reduces fiber length,whereas overexpression of GhFAD7A-1 results in longer fibers.The K2 domain of GhKNOX6 directly interacts with the POX domain of GhBLH1 to form a functional heterodimer,which interferes with the transcriptional activation of GhFAD7A-1 via the POX domain of GhBLH1.Overexpression of GhKNOX6 leads to a significant reduction in cotton fiber length,whereas knockout of GhKNOX6 results in longer cotton fibers.An examination of the hybrid progeny of GhBLH1 and GhKNOX6 transgenic cotton lines provides evidence that GhKNOX6 negatively regulates GhBLH1-mediated cotton fiber elongation.Our results show that the interplay between GhBLH1 and GhKNOX6 modulates regulation of linolenic acid synthesis and thus contributes to plant cell elongation.

Fatty acid analysis in the seeds of 50 Paeonia ostii individuals from the same population

Tree peony seeds are rich in α-linolenic acid(ALA), and the peony seed oil is now being produced in China. Paeonia ostii is the most widely used tree peony species for oil extraction, which is commercially called Fengdan and treated as a single cultivar. Here, 50 P. ostii individuals from the same population in northern China were randomly selected for fatty acids(FAs) analysis. Thirteen FAs were isolated, and the most abundant five were palmitic acid(5.31–6.99%), stearic acid(1.22–2.76%), oleic acid(18.78–28.15%), linoleic acid(11.86–26.10%), and ALA(41.11–57.51%). There were significant individual differences of plants in FA quality and quantity and the linoleic acid content in Plant No. 48 even exceeded the scope of 1–99%. Further statistical analysis indicated that most of the individual FAs, saturated FAs, unsaturated FAs, and total FAs levels showed significant positive correlations to each other, whereas the seed yield per plant was independent and not correlated to the factors mentioned above. Ward’s hierarchical clustering results grouped the 50 plants into four clusters based on FA contents and seed yield, and the seven plants in Cluster IV were identified as good candidates for oil production. Our results confirmed that the individual differences did occur in P. ostii and Fengdan cannot be simply treated as one uniform cultivar. Also, these results may help simplify the selection of plants for oil peony breeding and accelerate the development of the oil peony industry.

Docosahexaenoic Acid in Breast Milk Reflects Maternal Fish Intake in Iranian Mothers

To estimate essential fatty acid (FA) and long-chain polyunsaturated fatty acid (LCPUFA) concentrations in early breast milk (BM) in relation to habitual fish intake. BM was collected within 72-hours after delivery from consecutively included mothers, 60 in Guilan (coastal) and 60 in Kermanshah (inland) provinces. Mothers were interviewed to com-plete a food frequency questionnaire. The FA composition was measured with gas chromatography. Mothers in the coastal area had higher intake of fish/seafood. Consumption of saturated fat was higher in Kermanshah and olive intake was higher in Guilan. High fish/seafood intake was associated with higher docosahexaenoic acid (DHA) and lower arachidonic acid (AA)/DHA ratio in BM. There were no differences in linoleic and α-linolenic acid concentrations in BM between the provinces. N-3 FA and DHA concentration were significantly higher in Guilan than Kermanshah, but total n-6 FAs and AA did not differ and were high in both provinces. The ratios of total n-6/n-3 and AA/DHA in BM of mothers from Guilan were significantly lower than those in Kermanshah. The LCPUFA status in BM in two Iranian provinces was generally good and DHA was higher and the AA/DHA was significantly lower in mothers with high fish intake.

