Cyanopropyl Column Research Articles

Fundamental investigation of impact of water and TFA additions in peptide sub/supercritical fluid separations

The retention of three peptides was studied under analytical and overloaded conditions at different concentrations of trifluoroacetic acid (TFA) and water added to the co-solvent methanol (MeOH). Four columns with different stationary phase properties, i.e., silica, diol, 2-ethylpyridine and cyanopropyl (CN) columns, were evaluated in this investigation. The overall aim was to get a deeper understanding on how column chemistry as well as water and TFA in the co-solvent affect the analytical and overloaded elution profiles using multivariate design of experiments and adsorption measurements of co-solvent components. Multivariate experimental design modeling indicated that water had on average around five times higher effect on the retention than the addition of TFA. The results also showed that the retention increases with the addition of TFA and water to the co-solvent on all columns except the CN column, on which the retention decreased. When examining the effect of adding water to the co-solvent, evidence of a hydrophilic interaction liquid chromatography (HILIC)-like retention mechanism was found on the three other columns with more polar stationary phases. However, on the CN column water acted as an additive, decreasing the retention due to competition with the peptide for available adsorption surface. Adsorption isotherm measurements of the polar solvent MeOH showed that MeOH adsorbs much weaker on the CN column than on the other columns. Addition of TFA and water to the co-solvent substantially sharpened the elution profiles under both overloaded and analytical conditions. Adding a small amount of TFA (from 0 % to 0.05 %) to the co-solvent substantially improved the peak shape of the elution profiles, while further addition (from 0.05 % to 0.15 %) had only a minor effect on the elution profile shape. The reduced retention on the CN column could not be explained by TFA adsorption, which was very weak on all studied columns (retention factor, 0.05–0.15). One could therefore speculate that the ion-pairing complex formed between the peptide and TFA in the mobile phase, reduce the retention due to its reduced polarity. On the other columns displaying HILIC-like properties, the TFA probably just decreased the pH of the mobile phase, thereby promoting the partitioning of the peptide into the water-rich layer. Finally, peak deformation due to diluent–eluent mismatch was observed under overloaded conditions. This was most severe in the cases where MeOH adsorption to the stationary phase was strong and the peptides were only mildly retained. Adding 1,4-dioxan to the diluent resolved this issue.

Comparison of supercritical fluid chromatographic methods to predict the skin permeability of pharmaceutical and cosmetic compounds

Supercritical fluid chromatography (SFC) was explored as an alternative for liquid chromatography to predict the skin permeability of pharmaceutical and cosmetic compounds. Nine dissimilar stationary phases were applied to screen a test set of 58 compounds. The experimental retention factors (log k), in addition to two sets of theoretical molecular descriptors, were applied to model the skin permeability coefficient. Different modelling approaches, i.e. multiple linear regression (MLR) and partial least squares (PLS) regression, were used. In general, the MLR models performed better than the PLS models for a given descriptor set. The results obtained on a cyanopropyl (CN) column provided the best correlation with the skin permeability data. The retention factors obtained on this column were included in a simple MLR model, together with the octanol-water partition coefficient and the number of atoms (r² = 0.81, RMSEC = 0.537 or 20.5% and RMSECV = 0.580 or 22.1%). The overall best MLR model included the chromatographic descriptor from a phenyl column and 18 descriptors (r² = 0.98, RMSEC = 0.167 or 6.2% and RMSECV = 0.238 or 8.9%). This model showed a good fit, on top of very good predictive features. However, stepwise MLR models with a reduced complexity could also be determined, with the best performance parameters obtained with the CN-column based retention and eight descriptors (r² = 0.95, RMSEC = 0.282 or 10.7% and RMSECV = 0.353 or 13.4%). SFC thus provides a suitable alternative to the liquid chromatographic techniques previously applied to model the skin permeability.

Supercritical fluid chromatography with post-column addition of supporting electrolyte solution for electrochemical determination of tocopherol and tocotrienol isomers.

