Mobile Phase Mixture Research Articles

Determination of carotenoid pigments in several tree leaves by reversed-phase high-performance liquid chromatography

A rapid and simple reversed-phase high-performance liquid chromatographic method has been used to analyse all carotenoid pigments from leaves of five different trees. The pigments were separated on an octadecylsilane radial compression column, using a mobile phase mixtures of acetonitrile, methanol-water and ethyl acetate in three isocratic steps. This method resolves all higher plant photosynthetic pigments, carotenoids and also chlorophylls in less than 15 min, while achieving a relatively good separation of trans-lutein—the major carotenoid—from its isomers zeaxanthin and cis-lutein.

Separation of o-phthalaldehyde—mercaptoethanol derivatives of amino acids from blood plasma on reversed-phase Nova-Pak C 18 cartridges

A separation of 25 o-phthalaldehyde—mercaptoethanol derivatives of primary amino acids in plasma prepared from human blood has been developed for Waters 10 cm × 0.8 cm I.D., 4-μm Nova-Pak C 18 Radial-Pak cartridges. A binary gradient system with solvent-switching capability for the A pump is required. Computer methodologies have been utilized to develop mobile phase mixtures of phosphate buffer (pH 6.9), water, methanol and tetrahydrofuran. Advantages of the method include simple sample preparation, fast turnover time (67 min including the pre-column Autotag derivatization procedure) and exceptional column durability (several hundred analyses).

Role of π-π interactions in reversed-phase liquid chromatography

The retention of various saturated and unsaturated solutes in reversed-phase liquid chromatography (RPLC) was investigated for various saturated and unsaturated packings in mobile packings in mobile phase mixtures of various compositions of methanol and acetonitrile. Non-aqueous eluents containing acetonitrile have a lower solvent strength than eluents containing methanol for saturated alkanes, alcohols and triglycerides, as expected from the solvophobic theory of retention. Very peculiar retention behaviours are observed when unsaturated solutes are analysed on aromatic stationary phases and/or when mobile phase containing acetonitrile is used with either an aromatic stationary phase or with unsaturated solutes or both. All these observations can be rationalized by taking into account the effect of π-π interactions between the π-electron systems of the solute, the stationary phase and the mobile phase. The influence of the residual silanols on silica-based RPLC supports on the retention of alcoholic solutes in pure acetonitrile is demonstrated. The influence of the concentration of a charge-transfer complexing agent (silver nitrate) in the mobile phase on the retention of unsaturated triglycerides was studied in a methanolic eluent and compared with that in a mobile phase containing acetonitrile.

Determination of Amphotericin-B in Human Plasma by Reversed-Phase High Performance Liquid Chromatography Using a Short Octyl Column

Abstract Amphotericin-B is a polyene antifungal antibiotic used for the treatment of severe systemic fungal infections. For effective treatment of urinary fungaria and the prevention of significant adverse-effects, monitoring the concentration of Amphotericin-B in biological samples of humans (ingesting the drug) is required. In this experiment, Amphotericin-B was isolated from plasma endogenous substances by adding 200 μL of acetonitrile in 800 μL of plasma. This mixture was vortex mixed, 20 mg of zinc sulfate and 10 mg of monobasic potassium phosphate was added to the mixture. This mixture was again vortex mixed and followed by centrifugation. The supernatant was filtered through a 0.45 μm membrane and a 100 μL aliquot of this solution was injected onto the chromatographic system. A short column of 60 mm × 4.6 mm packed with 3 μm octyl particles was used with an isocratic elution of 50/50, acetonitrile/0.01M KH2PO4 (v/v). The pH of the mobile phase mixture was adjusted to 3.5 with H3PO4. The intact drug...

Retention studies in supercritical fluid chromatography

The effect of fluid density on solute retention using a mobile phase containing 3% (v/v) propan-2-vol in pentane has been investigated. A linear relationship was established between solute retention times and mobile (fluid) phase density. A new method for the determination of the critical temperature of mobile phase mixtures is described and the chromatographic integrity of gas, supercritical fluid and high-performance liquid chromatography is demonstrated.

