Changes in the Quality of Refined Fish Oil in an Accelerated Storage Study

ABSTRACT

The aim of this study was to prepare the refined oil derived from Sardinella longiceps by a step-wise procedure of degumming, bleaching, and deodorization. The refined sardine oil showed greater susceptibility toward oxidation than the crude oil, but the addition of ethyl acetate fractions of the red seaweeds Hypnea musciformis, Kappaphycus alvarezii, and Jania rubens with great antioxidant potential significantly increased the oxidative stability of the refined oil. The refined oil supplemented with the ethyl acetate fraction of these seaweeds indicated more noteworthy oxidative stability indices (≥ 0. 51 h) than α-tocopherol. The induction time was greater for refined oil added with H. musciformis (1.26 h) than butylated hydroxytoluene (1.04 h) and α-tocopherol (0.4 h). No traces of aromatic (δ 6.7–7.5 ppm), hydroperoxides, and aldehydes (δ 9–10 ppm) in the ¹H-NMR spectra of the refined oil supplemented with seaweed extracts at the baseline and after the accelerated storage study demonstrated that the seaweed extracts were able to prevent the formation of secondary oxidation products in the unsaturated system during storage. This study demonstrated the potential of seaweeds as natural alternatives to the synthetic antioxidants to prevent the rancidity of the refined fish for use in food and pharmaceutical industries.     

Eicosapentaenoic acid enrichment from sardine oil by argentation chromatography

Eicosapentaenoic acid (EPA) derived from chemically hydrolyzed sardine oil was concentrated by urea fractionation using methanol at different temperatures (2, 4, and 6 degrees C) and urea/fatty acid ratios (2:1, 3:1, and 4:1 w/w) and purified by argentation neutral alumina column chromatography. The individual fatty acids were determined as fatty acid methyl esters (FAME) by gas-liquid chromatography and gas chromatography-mass spectroscopy as FAME and N-acyl pyrrolidides. In the mass fragmentation pattern of FAME, the base peak was assigned to be the 1-methoxyethenol moiety (m/z = 74) obtained by McLafferty rearrangement. Formation of the cyclic tropylium ion (m/z = 91) in fatty acids with four or more double bonds was apparent in FAME-PUFAs. The base peak of N-acyl pyrrolidides was the McLafferty rearrangement ion, 1-(pyrrolidin-1-yl)ethenol (m/z = 113). The highest concentration of EPA (47.78%) was obtained at the crystallization temperature of 4 degrees C with a urea/fatty acid ratio of 4:1 (w/w) with 93.74% yield. After complexation of saturated and less unsaturated fatty acids by urea complexation, argentation chromatography resulted in an EPA of high purity (99.6%) with an overall recovery of 54.09% using 50% diethyl ether/n-hexane as eluting solvent. The peroxide (POV) and thiobarbituric acid (TBS) values were found to be highest (4.0 mequiv of O2/kg and 5.2 mg of malondialdehyde/kg, respectively) during urea fractionation at the higher crystallization temperature (6 degrees C) and higher urea/fatty acid ratio (4:1). Keywords: Sardine oil; eicosapentaenoic acid (EPA); fatty acid methyl esters (FAME); urea fractionation; argentation column chromatography.     

Synthesis and quantitative structure-activity relationships of oxanilates as chemical hybridizing agents for wheat (Triticum aestivum L.)

Chemical hybridizing agents (CHAs) can facilitate two-line breeding in heterosis programs of crops. Twenty-seven oxanilates having different aromatic substitutions were synthesized and screened as CHAs on two genotypes of wheat, PBW 343 and HD 2733, during two Rabi (winter) seasons, 2000-01 and 2001-02. The oxanilates prepared by thermal condensation of anilines with diethyl oxalate or by acylation with ethoxycarbonyl methanoyl chloride were sprayed at 1000 and 1500 ppm at the premeiotic stage of wheat, when the length of the emerging spike of the first node was 7-8 mm. Pollen sterility and spikelet sterility were measured in each treatment. Ethyl oxanilates 5, 6, and 25,containing 4-F, 4-Br, and 4-CF(3) aromatic substituents, respectively, induced greater than 98% spikelet sterility, the desired level, at 1500 ppm. Quantitative structure-activity relationship analysis revealed a direct relationship between F(p) and molecular mass but an inverse relationship between MR, E(S), and R in influencing the bioactivity. Several F(1) hybrids were developed using 5, and at least one showed heterosis.     

Synthesis and screening of anilides having olefinic and alkyl moiety in the side chain as chemical hybridizing agents for wheat (Triticum aestivum L.)