A supercritical fluid chromatography with electrochemical detection system was developed for the simultaneous determination of tocopherol and tocotrienol isomers. The supercritical fluid chromatography with electrochemical detection system was connected with an additional pump to create a flow path to add a supporting electrolyte solution. The supporting electrolyte solution was mixed with a mobile phase in a post-column fashion, enabling the independent control of the separation and detection. After optimization of the measurement conditions, vitamin E isomers and an internal standard substance (2,2,5,7,8-pentamethyl-6-hydroxychroman) were separated within 30min using a mixture of supercritical carbon dioxide and methanol (99:1, v/v) as a mobile phase and a cyanopropyl column (4.6mm inner diameter × 250mm length, 5 μm). For the electrochemical detection, methanol containing 1.0mol/L ammonium acetate was used as a supporting electrolyte solution, and the applied potential was set at +0.8 V. This analytical method showed good linearity (5-100 μg/mL) and repeatability (less than 2.5% relative standard deviation, n=6) and was applicable to the determination of tocopherol and tocotrienol isomers in nutrition supplements.

A Validated RP-HPLC Method for the Determination of Butamirate Citrate and Benzoic Acid in Syrup, Based on an Experimental Design Assessment of Robustness

A reversed-phase high-pressure liquid chromatography (RP-HPLC) method was developed and subsequently validated for the simultaneous determination of butamirate citrate (BC) and benzoic acid (BA) in cough syrup. The separation was performed employing a cyanopropyl column with a mobile phase consisting of 50%/50% v/v MeOH/NaH2PO4 * H2O 50 mM aqueous solution pH = 3.0. The quantitation was achieved with a diode array detector (DAD) at 210 nm. The method demonstrated a congenitally satisfactory separation, yet the acquired peaks were asymmetrical. This effect was eliminated by using 1% triethylamine in the buffer solution as a silanol blocker. In addition, the method was found to unequivocally assess the target analytes in the sample matrix and fulfilled the required specifications in relevance to specificity, linearity, accuracy, precision and stability of both the standard solutions and of the sample solutions. Lastly, an experimental design was designed in order to assess the robustness of the proposed assay. To this purpose, a graphical and a statistical approach were utilized and compared to identify the factors that should be strictly controlled during each execution of the method.

Concurrent chromatographic determination of pseudoephedrine and loratadine from combination syrups and tablets

BackgroundChromatographic separation of polar and nonpolar compounds when presented in combined dosage forms has always been considered as great analytical challenge. Separation and retention of both polar and nonpolar compounds by the same stationary phase can be a useful approach for analyses of complex samples with such a difference in chemical properties. Loratadine (nonpolar) and pseudoephedrine (polar) are typical examples of this situation.ResultsThe Box–Behnken design was used to optimize the separation process, an efficient separation of loratadine and pseudoephedrine was achieved within 6 min; employing a mixture of 16.0 mM ammonium acetate buffer (pH 4.5) and acetonitrile (23:77, v/v) as isocratic mobile phase, pumped at 1.0 mL/min through a Zorbax cyanopropyl column (250 mm × 4.6 mm, 5 μm), the analytes were detected at 250 nm. Under the same conditions, separation of sodium benzoate preservative co-formulated with the two analytes in syrup formulation was also achieved. The calibration curve demonstrated excellent linearity in the range of 24.6–123.2 μg/mL and 594.8–2974.0 μg/mL for loratadine and pseudoephedrine, respectively with determination coefficient (r2) > 0.999.ConclusionThe method’s accuracy bias < 2.0%, repeatability and intermediate precision (%RSD < 2.0%) were verified. In addition, system suitability parameters were found within the acceptable limits. Satisfactory results were obtained upon the application of the validated method to the analysis of commercial tablet and syrup formulations.

Liquid chromatography–electrospray ionization-tandem mass spectrometry for the determination of short-chain chlorinated paraffins in mixed plastic wastes

Wastes containing short-chain chlorinated paraffins (SCCPs) at concentrations above the Basel Convention low persistent organic pollutant content (LPC) values must be destroyed or irreversibly transformed in an environmentally-sound manner. Here, we developed a novel liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MSMS) method for determining the concentrations of SCCPs in mixed plastic wastes. Major SCCP homologues were identified with good separation and peak width by using a low-hydrophobicity cyano-propyl column and a mobile phase consisting of water and methanol containing ammonium acetate. Precursor ion peaks corresponding to the formation of stable SCCP homologue-adducts were observed, followed by two intense product ion peaks corresponding to separation of the adduct into acetate and the homologue. The formulation of a novel calibration standard with known SCCP homologue percentage composition supported the development of our novel mass spectrometric technique. The results obtained with the LC-ESI-MSMS system were quantitatively and qualitatively comparable with those obtained with a high-resolution mass spectrometry (HRMS) coupled to gas chromatography (GC) system. Homologue concentrations determined by LC-ESI-MSMS were significantly correlated with those determined by GC-HRMS in samples of commercial chlorinated paraffin mixture and mixed plastic waste, respectively. As a complementary technique to the highly accurate, but less versatile GC-HRMS approaches, the SCCP analysis by LC-ESI-MSMS is a practical way to identify mixed plastic wastes containing SCCPs at concentrations higher than the Basel Convention's LPC value.

