Separation of conjugated linoleic acid methyl esters by silver-ion high performance liquid chromatography in semi-preparative mode

Silver-ion HPLC (Ag-HPLC) has been utilized to separate a variety of unsaturated fatty acid methyl esters (FAMEs) by configuration, location or number of olefinic or acetylenic bonds. Two analytical Ag-HPLC columns connected in series and an isocratic solvent system of acetonitrile (ACN) in hexane were used to fractionate 10-15 mg samples of a mixture of two deuterium-labeled isomers of conjugated linoleic acid (Z9.E11- and Z9,Z11-octadecadienoic acid-17,17,18,18-d4). "Baseline" (> 95%) resolution of the two isomers, which decreased with increasing weights of sample injected, was maintained by careful adjustment of the percentage of ACN in the ACN/hexane solvent system. Chemical purities of the isolated FAME were > 96%.     

The HPTLC separation of malondialdehyde from peroxidised linoleic acid

Peroxidised linoleic acid reacts with acidified thiobarbituric acid to give a chromogen absorbing maximally at 532 nm. This TBA reactant is chromatographically identical to the TBA-MDA adduct formed from linolenic acid and a pure MDA standard. Considerably less MDA is produced by peroxidised linoleic acid compared to that formed from linolenic acid. In this respect the type of acid used for the colour reaction appears to be important.     

Self-aggregated nanoparticles from linoleic acid modified carboxymethyl chitosan: Synthesis, characterization and application in vitro

The purpose of the present research work was to study the formation of linoleic acid (LA) modified carboxymethyl chitosan (LCC). Another objective was to evaluate effect of linoleic acid degree of substitution on loading capacity (LC), ADR loading efficiency (LE) and in vitro release profile of LCC nanoparticles. The hydrogel nanoparticles can be prepared using linoleic acid modified carboxymethyl chitosan (LACMCS) after the sonication. The critical aggregation concentration (CAC) of the self-aggregate of LA modified CMCS (LCC) was determined by measuring the fluorescence intensity of the pyrene as a fluorescent probe. The CAC values were in the range of 0.061–0.081 mg/mL. Self-aggregated nanoparticles exhibited an increased LC and LE, decreased sustained release with an increasing ratio of the hydrophobic LA to hydrophilic CMCS. LCC nanoparticles loaded with ADR exerted in vitro anticancer activity against Hela cells that was comparable to the activity of free (non-entrapped in nanoparticles) ADR.     

Oxidation-stable linoleic acid by inclusion in α-cyclodextrin

Linoleic acid is essential to the skin’s barrier function. Its skin-care and restorative properties make it attractive for use in cosmetics and dermatology. However, its sensitivity to oxidation and tendency to go rancid prevent its use in cosmetic preparations. But in the form of a molecular inclusion compound with α-cyclodextrin, linoleic acid is effectively protected against oxidation. Investigations into the storage and light stability, olfactory tests and headspace analysis of the formulations give evidence of the stability of a suitable inclusion compound. Reversible complexation makes it possible for the first time to use linoleic acid in various cosmetic formulations and personal-care products.     

Studies of lipase-catalyzed polyesterification of an unactivated diacid/diol system

Preliminary enzymatic polymerization studies in the simple stoichiometric adipic acid/butane-1,4-diol system using lipase B from Candida antarctica, immobilized as Novozym 435®, suggest that in solvent-free conditions a step-growth mechanism operates involving the sequential addition of an AB synthon by esterification mode only. Conversely, in toluene as solvent there is a change to the more facile transesterification mode in line with the conventional polyesterification procedure, pointing to a change in specificity of the lipase. Evidence is drawn from qualitative studies using a series of synthetic intermediates, enabling authentication of product mixtures together with an indication of the comparative reactivity of species along the proposed reaction pathway. © 1998 John Wiley & Sons, Inc. J Polym Sci A: Polym Chem 36: 2069–2080, 1998     

Analysis of interferents by means a D-optimal screening design and calibration using partial least squares regression in the spectrophotometric determination of Cr(VI)