Induction of male sterility by deployment of chemical hybridizing agents (CHAs) holds immense potential in heterosis breeding of wheat. A total of 21 anilides having different aromatic substitutions and side-chain variation were synthesized and screened as CHAs on three genotypes of wheat viz., PBW 343, HW 2046, and HD 2733, at winter season. Various anilides having vinyl moiety in the acyl side chain were synthesized by condensation between substituted anilines with different esters or acid chlorides. Another lead in the form of N-alkyl anilines also became evident. The percent male sterility data caused by CHAs revealed the significant contribution of anilides containing vinyl double bond incorporated in the form of closed ring structure viz., furyl moiety as the side chain. 4'-Fluoro-furyl anilide (1) and 4'-bromo-furyl anilide (2) are found to be promising lead CHAs for the design of highly active molecules. QSAR analysis revealed a direct relationship of field effect exemplified by the Swain-Lupton constant F(p) for the aromatic substitution but an inverse relationship of molar refractivity MR for the side chain. The negative influences of parachor for the acyl domain have been underlined. The real guiding principle for selectivity of CHA action was found to be the pi value. The CHAs act by mimicking UDP-glucose, the key substrate in the synthesis of callose, or lead to an imbalance in acid-base equilibrium in pollen mother cells resulting in dissolution of callose wall by premature callase secretion.     

Development and utilization of novel SSRs in foxtail millet [Setaria italica (L.) P. Beauv.]

Although the foxtail millet [Setaria italica (L.) P. Beauv.] is recently regarded as a model crop for studying functional genomics of biofuel grasses, its genetic improvement to some extent was limited due to the non‐availability of molecular markers, particularly the microsatellite markers and the saturated genetic linkage map. Considering this, we attempted to generate a significant number of microsatellite markers in cultivar ‘Prasad’. Two hundred and fifty‐six clones were sequenced to generate 41.82‐kb high‐quality sequences retrieved from genomic library enriched with dinucleotide repeat motifs. Microsatellites were identified in 194 (76%) of the 256 positive clones, and 64 primer pairs (pp) were successfully designed from 95 (49%) unique SSR‐containing clones. The 67.4% primer designing ability, 100% PCR amplification efficiency and 45.3% polymorphic potential in the parents of F₂ mapping population established the efficacy of genomic microsatellites. All the 64 microsatellite markers displayed high level of cross‐species amplification (~67%) in 10 millets and non‐millets species. These experimental findings suggest the utility and efficacy of SSRs in diverse genotyping applications, resolving QTLs, phylogenetic relationships and transferability in several important grass species.     

Nutritional and Antioxidative Attributes of Green Mussel (Perna viridis L.) from the Southwestern Coast of India

ABSTRACT

The present study reports the biochemical composition of Perna viridis from the southwestern coast of India. A balanced essential to nonessential amino acid ratio (0.7:1.1) along with the optimum and balanced quantities of vitamins, mineral nutrients, and low cholesterol contents characterized P. viridis. The n-3/n-6 polyunsaturated fatty acid ratio of P. viridis was found to be 3.7:5.3 and therefore may serve as an alternative to balance the higher intake of n-6 fatty acids. High oxyradical scavenging capacity (24–32%) and total phenolics (2–5 mg GAE^?1) suggest the nutraceutical potential of P. viridis.     

Evaluation of chemical compounds for induction of male sterility in wheat (Triticum aestivum L.)

The short-list of eleven chemical hybridizing agents (CHAs) showing 98% or more induction of male sterility were identified following application at the pre-meiotic stage of wheat. Among ethyl oxanilates, 4-fluoro (CHA A1), 4-bromo (CHA A2), 4-trifluoromethyl (CHA A5), and 4-cyano (CHA A3) derivatives; and among pyridones, 4-chloro (CHA B3), 4-fluoro (CHA B1), 4-bromo (CHA B2), and 4-trifluoromethyl (CHA B6) derivatives were the most promising. These agents showed no adverse effects on plant growth and yield. Ethyl 4-fluoro oxanilate (CHA A1) was tested on 29 wheat genotypes at 1500ppm and induced 99.76± 0.37% male sterility. Ethyl 4-fluoro oxanilate residues were non-detectable in grain and husk and thus appeared to have no lasting residue effects. An erratum to this article can be found at     

Anti-Inflammatory Concentrate Enriched with Substituted Oligofucans Derived from Brown Seaweed Turbinaria conoides (J. Agardh) Kützing and Its Safety Assessment on Wistar Rats