A Dedicated Mass Spectrometer Tuning Method for SFC–MS and Evaluation of Two Different Linear Restrictor Types Used in the Operation of a Split Flow Pre-Back Pressure Regulator SFC–MS Interface

The operation of a SFC–MS system has been improved. The SFC–MS interface uses a post-column make-up flow of liquid whose supply line to the SFC–MS interface mixing tee is equipped with an injection valve. Prior to analysis, this arrangement permits a stable flow of a dilute tune solution of analyte(s) for the exact amount of time required to optimize mass spectrometry tuning. Advantageously, with this arrangement tuning is performed in the presence of the exact split flow and same composition of SFC mobile phase and make-up liquid to be used for subsequent SFC–MS operation. The SFC–MS interface was constructed to provide a pre-back pressure regulator split flow to the mass spectrometer using a linear restrictor. Two different linear restrictors constructed from either 50 µm i.d. PEEK tubing or 25 µm i.d. fused silica tubing were evaluated. For best electrospray performance, each restrictor required a different post-column make-up flow rate. The use of either restrictor retained a high degree of chromatographic fidelity for the SFC separation of six pharmaceutical compounds using a cyanopropyl column with a mobile phase of supercritical fluid carbon dioxide modified with methanol and aqueous ammonia solution. Each restrictor provided a different level of SFC–MS performance for each of the six pharmaceutical compounds tested. Based upon overall performance, a linear fused silica restrictor was selected to develop a chromatographic separation using on-line SFC–MS for three structurally very similar monensin sodium salt analogues that are not directly amenable to off-line SFC using UV/Vis detection.

DETERMINATION OF ARGININE AND ASCORBIC ACID IN EFFERVESCENT TABLETS BY MULTIDIMENSIONAL CHROMATOGRAPHY

This paper describes a fast and direct method for quantification of arginine and ascorbic acid (vitamin C) in effervescent tablets. HPLC-UV multidimensional chromatography (RP18 and cyanopropyl columns) was developed and validated. Phosphate buffer 10 mMpH 3.4 was used as mobile phase. Satisfactory resolution between the drugs was obtained with analysis time less than 8.0 min. Method was linear in the ranges of 140-320 mg.mL-1 and 240-560 mg.mL-1 for arginine and ascorbic acid, respectively. Precision showed RSD &lt;2.0%. Recovery was 99.5% and 100.0% for arginine and ascorbic acid, respectively. Robustness was confirmed through factorial analysis 22 (pH and mobile phase flow rate as factors).

An improved method for fast and selective separation of carotenoids by LC-MS.

Carotenoids are a large class of compounds that are biosynthesized by condensation of isoprene units in plants, fungi, bacteria, and some animals. They are characteristically highly conjugated through double bonds, which lead to many isomers as well susceptibility to oxidation and other chemical modifications. Carotenoids are important because of their potent antioxidant activity and are the pigments responsible for color in a wide variety of foods. Human consumption is correlated to many health benefits including prevention of cancer, cardiovascular disease, and age-related disease. Extreme hydrophobicity, poor stability, and low concentration in biological samples make these compounds difficult to analyze and difficult to develop analytical methods for aimed towards identification and quantification. Examples in the literature frequently report the use of exotic stationary phases, solvents, and additives, such as ethyl acetate, dichloromethane, and methyl tert-butyl ether that are incompatible with liquid chromatography mass spectrometry (LC-MS). In order to address these issues, we implemented the use of LC-MS friendly conditions using a low-hydrophobicity cyano-propyl column (Agilent Zorbax SB-CN). We successfully differentiated between isomeric carotenoids by optimizing two gradient methods and using a mixture of 11 standards and LC-MS in positive ionization mode. Three complex biological samples from strawberry leaf, chicken feed supplement, and the photosynthetic bacterium Chloroflexus aurantiacus were analyzed and several carotenoids were resolved in these diverse backgrounds. Our results show this methodology is a significant improvement over other alternatives for analyzing carotenoids because of its ease of use, rapid analysis time, high selectivity, and, most importantly, its compatibility with typical LC-MS conditions.