Using a central composite design, the signal of the process for the spectrophotometric determination of hexavalent chromium (λ = 543 nm) is maximised and its variability minimised using as complexing agent 1,5-diphenylcarbazide in sufficiently acid medium. To analyse the interference of various analytes (Mo(VI), V(V), Fe(III) and Mn(VII)) on the Cr(VI) as a function of concentration of interferent, a factorial design was prepared at three levels of each (zero, medium and high concentration), which implies performing 81 determinations. However, a D-optimal design with just nine experiments is sufficiently good to estimate the model proposed.The interference of these metals makes it impossible to determine Cr(VI) when they are present in the sample. To avoid prior separation steps, a multivariate regression by partial least squares, PLS, is proposed to calibrate the Cr(VI) in the presence of these analytes varying the concentration of the Cr(VI) between 0.1 and 0.9 μg ml⁻¹ and that of the interferents between 3 and 5 μg ml⁻¹. The average errors obtained were 4.5% and 3.29% fitted and in prediction, respectively, with a standard error in prediction (RMSEP) of 0.016% presenting absence of both constant and proportional bias.The detection limit with the PLS regression in the presence of interferents is 0.1 μg ml⁻¹ with a probability of false positive equal to 5% and less than 5% for false negative. The capability of detection is similar to that obtained with the univariate calibration (absorbance at 543 nm) in absence of interferents.With the PLS regression it is possible to discriminate 0.085 μg ml⁻¹ of Cr(VI) in a sample with 0.5 μg ml⁻¹ of Cr(VI) with probabilities of false compliance and false non-compliance equal to 0.05. For the univariate calibration without interferents, it was established at 0.0971 μg ml⁻¹ of Cr(VI) for the same nominal concentration.In relation to interference of V(V), Fe(III) and Mn(VII), the PLS calibration could be an efficient alternative to the separation step for Cr(VI) spectrophotometric determination using 1,5-diphenylcarbazide.     

Microwave-Assisted Synthesis of 5′-O-methacryloylcytidine Using the Immobilized Lipase Novozym 435

Nucleobase building blocks have been demonstrated to be strong candidates when it comes to DNA/RNA-like materials by benefiting from hydrogen bond interactions as physical properties. Modifying at the 5′ position is the simplest way to develop nucleobase-based structures by transesterification using the lipase Novozym 435. Herein, we describe the optimization of the lipase-catalyzed synthesis of the monomer 5′-O-methacryloylcytidine with the assistance of microwave irradiation. Variable reaction parameters, such as enzyme concentration, molar ratio of the substrate, reaction temperature and reaction time, were investigated to find the optimum reaction condition in terms of obtaining the highest yield.     

Usefulness of D-optimal designs and multicriteria optimization in laborious analytical procedures. Application to the extraction of quinolones from eggs

An analytical method has been developed to extract ciprofloxacin and enrofloxacin from eggs. The aim of this work is to determine the experimental conditions of extraction providing high recoveries with small standard deviations. An experimental design based on the D-optimality criterion and replicated three times was built to evaluate the effect of five factors related to the extraction which is the most inaccurate stage of the procedure. This non-classical design is needed because there are several practical constraints: (i) the extraction procedure is time-consuming, quinolones are not stable and the design must be performed in a single working session. (ii) The tube capacity of the centrifuge is 6, so the number of experiments will be 6 or a multiple of 6. In the optimal experimental conditions, the extraction is performed once with 5 ml of methanol. Then, fatty acids are removed with a mixture of hexane/ether. Analytes are finally separated and detected by HPLC-fluorescence without the additional step of purification by solid-phase extraction (SPE). Under these conditions, the mean recovery is 64% and 70% and the standard deviation 5% and 4% for ciprofloxacin and enrofloxacin, respectively. The capability of decision, CCalpha, is 3.1 and 2.8 microg kg(-1) of ciprofloxacin and enrofloxacin, respectively. The capability of detection, CCbeta, is 7.8 and 7.0 microg kg(-1) of ciprofloxacin and enrofloxacin, respectively. In both cases the probabilities of false positive, alpha, and of false negative, beta, were fixed at 0.05.     

正交设计和均匀设计用于硒-金膜修饰电极制备条件的选择

硒代胱氨酸在硒 金(Se Au)膜修饰玻碳电极上产生两个灵敏的氧化还原峰:峰II(-500mV左右)和峰III(-327mV左右),以峰II的峰电流作为评价指标,采用正交设计与均匀设计相结合的方法对Se Au膜修饰电极的制备条件进行优化得到最佳优化条件:底液0.1mol LKCl;沉积电位为-850mV;沉积时间为60s;SeO2浓度为8.3×10-3mol L;AuCl3浓度为8.9×10-4mol L。均匀设计的数据应用Matlab计算机软件处理。依此制备的Se Au膜修饰电极性能稳定,用于硒代胱氨酸伏安特性研究有良好的重现性。     

Optimization of supercritical fluid extraction of geniposidic acid from plantain seeds using response surface methodology

The process parameters of supercritical CO₂ (SCCO₂) plus modifer for the extraction of geniposidic acid from plantain seeds were studied using a Box–Behnken design. The effects of independent variables, that is, ethanol concentration (0–70%, ethanol:water, v/v), extraction pressure (10–30 MPa), and temperature (50–80°C) on the yield of geniposidic acid were evaluated. Results indicated that the data could be well fitted to a second-order polynomial model. The effects of ethanol concentration and temperature, as well as the interaction between ethanol concentration and temperature were significant (p < 0.05). The yield (8.9 mg/g) of modified SCCO₂ extraction at optimal conditions was compared with that obtained by Soxhlet extraction or ultrasound assisted extraction.     