ABSTRACT

Aqueous extract of the seaweed Turbinaria conoides was purified to obtain an oligofucan-enriched seaweed concentrate (OESC). Oligofucans isolated were characterized as two types with (→1)-fucose-(2,3-diSO₃^?)-(1→4)-fucose-(2-SO₃^?)-(1→3)-fucose-(2,3-diSO₃^?)-(1→4)-fucose-(2-SO₃^?, 3-OAc)-(4→) and (→1)-fucose-(3-SO₃^?)-(1→4)-fucose-(2-SO₃^?)-(1→4)-fucose-(3-SO₃^?)-(1→4)-fucose-(2-SO₃^?)-(4→) motifs. A 90-day accelerated shelf-life study (50°C) showed that OESC maintained its antioxidant properties (free radical scavenging, reducing, lipid peroxidation inhibition, and chelating activities) even after 30 days. In vitro COX-2 and 5-LOX inhibitory properties of OESC (67.2 and 95.2%, respectively) showed no significant variation even at the 30th day. OESC significantly mitigated the carrageenan-induced inflammation in rats at 0–2 h (59.7–70.3% inhibition), which were greater compared to the synthetic NSAID aspirin. The safety of OESC has been assessed by acute (14 days) and subchronic (90 days) oral toxicity studies, which showed no toxicity-related significant changes in renal or hepatic function, hematological indices, and serum biochemical parameters in the OESC-treated Wistar rats. No histopathological alterations were observed in the vital organs of rats treated with OESC. LD₅₀ and sub-chronic no-observed-adverse-effect level (NOAEL) for this concentrate were found to be > 5,000 and 2,000?mg/kg BW, respectively. Hence, oligofucan-enriched seaweed concentrate is safe to consume without any adverse effects in the body.     

Antioxidative Status of a Nutrient Enriched Formulation of Perna viridis in a Time Series Shelf-Life Study

ABSTRACT

The freeze-dried concentrate of green mussel, Perna viridis, was added with naturally derived antioxidative oleoresins of rosemary and turmeric along with trace amounts of other natural additives to prepare different treatments and was subjected to accelerated shelf life study (50°C) to evaluate its oxidative stability. The total oxyradical scavenging activities of samples with higher concentrations of turmeric over rosemary demonstrated the prominent role of the turmeric extracts to arrest free radical chain reactions. Time dependent antioxidative status in relation to the nutritional compositions was evaluated to identify the oxidative changes for different treatments. The addition of natural additives with polyphenolic moieties in the treatments shielded the mussel’s unsaturated fatty acids against oxidation as evident by higher polyunsaturated fatty acids in the two treatments as compared to control after 90 days of accelerated shelf-life study. Higher n-3/n-6 ratios after 90 days’ storage were also observed in these treatments. A threshold level of combined turmeric (0.8 g/100 g) and rosemary (0.4 g/100 g) was sufficient for optimum antioxidant activity and to maintain nutritional composition of the green mussel extract at desired levels.     

Enrichment of eicosapentaenoic acid from sardine oil with Delta5-olefinic bond specific lipase from Bacillus licheniformis MTCC 6824

Lipase derived from Bacillus licheniformis MTCC 6824 was purified to homogeneity by anion exchange chromatography on Amberlite IRA 410 (Cl-) and gel filtration using Sephadex G-100 as judged by denaturing polyacrylamide gel electrophoresis. The purified lipase was used for hydrolysis of triacylglycerol in sardine oil to enrich Delta5-polyunsaturated fatty acids (Delta5-PUFAs) namely, arachidonic acid (5,8,11,14-eicosatetraenoic acid, ARA, 20:4n-6) and eicosapentaenoic acid (5,8,11,14,17-eicosapentaenoic acid, EPA, 20:5n-3). The individual fatty acids were determined as fatty acid methyl esters (FAMEs) by gas-liquid chromatography and gas chromatography-mass spectroscopy as FAMEs and N-acyl pyrrolidides. The enzyme exhibited hydrolytic resistance toward ester bonds of Delta5-PUFAs as compared to those of other fatty acids and was proved to be effective for increasing the concentration of EPA and ARA from sardine oil. Utilizing this fatty acid specificity, EPA and ARA from sardine oil were enriched by lipase-mediated hydrolysis followed by urea fractionation at 4 degrees C. The purified lipase produced the highest degree of hydrolysis for SFAs and MUFAs (81.5 and 72.3%, respectively, from their initial content in sardine oil) after 9 h. The profile of conversion by lipase catalysis showed a steady increase up to 6 h and thereafter plateaued down. Lipase-catalyzed hydrolysis of sardine oil followed by urea adduction with methanol provided free fatty acids containing 55.4% EPA and 5.8% ARA, respectively, after complexation of saturated and less unsaturated fatty acids. The combination of enzymatic hydrolysis and urea complexation proved to be a promising method to obtain highly concentrated EPA and ARA from sardine oil.