A newly validated high-performance liquid chromatography method with diode array ultraviolet detection for analysis of the antimalarial drug primaquine in the blood plasma.

Primaquine (PQ) diphosphate is an 8-aminoquinoline antimalarial drug with unique therapeutic properties. It is the only drug that prevents relapses of Plasmodium vivax or Plasmodium ovale infections. In this study, a fast, sensitive, cost-effective, and robust method for the extraction and high-performance liquid chromatography with diode array ultraviolet detection (HPLC-DAD-UV ) analysis of PQ in the blood plasma was developed and validated. After plasma protein precipitation, PQ was obtained by liquid-liquid extraction and analyzed by HPLC-DAD-UV with a modified-silica cyanopropyl column (250mm × 4.6mm i.d. × 5μm) as the stationary phase and a mixture of acetonitrile and 10mM ammonium acetate buffer (pH = 3.80) (45:55) as the mobile phase. The flow rate was 1.0mL·min-1, the oven temperature was 50OC, and absorbance was measured at 264nm. The method was validated for linearity, intra-day and inter-day precision, accuracy, recovery, and robustness. The detection (LOD) and quantification (LOQ) limits were 1.0 and 3.5ng·mL-1, respectively. The method was used to analyze the plasma of female DBA-2 mice treated with 20mg.kg-1 (oral) PQ diphosphate. By combining a simple, low-cost extraction procedure with a sensitive, precise, accurate, and robust method, it was possible to analyze PQ in small volumes of plasma. The new method presents lower LOD and LOQ limits and requires a shorter analysis time and smaller plasma volumes than those of previously reported HPLC methods with DAD-UV detection. The new validated method is suitable for kinetic studies of PQ in small rodents, including mouse models for the study of malaria.

Development and Validation of a New Method to Quantify Pilocarpine in Tablets by HPLC-DAD

Introduction: Pilocarpine, a natural alkaloid that can be obtained from species of Pilocarpus, has been used to treat glaucoma, ocular hypertension and xerostomia. The development and validation methods for analysis of pilocarpine and its degradation products are quite challenging since the physicochemical properties between them are relative small. Methods: Analytical tests in HPLC-DAD were performed and 13 different conditions were take into acount, employing three columns (silica based C18, modified-silica cyanopropyl and Zic-HILIC), two buffer solutions, two organic solvents, pH ranging from 5.1 to 5.45, three temperatures and flow rate from 0.8 to 2.0 mL.min-1, in RP or HILIC. The chromatographic assays for pilocarpine hydrochloride in aqueous solution allowed to establish the best chromatographic parameters in terms of signal symmetry and retention as well as to separate pilocarpine from its degradation products with great selectivity, efficiency and resolution. It was possible to validate a new method to dosage pilocarpine hydrochloride in tablets using a modified-silica cyanopropyl column, mobile phase in isocratic mode with mixture of acetonitrile/ potassium dihydrogen phosphate buffer 10 mM (60:40, v/v), pH 5.30, oven temperature of 50 oC and flow rate of 1.0 mL.min-1. Conclusion: The precision, accuracy, linearity, robustness, recovery, short retention time, isocratic elution mode, quantification (250 ng.mL-1) and identification (75 ng.mL-1) limits allowed establishing a routine estimation for pilocarpine in tablets. The pilocarpine hydrochloride content it was in the range as specified by the Salagen® producer (4.81 ± 0.07 mg/ tablet, 96.16 ± 1.33%) and by the formulation in carbomer 940® + chitosan tablets (4.80 ± 0.02 mg/ tablet, 95.99 ± 0.49%) and in sodium carboxymethylcellulose + chitosan tablets (4.95 ± 0.04 mg/ tablet, 99.08 ± 0.79%). Keywords: Pilocarpine, pharmaceutical analysis, tablets, HPLC-DAD, cyanopropyl column.

Retention, separation selectivity and system efficiency of selected basic psychotropic drugs on different RPLC columns

Abstract Retention parameters of psychotropic drug standards were determined on different columns, i.e., Octadecyl silica, Phenyl, Phenyl-Hexyl, Polar Reverse Phase, Pentafluorophenyl, and Cyanopropyl using aqueous eluent systems containing methanol or acetonitrile as organic modifiers, acetate buffer at pH 3.5 and addition of silanol blocker − diethylamine (DEA). The retention, separation selectivity, and sequence of elution were different when using eluents containing various organic modifiers. The significant differences were observed in retention parameters with a change of the used stationary phase. The various properties of stationary phases resulted in differences in analyte retention, peaks shape, systems efficiency and separation selectivity. The best shape of peaks were on Cyanopropyl (CN) column and the highest efficiency for most investigated psychotropic drugs were obtained on Phenyl-Hexyl and Polar RP columns.