Optimization of a solid-phase extraction procedure in the fluorimetric determination of sulfonamides in milk using the second-order advantage of PARAFAC and D-optimal design

The objective of this work is to optimize a solid-phase extraction procedure for the simultaneous determination of sulfadiazine, sulfamerazine, and sulfamethazine in milk by fluorimetric detection. For this task, an alternative strategy is employed, which allows one to reduce noticeably the number of experiments without losing the quality of the estimations. It consists of the use of a D-optimal design together with PARAFAC decomposition for the calculation of the response in the experimental design. Effects of amount of cartridge sorbent, kind of milk, volume of conditioning solutions, kind of wash and elution, and kind of mixture of sulfonamides have been evaluated, for maximizing sulfonamide mean recovery and minimizing its standard deviation. Since milk without sulfonamides may give some matrix effect over the fluorescence signal, its behavior has also been studied. Optimal conditions have been selected where the ratio between sulfonamide recovery and milk without sulfonamides was the highest, which are 500 mg of cartridge sorbent, acid wash, and elution and 3 mL of conditioning solutions. The type of milk and mixture of sulfonamides not significant. This makes the procedure suitable for the combined determination of sulfadiazine, sulfamerazine, and sulfamethazine in any kind of milk. Finally, an experimental procedure is proposed, obtaining a sulfonamide mean recovery equal to 68.5% with values of standard deviation between 7 and 8 μg kg⁻¹.     

Revisiting the design of experiments for copolymer reactivity ratio estimation

Tidwell and Mortimer used the D-optimal criterion to design terminal model reactivity ratio estimation experiments. In this paper, two problems are investigated which were not addressed by Tidwell and Mortimer. The first problem is the design of reactivity ratio estimation experiments subject to composition constraints. It is shown that all the key information contained in the D-optimal criterion can be summarized in two equations: one which is a function of r?₁, and one which in a function of r?₂. These two equations can be used to design reactivity ratio estimation experiments under composition constraints which are within 3 mol % of the D-optimal feed compositions. The second problem is the design of penultimate reactivity ratio estimation experiments. The D-optimal criterion for the penultimate model was derived and optimized for two existing systems as well as a large number of hypothetical systems. The D-optimal criterion for the penultimate model was found to be much more complicated than the terminal model criterion. For this reason, the optimal penultimate reactivity ratio estimation experiments can not be approximated by simple equations. Anyone interested in designing such experiments must derive and optimize the D-optimal criterion. © 1993 John Wiley & Sons, Inc.     

Determination of homogentisic acid in urine for diagnosis of alcaptonuria: Capillary electrophoretic method optimization using experimental design

Homogentisic acid (HGA) is a diagnostic metabolite that accumulates in the urine and tissues of patients with alkaptonuria which is a rare autosomal recessive disease. HGA is a specific metabolite in urine and serum, which is used for diagnosis of alkaptonuria. This study presents an inexpensive and easy capillary electrophoretic method for the quantitative determination of HGA in urine samples. The method was optimized using full factorial experimental design. The optimal separation electrolyte and separation voltage were revealed as 45 mmol/L phosphate buffer at pH 7.0 and 22 kV, respectively. Under these conditions the presence of HGA was detected in 6 min. Repeatability of migration times and corrected peak areas of HGA (as RSD) were 0.37 and 1.99, respectively. The detection limit was 0.56 μg/mL, 3 times of the average noise, and the quantification limit was 1.85 μg/mL, 10 times the average noise for HGA. Urine samples were directly injected to the capillary without any pretreatment step.     

Optimization of sylvite transformation into arcanite using experimental design methodology

This study is an application of the experimental design methodology for optimizing a potassium sulfate synthesis reaction. The latter is a two‐stage reaction through an intermediate product (Schoenite: K₂SO₄.MgSO₄·6H₂O). To determine optimal experimental conditions of the first stage, we have conducted a fractional factorial design and a central composite one. The optimal conditions of the second stage were determined only by means of a fractional factorial design. Several physico‐chemical techniques were used to implement this study, namely potentiometry, complexometry, gravimetry and X‐ray diffraction. This work has showed that this double decomposition reaction, when performed under the determined optimal conditions, gives good quality potassium sulfate (purity more than 95%) with a maximal yield. Copyright © 2008 John Wiley & Sons, Ltd.     

Identification of radicals spin-trapped in autoxidized linoleic acids: A high performance liquid chromatography and electron spin resonance study

Radicals generated in linoleic acid and deuterated linoleic acid have been trapped by the spin trap 2-methyl-2-nitroso-propane (MNP) and identified by electron spin resonance (ESR) and ENDOR spectroscopy. The formation of two distinct secondary alkyl radical adducts (one conjugated, the other non-conjugated, as shown by their UV absorption spectra) was demonstrated by 11, 11-dideuterio-9-cis-12-cis-linoleic acid using HPLC and ESR spectroscopy.     