Determination of ascorbic acid, dehydroascorbic acid and ascorbic acid-2-phosphate in infiltrated apple and potato tissue by high-performance liquid chromatography

A high-performance liquid chromatography procedure for the determination of ascorbic acid-2-phosphate (AAP), ascorbic acid (AA), and dehydroascorbic acid (DHAA) in raw apple and potato, treated with AAP and AA to prevent browning, was developed. These compounds were extracted with a mixture of mobile phase and 2.5% metaphosphoric acid and separated on an aminopropyl bonded-phase silica column. DHAA was determined as AA following reduction with dithiothreitol. The method was evaluated with spiked samples and found to be accurate and reproducible at concentrations as high as 0.9 m M AA or AAP.

A method for the preparation of binary mobile phase mixtures for capillary supercritical fluid chromatography

Journal of High Resolution ChromatographyVolume 12, Issue 1 p. 51-52 Short Communication A method for the preparation of binary mobile phase mixtures for capillary supercritical fluid chromatography D. E. Raynie, D. E. Raynie Department of Chemistry, Brigham Young University, Provo, UT 84602, USASearch for more papers by this authorS. M. Fields, S. M. Fields Department of Chemistry, Brigham Young University, Provo, UT 84602, USASearch for more papers by this authorN. M. Djordevic, N. M. Djordevic Department of Chemistry, Brigham Young University, Provo, UT 84602, USASearch for more papers by this authorK. E. Markides, K. E. Markides Department of Chemistry, Brigham Young University, Provo, UT 84602, USASearch for more papers by this authorM. L. Lee, Corresponding Author M. L. Lee Department of Chemistry, Brigham Young University, Provo, UT 84602, USADepartment of Chemistry, Brigham Young University, Provo, UT 84602, USASearch for more papers by this author D. E. Raynie, D. E. Raynie Department of Chemistry, Brigham Young University, Provo, UT 84602, USASearch for more papers by this authorS. M. Fields, S. M. Fields Department of Chemistry, Brigham Young University, Provo, UT 84602, USASearch for more papers by this authorN. M. Djordevic, N. M. Djordevic Department of Chemistry, Brigham Young University, Provo, UT 84602, USASearch for more papers by this authorK. E. Markides, K. E. Markides Department of Chemistry, Brigham Young University, Provo, UT 84602, USASearch for more papers by this authorM. L. Lee, Corresponding Author M. L. Lee Department of Chemistry, Brigham Young University, Provo, UT 84602, USADepartment of Chemistry, Brigham Young University, Provo, UT 84602, USASearch for more papers by this author First published: January 1989 https://doi.org/10.1002/jhrc.1240120117Citations: 12AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat No abstract is available for this article.Citing Literature Volume12, Issue1January 1989Pages 51-52 RelatedInformation

Dimetridazole Residues in Pork Tissue. I. Assay by Liquid Chromatography with Electrochemical Detector

A liquid chromatographic (LC) method with electrochemical detection in the reductive mode was developed for the quantitative determination of dimetridazole (DMZ) and its major metabolite (HMMNI) at residue levels in pork tissue. For blood plasma, a sample is precipitated with 2 volumes of acetonitrile and centrifuged, and a diluted aliquot of the supernatant liquid is chromatographed. For muscle, a 10 g sample is extracted 3 times with dichloromethane. After evaporation of the combined extracts, the residue is redissolved in a mixture of hexane and mobile phase (0.3% TEA in 0.6M ammonium acetate pH 5.0 and acetonitrile, 85 + 15) and centrifuged, and an aliquot of the lower phase is chromatographed. Chromatography is accomplished using valve switching with 2 liquid circuits, employing the same mobile phase for both. The sample is deaerated by sparging with helium under slight positive pressure to prevent rediffusion of the oxygen. The sample is first loaded into a deoxygenator and the flow is stopped for complete deoxygenation. The flow is then resumed to transfer the sample into the first, low back-pressure column (ODS, 10 microns, 4.6 x 200 mm). Switching the valve at this point removes the deoxygenator from the circuit and connects the first column to a second one (ODS, 5 microns, 4.6 x 150 mm) in tandem. After the effluent is passed through a second deoxygenator to reduce the residual oxygen in the mobile phase, it is monitored by an electrochemical detector with a screened wall jet cell and a gold mercury electrode, set at -1.2 V.(ABSTRACT TRUNCATED AT 250 WORDS)