Docosahexaenoic Acid Ester Degradation Measured by FTIR-ATR with Correlation Spectroscopy

Highly unsaturated fatty acids such as docosahexaenoic acid and linolenic acid are prone to oxidation with a resulting loss of bioactivity and generation of malodorous degradation compounds. Degradation proceeds by formation of the corresponding hydroperoxyl free radical with subsequent oxidative cleavage at the double bond site. FTIR-ATR was used to detect degradation through spectral changes that were correlated to oxidative degradation. The spectrometer was equipped with a heated stage so that a series of spectra could be collected while the sample was exposed to elevated temperatures. The appearance of new bands at 972.8 cm﹣1, 2934 cm﹣1, and 3498 cm﹣1 with the disappearance of bands at 703.6 cm﹣1 and 3013 cm﹣1 were observed after heating and these spectral changes were evaluated by correlation spectroscopy analysis. These results provide a method to quickly detect the oxidative degradation of polyunsaturated fats. This approach is expected to be applied for the quality assessment of feed formulations fortified with omega-3 fatty acids.

Determination of Fatty Acid Composition by GC-MS and Potential Antioxidant and Antibacterial Properties of Rumex hastatus and Cytisus scoparius Seeds Grown in Kashmir

Rumex hastatus D. Don and Cytisus scoparius Linn. are medicinally important plants used in Ayurveda and food, areenriched with remarkable nervine, diuretic, sedative and various other therapeutic properties. In this study Soxhlet extractionproduces higher yield of seed extracts. Gas chromatography-mass spectrometry was used to determine fatty acid composition ofpetroleum ether extracts (PE). The major fatty acids were Linolenic acid 53.53% in R. hastatus and linoleic acid 59.42% in C.scoparius. Palmitic acid was also found in significant amount (16.58% and 19.33%) in R. hastatus and C. scoparius respectively. Thechloroform (RCF, CCF), ethyl acetate (REA, CEA), acetone (RAT, CAT) and methanol (RMT, CMT) extracts of R. hastaus and C.scoparius respectively, were examined for their antioxidant activity using 2,2-di-phenyl-1-picrylhydrazyl (DPPH), nitrobluetetrazolium (NBT), hydrogen peroxide (H2O2), 2,2ˈ-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) radical scavengingassay and antibacterial activity against Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus by Agar disc diffusionmethod. All the extracts showed good radical scavenging and antibacterial activities compared to butylated hydroxyltoluene andstreptomycin respectively. However, dominant radical scavenging effect was observed at 100 μg/mL and 200 μg/mL concentrationsof extracts and dominant zone of inhibition was shown by CAT, CMT, RAT and REA extracts. This study exhibited beneficialproperties of these plants as an ingredient (unsaturated fatty acids) for the formulation of various functional foods and a source ofnatural antioxidant and antibacterial agents for drug, design and development.

Correlation of Various Foods Intakes and Plasma Levels of Omega Fatty Acids in Healthy Japanese Old Men

Background: Trans fatty acids are considered to impair health and some ω fatty acids are protective against atherosclerosis and diabetes mellitus. Trans fatty acids are said to be formed by the partial hydrogenation of vegetable oils. Some amounts are produced in digestive organs of ruminants and present in dairy products or meat. It is important how much these intaken fatty acids influence their plasma levels. Methods: Plasma levels of fatty acids including transforms of healthy old men are measured by gas chromatography and correlations between various foods intakes and plasma levels of trans fatty acids, and ω fatty acids are examined. Results: Intake of fish resulted in increase in plasma levels of docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) but intake of meat such as beef, cow and chicken meat did not increase plasma levels of arachidonic acid (AA). Intakes of oils increased plasma levels of dihomo-g-linolenic acid significantly and AA to some extent. Conclusion: Plasma levels of EPA and DHA increased upon intakes of fish in Japanese old men. Oil intake but not meat intake increased DGLA significantly. These results may explain low incidence of cardiovascular diseases in Japanese people compared with American people whose plasma levels of DHA and EPA are lower.