亚油酸酰化胰岛素的制备及飞行质谱分析

胰岛素侧链氨基的酰化是胰岛素化学修饰较常用的方法。天然胰岛素分子中有三个裸露的氨基,即G lyA1、PheB1的α-氨基和LysB29的ε-氨基,都可作为胰岛素化学修饰的位点。酰化试剂通常经由N-羟基琥珀酰亚胺酯对胰岛素进行修饰。N-羟基琥珀酰亚胺酯可由羧酸与N-羟基琥珀酰亚胺(HOS     

Integrated quality by design (QbD) and design of experiments (DoE) approach for UFLC determination of telaprevir in rat serum

An ultrafast liquid chromatographic bioanalytical method was developed and validated for the determination of telaprevir in Wistar albino rat serum. Principles of quality by design (QbD) were implemented for enhancing the bioanalytical liquid–liquid extraction of telaprevir from rat serum. A Box–Behnken design was utilized in the studies by selecting extraction time, centrifugation speed, and vortex time as the critical method variables for evaluating their effect on the critical analytical attribute, i.e., %recovery of telaprevir. Chromatographic separation was achieved within a run time of 10?min using a C-18 column and mobile phase comprising of methanol:borate buffer of pH 9 (90:10 v/v) flowing at 1.2?mL/min. Photodiode array detection was performed at 270?nm. Results of validation studies were satisfactory. The method was linear over a concentration of 25–10,000?ng/mL. Limit of detection for the developed method was 10?ng/mL. Further, design of experiments (DoE) used for inter-day accuracy and precision study suggested superior method reliability. This integrated QbD- and DoE-based approach ensured the development of a validated and reliable analytical method for optimum bioanalysis of telaprevir in biological matrix.     

Preparation and characteristics of linoleic acid-grafted chitosan oligosaccharide micelles as a carrier for doxorubicin

The linoleic acid (LA)-grafted chitosan oligosaccharide (CSO) (CSO-LA) was synthesized in the presence of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC), and the effects of molecular weight of CSO and the charged amount of LA on the physicochemical properties of CSO-LA were investigated, such as CMC, graft ratio, size, zeta potential. The results showed that these chitosan derivatives were able to self-assemble and form spherical shape polymeric micelles with the size range of 150.7–213.9 nm and the zeta potential range of 57.9–79.9 mV, depending on molecular weight of CSO and the charged amount of LA. Using doxorubicin (DOX) as a model drug, the DOX-loaded CSO-LA micelles were prepared by dialysis method. The drug encapsulation efficiencies (EE) of DOX-loaded CSO-LA micelles were as high as about 75%. The sizes of DOX-loaded CSO-LA micelles with 20% charged DOX (relating the mass of CSO-LA) were near 200 nm, and the drug loading (DL) capacity could reach up to 15%. The in vitro release studies indicated that the drug release from the DOX-loaded CSO-LA micelles was reduced with increasing the graft ratio of CSO-LA, due to the enhanced hydrophobic interaction between hydrophobic drug and hydrophobic segments of CSO-LA. Moreover, the drug release rate from CSO-LA micelles was faster with the drug loading. These data suggested the possible utilization of the amphiphilic micellar chitosan derivatives as carriers for hydrophobic drugs for improving their delivery and release properties.     

Effect of conjugated linoleic acid grafting on the hemocompatibility of polyacrylonitrile membrane

Polyacrylonitrile (PAN) membrane was hydrolyzed with NaOH_(aq) and grafted with conjugated linoleic acid (CLA) via esterification with 1,3‐propanediol. The resulting CLA grafted PAN membranes were characterized using Fourier transform infrared spectrometry (FT‐IR) and X‐ray photoelectronic spectroscopy (XPS). The effects of CLA grafting on the blood coagulation, platelet aggregation, and oxidative stress were evaluated using human blood. The complete blood count (CBC) and coagulation time (CT) was evaluated in vitro for hemocompatibility. After CLA grafting, the proliferation of human umbilical vein endothelial cells (HUVECs) on the membranes were improved. In addition, the production of reactive oxygen species (ROS) was measured by the chemiluminescence (CL) method to evaluate the oxidative stress. The results showed that the CLA‐grafted PAN membrane could keep the CBC values more stable than unmodified PAN membrane. The CLA‐grafted PAN membranes also showed longer CT. CLA‐grafted PAN membrane could keep the CL counts of hydrogen peroxide and superoxide values more stable than unmodified PAN membrane. These results suggest that a CLA‐grafted PAN membrane could offer protection for patients against oxidative stress and would be helpful for reducing the dosage of anticoagulant during hemodialysis. Copyright © 2006 John Wiley & Sons, Ltd.