Separation and identification of lipid classes by normal phase LC‐ESI/MS/MS on a cyanopropyl column

In order to establish a versatile and convenient method for the analysis of lipids, electrospray ionization tandem mass spectrometry (ESI‐MS/MS) was applied to a HPLC separation on a cyanopropyl‐bonded stationary phase. A binary gradient mobile phase system consisting of hexane, toluene, methanol and a stable electrospray yielding sodium adduct ions could be used to generate specific product ions in MS/MS mode. By applying the LC/ESI‐MS/MS method on an egg yolk sample, 29 different molecular species of phosphatidylethanolamines, phosphatidylcholines, and lysophosphatidylcholines could be detected within 25 min.

Application of LC and HPTLC-densitometry for the simultaneous determination of two multicomponent mixtures containing pyridoxine hydrochloride and cyanocobalamine

Summary Two methods were presented for the simultaneous determination of two multicomponent mixtures containing pyridoxine hydrochloride (B6) and cyanocobalamine (B12) with benfotiamine (BN) [mix. 1], or thiamine nitrate (B1) and diclofenac sodium (DC) [mix. 2]. The first high-performance liquid chromatographic (HPLC) method depends on the use of a cyanopropyl column at a flow rate of 1.5 mL min−1 with a mobile phase consisting of acetonitrile-25 mM ammonium acetate, pH 3.7 (10:90, v/v), for mix. 1 or acetonitrile-25 mM ammonium acetate, pH 5.0 by gradient elution, changing the proportion of the system linearly with a time schedule program, for mix. 2. Quantitation was achieved by UV detection at 220 nm. The second method was based on HPTLC separation followed by densitometric measurement of the spots at 254 nm. The separation was achieved on HPTLC silica gel F254 plates using chloroform-ethanol-water-acetic acid (5:8:2:0.5, v/v/v/v) for mix. 1 and chloroform-ethanol-water-acetic acid (1.5:8:2:0.5, v/v/v/...

Comparison of GC stationary phases for the separation of fatty acid methyl esters in biodiesel fuels

The fatty acid methyl ester (FAME) content of biodiesel fuels has traditionally been determined using gas chromatography with a polar stationary phase. In this study, a direct comparison of the separation of FAMEs present in various biodiesel samples on three polar stationary phases and one moderately polar stationary phase (with comparable column dimensions) was performed. Retention on each column was based on solubility in and polarity of the phase. Quantitative metrics describing the resolution of important FAME pairs indicate high resolution on all polar columns, yet the best resolution, particularly of geometric isomers, is achieved on the cyanopropyl column. In addition, the separation of four C18 monounsaturated isomers was optimized and the elution order determined on each column. FAME composition of various biodiesel fuel types was determined on each column to illustrate (1) chemical differences in biodiesels produced from different feedstocks and (2) chemical similarities in biodiesels of the same feedstock type produced in different locations and harvest seasons.

Separation of &lt;i&gt;cis/trans&lt;/i&gt; fatty acid isomers on gas chromatography compared to the Ag-TLC method

The present study investigates the separation of the cis/trans isomers of fatty acids on the 75 m SP2560 column under different gas chromatographic (GC) conditions including an isothermal program and a time-temperature program. The time-temperature program showed improved separation of cis/trans isomers of C 14:1 , C 16:1 , C 18:1 , C 18:2 and C 18:3 fatty acids along with short chain fatty acids compared to the isothermal program. The separation of trans/trans isomers of C 18:1 fatty acids including elaidic acid (C 18:1 ∆9t) and vaccenic acid (C 18:1 ∆11t) was difficult with the time-temperature program. The thin layer chromatography impregnated with silver nitrate (Ag- TLC) method was performed to separate cis/trans fractions and GC analysis was carried out with the trans fraction. But GC analysis showed a co-elution of trans isomers of C 18:1 fatty acid. Thus the study shows that a time-temperature programmed GC method with the highly polar cyanopropyl column is sufficient to resolve trans fatty acids along with short chain fatty acids when a large number of samples has to be analyzed.