Use of unmodified silica with buffered aqueous mobile phase mixtures for selective chromatography of basic drugs

The characteristic selectivity of unmodified silica using mixed aqueous-methanolic mobile phase systems for the separation of basic drugs was studied. It was found that low buffer concentration-high methanol eluents show increased chromatographic performance over mobile phase systems lower in methanol and higher in ion strength. Regarding an assumed ion-exchanger retention mechanism, the capacity factors of an amino group containing drugs is primarily dependent on the concentration of the cation and the pH-value of the eluent. By increasing the pH-value of the eluent, elution order of primary and secondary to tertiary amines is reversed. By only using eluents with buffering capability, a very high stability and reproducibility of retention of the basic drugs could be observed. Comparing three different brands of unmodified silica materials, with different pore size and specific surface area, only small differences in the selectivity for the tested drugs could be found.

Synthesis, Identification, and Acute Toxicity of Some 7V-AlkyI Derivatives of 3,4-Methylenedioxyamphetamine

A series of N-alkyl derivatives of 3,4-methylenedioxyamphetamine (MDA) was prepared in an effort to characterize these potential drugs of abuse. These secondary amines were synthesized via reductive amination of the corresponding ketone with alkylamines. The ultraviolet absorption spectra for these compounds produced almost equally intense absorbance at 234 and 285 nm. The compounds were separated by liquid chromatography using reverse phase (C18) procedures with a ternary mobile phase mixture. Toxicity studies showed all of the compounds to have LD50 values similar to N-methyl MDA (MDMA).

Determination of Sorbic, Dehydroacetic, and Propionic Acids in Cheese by Liquid Chromatography and Gas Chromatography

Abstract A convenient and accurate method is presented for the determination of sorbic, dehydroacetic, and propionic acids in cheese by liquid chromatography and gas chromatography. The volatile acids are separated from cheese by steam distillation. Sorbic and dehydroacetic acids are determined by liquid chromatography using a reverse phase column and a mobile phase mixture (30 + 70) of methanol and 0.02M phosphate buffer, pH 7.2, containing 0.005M tetrabutyl ammonium. The effluent is monitored at 235 nm. After the distillate is acidified with formic acid, propionic acid is determined by gas chromatography with a flame ionization detector using a Porapak QS column. Average recoveries of sorbic, dehydroacetic, and propionic acids ranged from 92.3 to 99.5%. By this method, the acids could be determined without using organic solvent during extraction and cleanup.

Diastereospecific HPLC Analysis of the Antiulcer Drug, Enprostil, in a Soft Elastic Gelatin Capsule Formulation

Abstract Enprostil, an E-type prostaglandin analog, is formulated as a 0.3 mM propylene carbonate solution filled into soft elastic gelatin capsules. A characteristic structural feature of enprostil is the presence of two unresolved chiral centers, yielding a drug molecule that exists as two diastereomeric pairs of enantiomers. This report describes an HPLC method for quantitatively determining the relative amounts of enprostil diastereomers in soft elastic gelatin capsules. Using a Spherisorb ODS 5 μym, 250 × 4.5 mm column and a mobile phase mixture of 59:40:1 methanol:water:tetrahydrofuran separates the enprostil diastereomeric pairs and also resolves the enprostil diastereomers from potential interferences arising from placebo and drug degradation. The method performs well as evidenced by the usual statistical tests for: response linearity, recovery efficiency, precision, and lower quantitation limit. Also, the demonstrated dependence of chromatographic retention on mobile phase composition and system ...

