Methods For Determination Of Lipids Research Articles

Rapid method for lipid determination in Chlorella sp. based on Nile Red fluorescence

Microalgae are considered excellent organisms for biofuel source for abundant neutral lipids. However, the traditional gravimetric method has the problems of long measuring time and dangerous reagent. Therefore, a rapid and safer quantification method Nile red method was applied in Chlorella sp. L166. This study exams the influenced factors for Nile red method: emission wavelength, excitation wavelength, voltage, slit, Nile red concentration and staining time were 534 nm, 576 nm, 700 V, 5.0 nm, 4.0 μg/mL and 10 min, respectively. The fluorescence intensity of different concentration algal solution (OD680 = 0.2–1.2) showed a good correlation. Compared with the gravimetric method, there was no significant difference between these two methods. Fluorescence intensity stained with Nile red got reproducible results. The method could determine the lipid concentration in a short-time (measurement time was reduced by 13 times) and its safer, it might be a promising alternative for rapid lipid measurement in Chlorella sp.

A new fluorimetric method for simultaneous determination of lipid and protein hydroperoxides in muscle foods with the use of diphenyl-1-pyrenylphosphine (DPPP)

A new fluorimetric assay for simultaneous determination of lipid and protein hydroperoxides in muscle foods by the use of diphenyl-1-pyrenylphosphine (DPPP) was proposed in the study. Non-fluorescent DPPP that reacts with hydroperoxides stoichiometrically to yield fluorescent DPPP oxide, was used as a fluorescent dye to determine lipid and protein peroxidation in fish samples during frozen storage. The novel sensitive DPPP-assay developed for analysis under fluorescence microscopy conditions was based on this reaction. A good linear correlation between the amount of lipid hydroperoxides measured by the developed DPPP-assay and the peroxide value measured by iodometric titration was observed. Also, increase in fluorescence intensity from the reaction of DPPP with protein hydroperoxides in fish samples showed linear relation to the amount of cumene hydroxide added over a wide concentration range. The method was successfully applied on fish samples and can be used for fast assessment of primary oxidation stress in muscle foods.

A novel, sensitive and non-destructive method for quantitative determination of lipid in live Eriocheir sinensis using low-field 1H Nuclear magnetic resonance.

In this study, lipid content of live Eriocheir sinensis has been quickly and accurately determined by low‐field 1H Nuclear magnetic resonance (LF‐1H NMR). The experimental parameters of LF‐1H NMR have been optimized and the validity of the established standard method has been confirmed with traditional Soxhlet extraction method. Results show that the lipid signal intensity is strongly correlated with its content and exhibits a good linear correlation (Y = 0.0376 + 4.899X, R 2 = 0.9999), thus demonstrating favorable accuracy and sensitivity for the quantitative determination of lipid content. In conclusion, the lipid content of live E. Sinensis can be directly obtained based on an established method, indicating a great application potential in food and other fields.

Improved production of fatty acids by Saccharomyces cerevisiae through screening a cDNA library from the oleaginous yeast Yarrowia lipolytica.

Biological production of fatty acid (FA)-derived products has gained increasing attention to replace petroleum-based fuels and chemicals. FA biosynthesis is highly regulated, and usually it is challenging to design rational engineering strategies. In addition, the conventional 'one sample at a time' method for lipid determination is time consuming and laborious, and it is difficult to screen large numbers of samples. Here, a method for detecting free FAs in viable cells using Nile red staining was developed for use in large-scale screening. Following optimization of the method, it was used for screening a cDNA library from the oleaginous yeast Yarrowia lipolytica for identification of genes/enzymes that were able to enhance free FA accumulation in Saccharomyces cerevisiae. Several novel enzymes resulting in increasing FA accumulation were discovered. These targets include a GPI anchor protein, malate dehydrogenase, glyceraldehyde 3-phosphate dehydrogenase, FA hydroxylase, farnesyltransferase, anoctamin, dihydrolipoamide dehydrogenase and phosphatidylethanolamine-binding protein. The best enzyme resulted in a 2.5-fold improvement in production of free FAs. Our findings not only provide a novel method for high-throughput evaluation of the content of free FAs, but also give new insight into how enzymes from Y. lipolytica may increase the production of fatty acids in S. cerevisiae.