SIMULTANEOUS DETERMINATION OF TWO MULTICOMPONENT MIXTURES CONTAINING PHENOBARBITONE AND EPHEDRINE HYDROCHLORIDE USING HPLC AND CHEMOMETRIC ASSISTED SPECTROPHOTOMETRIC METHODS

Three methods for the simultaneous determination of two multicomponent mixtures containing phenobarbitone and ephedrine hydrochloride with theophylline, and postafene hydrochloride [mix 1]; or dihydroxypropyltheophylline and chlorpheniramine maleate [mix 2] were proposed. The HPLC method depended on the use of gradient mobile phase system consisting of (A) acetonitrile and (B)12 mM ammonium acetate; pH 4.5 with cyanopropyl column at a flow rate of 1.5 mL min−1. Quantitation was achieved with UV detection at 215 nm, based on peak area. The two other methods were multivariate spectrophotometric calibration methods: partial least squares (PLS-1) and principal component regression (PCR). These approaches were successfully applied to quantify each component in the mixture. Analytical figures of merit (FOM) were calculated for both PLS-1 and PCR. The proposed methods were successfully applied for the analysis of pharmaceutical formulations containing the two multicomponent combinations and the results were compared.

Separation of Asphaltenes by Reversed-Phase Liquid Chromatography with Fraction Characterization

The use of a 4.6 × 250 mm, 5 μm cyanopropyl column is effective for the liquid chromatography (LC) separation of asphaltenes with sequential ultraviolet (UV) and florescence detection. The mobile-phase composition is an optimized gradient from acetonitrile (MeCN) and water to N-methyl-2-pyrrolidone (NMP) and tetrahydrofuran (THF). A low flow rate of 0.5 mL min–1 is used to maintain lower operating pressure to minimize aggregate formation. Using a 0.02 g L–1 asphaltene sample for preliminary optimization, three peaks, with two partially resolved, are evident in the fluorescence chromatogram. The UV chromatogram revealed an extra weakly retained peak, suggesting aggregates that quench fluorescence. Aggregation of asphaltenes increases with time up to about 10 h and is dependent upon the choice of sample solvent. On the basis of the reversed-phase mobile-phase gradient, the relative polarity of the peaks from least to most retained can be estimated over the polarity index (P′) range from about 6.3–4.3 on a scale of 0.1 for hexane (least polar) to 10.6 for water (most polar). The sample concentration is increased to 1 g L–1 for separation and collection of 12 fractions. Selected fractions are subjected to characterization using atmospheric pressure chemical ionization mass spectrometry (APCI–MS) using a linear quadrupole ion trap (LQIT). The variation of the molecular-weight distribution of the asphaltenes for the 12 fractions is fairly constant, indicating that the retention mechanism is not controlled by size exclusion but likely a partitioning/adsorption mechanism.

Analysis of vanilla extract by reversed phase liquid chromatography using water rich mobile phases

Vanillin is the major constituent of vanilla extract, a flavoring ingredient used in food and beverages. Natural vanilla extract prepared from the bean of the tropical orchid, Vanilla planifolia, is expensive due to the limited supply of the vanilla bean. For this reason, synthetic vanilla extracts are widely used. Synthetic vanilla extracts are less complex and usually contain vanillin, ethyl vanillin, and other related compounds that are prepared from inexpensive starting materials. Several liquid chromatographic methods have been developed to quantitate coumarin, vanillin, and ethyl vanillin in vanilla extract. The use of water rich mobile phases in reversed phase liquid chromatography (RPLC), e.g., 1% butanol in water with 0.2% acetic acid with C18, C8, and cyanopropyl columns, has been investigated as a potential method to characterize the composition of synthetic vanilla extracts. Better resolution is achieved in the separation of vanillin compounds when hydrophobic alcohols are used as organic modifiers. This can be attributed to butanol partitioning into the bonded phase, which provides a more extended ordered surface increasing the contact surface area of the stationary phase and thereby increasing the selectivity of the separation. Using water rich mobile phases, constituents of vanilla extract in 36 commercial products obtained from stores in the local area were identified demonstrating the efficacy of the proposed RPLC method.

Separation of Lipid Classes by HPLC on a Cyanopropyl Column

A new method for the separation and identification of lipid classes by normal-phase HPLC on a cyanopropyl column is described. The use of a simple binary gradient, with toluene as a component, provided a rapid separation of non-polar as well as phospholipid classes. The inherent small differences in performances between possible non-polar eluent components of the gradient, such as hexane, heptane, and iso-octane, had a pronounced impact on retention times for individual phospholipid classes. Separation of molecular species within a lipid class could also be observed.