Feeding Calcium Salts of Linseed Oil on Productive Performance and Milk Fatty Acid Profile in Grazing Dairy Cows

The objective of this study was to determine the effect of supplying calcium salts of linseed oil (Ca-FA) rich in omega-3 (α-linolenic acid) on the production and chemical composition of milk and its nutraceutical value in dairy cows in early lactation. The trial lasted 12 weeks (2 weeks for adaptation to lipids and 10 weeks of data collection). A total of 36 Holstein dairy cows with 58.0 ± 17.0 days in milk (DIM), 594.1 ± 92.4 kg BW, 2.6 ± 1.5 parity and 38.9 ± 9.3 kg milk day-1 were used in a randomized complete block design. The treatments were: 1) Omega-3 (O3): 5.2 kg DM day-1 of concentrate including 0.7 kg DM of Ca-FA + 13.5 kg DM day-1 of partial mixed ration (PMR) + 12 kg DM day-1 of alfalfa pasture (Medicago sativa) and 2) Control (C): diet similar to O3 but lipid supplementation was replaced by cracked corn grain so that the diets were isoenergetic. No treatment effect was detected (P > 0.05) for any milk production and composition variables, except for urea in milk that was slightly higher in O3 (P = 0.02). The treatment × week interaction was significant (P < 0.05) for fat yield and content, with differences (P < 0.01) only in the 3rd week of the data collection period in favor of group C (1.39 vs. 1.13 kg·day-1 and 3.86% vs. 3.23% for fat yield and content, respectively). Total DMI and PMR were similar (P > 0.05) between treatments. Concentrate intake was higher (P < 0.01) in C compared with O3. Pasture DMI tended (P = 0.06) to be greater for cows that received the O3 treatment compared with C. Total metabolizable energy (ME) intake was similar (P = 0.44) between treatments. No treatment effect was detected (P > 0.05) in rumen environment parameters. Supplementation with Ca-FA reduced (P < 0.05) the hypercholesterolemic fraction of milk (C12:0, C14:0 and C16:0, -13.6%, -7.4% and -9.0%, respectively). The concentration of α-linolenic acid (C18:3n-3) increased (108%, P < 0.01) in O3 group compared with group C. The absence of negative effects of lipids on the fat content of milk and ruminal fermentation suggests that protection by saponification was effective. The supplementation with Ca-FA (0.85 kg·day-1) improved the healthy value of the milk.

Gradient Temperature Raman Spectroscopy of Fatty Acids with One to Six Double Bonds Identifies Specific Carbons and Provides Systematic Three Dimensional Structures

Specialized pro-resolving mediators provide promising targets for new drugs and natural products. Much work has been accomplished on the structure/ function of the lipoxygenase and cyclooxygenase enzymes but not on the substrates. A better visualization of three-dimensional lipid structures will allow increased refinement of the interactions that produce the pro-resolving mediators, and lead to improvements in synthetic pathways. We present systematic analysis of oleic (18:1n-9), linoleic (18:2n-6), alpha-linolenic (18:3n-3), arachidonic (20:4n-6), docosapentaenoic (22:5n-3), and docosahexaenoic (22:6n-3) acids. Continuous gradient temperature Raman spectroscopy (GTRS) applies the temperature gradients utilized in differential scanning calorimetry to Raman spectroscopy. GTRS can identify and differentiate specific carbon chain sites, finally allowing Raman analysis to explain why the long-chain polyunsaturated fatty acids (LC-PUFA) exhibit such extreme functional differences despite minimal changes in chemical structure. Detailed vibrational analysis of the important frequency ranges 1450 - 1200 cm-1 (includes CH2 bending and twisting) and 1750 - 1425 cm-1 (includes C=C stretching and C-C stretching plus H-C in-plane rocking) shows for the first time that each molecule has its own characteristic set of modes with only some redundancy/commonality. The number and frequency of modes correlates with three-dimensional molecular structure, not the degree of unsaturation. The high degree of specificity of lipoxygenase and cyclooxygenase enzymes should be reconsidered in light of the fact that individual sites on the polyunsaturated fatty acid chain are nonequivalent, and each LC-PUFA molecule has an individual, specific three dimensional structure incorporating torsion.