Rapid determination of total lipids in fish samples employing extraction/partitioning with acetone/ethyl acetate solvent mixture and gravimetric quantification

A simple, non-laborious, fast, cheap, robust and environment friendly method for lipid determination in fish samples was developed. An amount of 5 g of fish tissue homogenate in a centrifuge tube was extracted with 5 mL of acetone/ethyl acetate mixture (6:4, v/v) using vigorous shaking and after addition of inorganic salts (2 g MgSO4 and 0.5 g NaCl) and agitation the organic phase was separated by centrifugation. An aliquot of the organic phase was air-dried and then oven-dried and the lipid content of the fish homogenate was determined gravimetrically on a wet weight basis. The performance of the proposed method was compared with a well-established method according to Smedes (1999) by analysing 66 fish samples with a wide range of lipid content (pollock, chub and salmon). In comparison with the method developed by Smedes, similar repeatability and good agreement between the results from both methods for all analysed samples was achieved. The linearity of the proposed method with a correlation coefficient of 0.9997 was obtained between 0.25 and 15% and limits of detection (LOD) and quantification (LOQ) were 0.11% and 0.34% of total lipids in fish homogenate, respectively. The accuracy of the method was verified by the analysis of NIST standard reference material SRM 1946 (Lake Superior Fish Tissue) and the obtained recovery (at 10.17% of extractable fat) was 99.3% with a relative standard deviation (RSD) of 0.5% (n = 3).

Development and validation of TLC-densitometric method for determination of lipid A adjuvant as a bulk and in solid fat nanoemulsions.

A simple, sensitive, selective and precise high-performance thin-layer chromatographic method was developed for determination of lipid A (MPLA) adjuvant as a bulk and in solid fat nanoemulsions. Chromatographic separations were performed on thin-layer chromatography aluminum plates precoated with silica gel 60 F-254 as stationary phase and chloroform-methanol-ethyl acetate solution (10:2:4, v/v/v) as mobile phase. With this solvent system, compact spots for MPLA at Rf value 0.80 ± 0.02 were obtained. Densitometric analysis of MPLA was carried out in absorbance mode at 357 nm. Linear regression analysis for the calibration plots showed good linear relationship with r = 0.9996 in the concentration range of 20-100 ng/spot. The mean values (±SD) of slope and intercept were found to be 7.355 ± 0.006 and 109.52 ± 0.170, respectively. Limits of detection (LOD) and quantitation (LOQ) were observed at 3.096 and 9.382 ng/spot, respectively.The method was validated for precision, accuracy, robustness and recovery as per the International Conference on Harmonization guidelines. Statistical analysis proved that the developed method for quantification of MPLA as a bulk and in solid fat nanoemulsions is reproducible, selective and economical. This method could be applied for quantitative assay of MPLA in lipid-based vaccine formulations.

WEFTA Interlaboratory Comparison on Total Lipid Determination in Fishery Products Using the Smedes Method

Lipid determination by the Smedes method was tested in an interlaboratory trial performed by nine laboratories from seven countries belonging to the West European Fish Technologists Association Analytical Methods Working Group. Five samples of fish and fishery products with different lipid contents, including two blind duplicates, were distributed among the participants. All laboratories applied a slightly modified Smedes method, which included extraction of lipids by cyclohexane and isopropanol, transfer of lipids to the cyclohexane phase by addition of water, phase separation by centrifugation, and gravimetric lipid determination. The results indicate that the RSD for reproducibility (RSD(R)) was between 4.11 and 6.31% for samples with moderate (7%) and high (14%) lipid content, depending on the sample. Larger SDs among the laboratories were obtained for a cod sample with low lipid content of 0.5%. The method is judged to be suitable as a routine method for lipid determination in fish and fishery products.

Evaluation of the Bligh & Dyer lipid determination method

The Bligh & Dyer method for lipid determination has been used for many years in several different ways. To discover the success of this old method and assess the impact of deviations from it on the analytical result, the development of the method was evaluated using the equilibrium theory. In addition the actual phase compositions and phase volumes of their experiments were determined. Absorption of organic phase to the tissue appeared to be one of the main sources of incomplete extraction. The amount of lipid remaining in the aqueous phase is likely to be negligible, just like adsorption of lipids to the tissue. The kinetics of the extraction were promoted both by a multi-step extraction approach, and the methanol content. The latter is also expected to be a key parameter in determining the yield of lipid extraction.