Practical procedure for the optimization of reversed- phase separations with quaternary mobile phase mixtures

In this paper we described an extension of the previously published one-parameter ternary mobile phase optimization to include quaternary mobile phases. Whereas in ternary optimization only mixtures of two from the three isoeluotropic binary phases of water with methanol, tetrahydrofuran and acetonitrile are optimized, with quaternary optimization a blend of all three binary phases is searched for the best separation conditions. This two-parameters optimization formulates first a linear estimate of the retention based on flat planes, fitted through a minimum number of three initial runs. The predicted optimum is verified and an observed differences is used to refine the retention surfaces. After a few such iterations, the true shape is found and the procedure can be stopped. Practical examples were run to demonstrate the possibilities of the procedures.

A Modified, Sensitive Liquid Chromatographic Method for Measurement of Propranolol with Fluorescence Detection

A simple, sensitive, and rapid fluorometric-high-perform-ance liquid Chromatographic method for quantitating human and animal plasma and blood levels of propranolol has been developed. The procedure, which requires microliter volumes of triethylamine in the mobile phase mixture and microliter volumes of plasma or blood (50–100 μL), may be applied in routine clinical analysis as well as in pharmacokinetic studies using small laboratory animals. A single, selective extraction step precedes the analysis. The limiting concentration that can be detected is 1ng/mL.

Stability Indicating high Performance Liquid Chromatographic Procedure for the Analysis of Prostaglandin E2Raw Material and Tablets

Abstract A procedure is described for the quantitative analysis of prostaglandin E2, its isomers and degradation products. The high performance liquid chromatographic method uses a chloroform/hexane mobile phase (70/30 v/v) mixture and a 250 × 4.6 mm, 5 micron cyano column, with testosterone as the internal standard. The time required for chromatography is approximately 15 minutes. The method gives a R.S.D. of 1.7% for the assay of the drug in tablets.

A simple procedure for the rapid optimization of reversed-phase separations with ternary mobile phase mixtures

A simple rapid procedure is described for estimating optimum compositions of ternary mobile phase mixtures for the separation of samples by reversed-phase liquid chromatography (RPLC). Retention data in two iso-eluotropic binary mobile phase mixtures (mixtures with equal retention times) are required to initiate the procedure. The logarithm of the capacity factor is assumed to vary linearly with the composition of isoeluotropic ternary mixtures formed by mixing the two limiting binaries. Using the product of resolution factors of adjacent peaks as the criterion, an optimum ternary composition is then calculated. After a chromatogram has been obtained with the predicted optimum ternary mobile phase, the procedure is repeated until no further improvement can be achieved. Examples of the application of the present procedure are described to illustrate its effectiveness.

High-Pressure Liquid Chromatographic Separation of Diphenhydramine and Some of its Metabolites: Effects of Eluent Salt Concentration on Chromatographic Characteristics

Abstract With appropriate variation of the pH, alkali halogenide concentration and cation and/or anion species in the mobile phase (mixtures of water and methanol), it was possible to resolve diphenhydramine, N-desmethyldiphenhydramine, N,N-didesmethyldiphenhydramine, benzhydrol, benzophenone and 2-methyldiphenylmethoxy acetic acid on a reversed-phase column. The retention volumes of the three amines decreased by increasing the following parameters: salt concentration, methanol concentration and cation and/or anion size. The retention volumes of the other compounds were not influenced. The good separation and the simplicity make this method attractive for use in metabolism studies.

Development and Validation of RP-HPLC method for determination of Modafinil in bulk and dosage form(3ò¬

A reverse phase high performance liquid chromatographic method was developed for the determination of Modafinil in bulk and dosage form. The separation was effective on a Hypersil ODS C18 column (250 mm x 4.6 mm; 5μ) using a mobile phase mixture of Buffer:Acetonitrile in a ratio of 55:45 (v/v) at a flow rate of 1.0ml/min. The detection was made at 220nm. The retention time of modafinil was found to be 4.80±0.06 min. Calibration curve was linear over the concentration range of 20-120μg/ml of modafinil. The proposed method was validated as per the ICH guidelines. The method was accurate, precise, specific and rapid and thus found to be suitable for the quantitative analysis of modafi-nil in the bulk and dosage form.Key Words: Method development; validation; Modafinil; Tablets; Hypersil C18 Column; RP-HPLC.DOI: http://dx.doi.org/10.3329/icpj.v1i4.10062International Current Pharmaceutical Journal 2012, 1(4): 77-80