甘蓝型油菜高亚麻酸CMS的快速选育方法及效果研究

为了加速甘蓝型油菜高亚麻酸杂交育种进程。以高亚麻酸油菜种质为试验材料,采用快速选育方法进行油菜高亚麻酸CMS及保持系的选育。结果表明:选育周期从5~6年缩短为4年,亚麻酸含量由11.85%提升至16.96%,含油量由45.0%提升至48.8%,单株亚麻酸产出量由0.98 g提升至1.53 g。与原CMS(陕6A)相比,“YM-1A”的含油量、单株产量、亚麻酸含量、异交结实系数、单株亚麻酸产量分别提高8.45%、4.49%、59.43%、12.8%、80.65%;而自交结实系数降低56.86%。因不育性、农艺、品质等性状显著提高,YM-1A在甘蓝型油菜高亚麻酸杂交育种中具有较高的利用价值。

γ-亚麻酸通过下调氧化应激反应抑制NLRP3介导的焦亡并改善脑梗塞小鼠认知障碍

目的:探究γ-亚麻酸(GLA)对脑梗塞小鼠认知障碍的影响及机制。方法:选取18只雄性小鼠,分为对照组6只、模型组6只和GLA组6只,对模型组以及GLA组以大脑中动脉闭塞术进行脑栓塞小鼠认知障碍模型制作,对照组进行假手术,即手术过程相同但不插入线栓。对照组和模型组每日以1mg/kg生理盐水灌胃,GLA组每日以1mg/kg GLA灌胃14d。以Morris水迷宫评估小鼠的认知障碍情况。处死小鼠并取小鼠大脑组织,以HE染色观察小鼠脑梗死面积,以TUNEL染色观察小鼠神经元凋亡情况,以Western Blot实验检测小鼠海马体中P22、P47、NLRP3、IL-1β、GSDMD以及Caspase-1蛋白表达水平。结果:模型组小鼠的逃逸潜伏期短于GLA组以及对照组,GLA组则短于对照组,模型组的穿越环次数最多,GLA组次之,对照组最少;模型组大脑梗死面积以及神经元凋亡数目最多,GLA组次之,对照组大脑无梗死且无神经元凋亡;模型组P22、P47、NLRP3、IL-1β、GSDMD以及Caspase-1蛋白表达高于GLA组和对照组,GLA组的蛋白表达水平则高于对照组。结论:GLA通过下调氧化应激反应抑制NLRP3介导的焦亡并改善脑梗塞导致的小鼠认知障碍。

γ-亚麻酸对糖尿病小鼠记忆衰退的影响及机制

目的 探究γ-亚麻酸(GLA)对糖尿病小鼠记忆衰退的影响及机制。方法 选取16只8周龄野生型小鼠,雌雄各半,分成GLA组及对照组,对所有小鼠使用1 g/kg D-果糖加10%饮用水饮食(持续40 d)诱导糖尿病小鼠记忆衰退模型。对照组小鼠给予每日1 mg/kg生理盐水灌胃,GLA组小鼠给予每日1 mg/kg GLA灌胃。干预7 d后,以Morris水迷宫评估小鼠的学习和记忆障碍情况,处死小鼠取大脑皮层及海马组织,以HE染色检查二者病理改变,以透射电镜观察小鼠海马神经元超微结构变化,以Western Blot实验检测小鼠海马组织中GRP78、GRP94、NOX2和NOX4蛋白表达水平。结果 GLA组小鼠的逃逸潜伏期短于对照组,穿越环次数高于对照组(P<0.05)。GLA组小鼠大脑皮层和海马组织病理变化以及海马神经元超微结构损伤程度均较对照组轻。GLA组小鼠海马组织GRP78、GRP94、NOX2和NOX4蛋白表达水平低于对照组(P<0.05)。结论 GLA能通过抑制海马神经元内质网应激反应,进而改善糖尿病小鼠记忆衰退。