Lipopolysaccharide determination by reversed-phase high-performance liquid chromatography after fluorescence labeling

A sensitive method for the quantitative determination of lipid A, the covalently bound hydrophobic component of lipopolysaccharides (endotoxins), has been developed. The determination of lipid A was based on the quantitative measurement of β-hydroxymyristic acid and β-hydroxylauric acid by reversed-phase HPLC. β-Hydroxy acids were liberated from ester and amide linkages in endotoxins by acid catalyzed methanolysis. The resulting methyl esters were derivatized with 9-anthracene-carboxyl chloride, 9-fluorene-carboxyl chloride and 4-(1-pyrenyl)butyric acid chloride and quantified with a fluorescence detector. The effectiveness of the three derivatizing agents was compared. As internal standards-β-hydroxytridecanoic acid [ β-OH(13:0)] and β-hydroxypentadecanoic acid [ β-OH(15:0)] ethyl esters were used. The limits of detection of β-hydroxymyristic acid were 0.5 pg for the 9-anthroyl and 2 pg for the fluorenoyl and 4-(1-pyrenyl)butyroyl ester per sample (signal-to-noise ratio of 3). The detection limits of lipopolysaccharide from a smooth strain ( Escherichia coli 0111:B4) was 20 pg, while that from two rough strains ( E. coli Nissle 1917 and Salmonella typhimurium SL 1181) was 5 pg per sample after methanolysis and derivatization with 9-anthroyl chloride.

Three extraction methods for determination of lipids in fish meal: Evaluation of a hexane/isopropanol method as an alternative to chloroform‐based methods

AbstractThree different extraction methods were compared for the determination of lipids in both oxidised and antioxidant‐stabilised brown fish meal. The three methods under investigation were those of Smith‐Ambrose‐Knobl using chloroform/methanol/water, Bligh and Dyer also using chloroform/methanol/water but monophasic, and Hara and Radin using hexane/isopropanol. The fat content, determined gravimetrically, was significantly lower (P < 0.05) when hexane/isopropanol was used for lipid extraction from both oxidised and antioxidant‐stabilised fish meal, than when either of the chloroform systems was used. Determination of lipid classes in the lipid extracts using silica gel thin‐layer chromatography on Chromarods‐SIII, with flame ionisation detection by latroscan, suggested that the lower lipid recovery by the hexane/isopropanol method may be due to a less effective extraction of the more polar lipid classes. including oxidation products, present in brown fish meal.

Methods of Lipid Analysis

A protocol for sample preparation before the determination in or isolation of lipids from human milk is presented. Methods for the direct determination of lipids including the creamatocrit, spectrophotometric (turbidi‐metric), and enzymatic methods are discussed. Procedures for the isolation of lipids including the Roese‐Gottlieb, modified Folch, and dry column extractions are compared. Finally, techniques for the separation and identification of lipid classes are described.

A rapid method for the accurate determination of lipid in animal tissues.

Download options Please wait... Article information DOI https://doi.org/10.1039/AN9729700562 Article type Paper Download Citation Analyst, 1972,97, 562-568 BibTex EndNote MEDLINE ProCite ReferenceManager RefWorks RIS Permissions Request permissions A rapid method for the accurate determination of lipid in animal tissues T. Atkinson, V. R. Fowler, G. A. Garton and A. K. Lough, Analyst, 1972, 97, 562 DOI: 10.1039/AN9729700562 To request permission to reproduce material from this article, please go to the Copyright Clearance Center request page. If you are an author contributing to an RSC publication, you do not need to request permission provided correct acknowledgement is given. If you are the author of this article, you do not need to request permission to reproduce figures and diagrams provided correct acknowledgement is given. If you want to reproduce the whole article in a third-party publication (excluding your thesis/dissertation for which permission is not required) please go to the Copyright Clearance Center request page. Read more about how to correctly acknowledge RSC content. Social activity Tweet Share Search articles by author T. Atkinson V. R. Fowler G. A. Garton A. K. Lough

A Turbulent Drier-Extractor and a Wide-Interface Extractor

Abstract A simpler modification of an original turbulent drier-extractor with provision for measuring moisture is presented, as well as methods for determination of lipids in serum and in urine in the wide-interface extractor, and a new use for the wide-interface extractor in washing extracts from the drier-extractor. The two extraction units described may also serve as a reference standard and may be preferred for routine lipid assays.

Rapid method for determination of lipid bound radioiodine in blood.

SummaryA method is reported for determination of lipid-bound radioiodine in blood by ion exchange resin technic. The method has been shown to be reproducible, accurate, rapid, and economical.