Ultra-performance Liquid Chromatography Column Research Articles

Development and Validation of an Ultra-Performance Liquid Chromatography Method for the Determination of Wedelolactone in Rat Plasma and its Application in a Pharmacokinetic Study.

Wedelolactone is a coumarin ether with significant hepatoprotective effects. However, there are few pharmacokinetic studies of wedelolactone, which will affect the studies of its efficacy and potential toxicity. In this study, a selective ultra-performance liquid chromatography (UPLC) method was developed to confirm the pharmacokinetic parameters of wedelolactone in rat plasma. The chromatographic separation was carried out on a Kromasil C18 UPLC column (250 × 4.6 mm; 5.0 μm) by gradient mobile phase of methanol-water containing 0.5% acetic acid (v/v). Perfect linearity was obtained and the samples were stable under different conditions. The intra-day and inter-day precisions (relative standard deviation, %) were within 3.81% and accuracies (relative error, %) ranged from −4.01% to 7.12%. The extraction recoveries in rat plasma ranged from 95.98% to 108.93%. This rapid method was successfully applied in the pharmacokinetic study of wedelolactone in rat plasma. Following the oral administration of 5.00 mg/kg wedelolactone, the wedelolactone was rapidly absorbed. Pharmacokinetic parameters were used to quantitatively describe the dynamic changes of wedelolactone in vivo, providing a theoretical basis for pharmacological research on drugs and preclinical medication. The study of wedelolactone can provide a theoretical basis and quick analysis for the study of other traditional Chinese medicine. This may lead to breakthroughs in the pharmacokinetic study of complex Chinese medicines.

Separation of hexabromocyclododecane diastereomers: Application of C18 and phenyl-hexyl ultra-performance liquid chromatography columns

This study was performed to establish the proper liquid chromatographic conditions for the separation of hexabromocyclododecane (HBCD) diastereomers. Column selectivity towards HBCD diastereomers was evaluated for C18 and phenyl-hexyl stationary phases. First, the baseline separation of the primary HBCDs (α-, β-, and γ-HBCD) was obtained using the ultra-performance liquid chromatography (UPLC) column with C18 stationary phase chosen in most previous studies for HBCD analysis; however, co-elution of δ- and ε-HBCD with the primary HBCD diastereomers was observed. To prevent the interference from δ- and ε-HBCD, we adopted a phenyl-hexyl UPLC column to resolve the HBCD diastereomers. The phenyl-hexyl UPLC column showed significantly different selectivity for the HBCD diastereomers compared with the C18 column, which allowed the clear isolation of δ-HBCD and ε-HBCD from the primary HBCD diastereomers. In addition, by checking the retention times of all HBCD diastereomers using both C18 and phenyl-hexyl columns, we confirmed the presence of δ-, ε-, η-, and θ-HBCDs in two technical HBCD mixtures.

Column performance study of different variants of liquid chromatographic technique: an application on pharmaceutical ternary mixtures containing tetryzoline.

High-performance liquid chromatography (HPLC), ultra-performance liquid chromatography (UPLC) and rapid resolution liquid chromatographic (RRLC) methods have been developed and validated for the separation and quantitation of both or either of two ternary mixtures present in ophthalmic solutions. The first mixture contains chloramphenicol, dexamethasone sodium phosphate and tetryzoline HCl (TZH); while the second one contains ofloxacin, prednisolone acetate and TZH. Both preparations contain benzalkonium chloride as a preservative. The columns used were a HPLC column (C18 5 µm particle size), a RRLC column (C18 2.6 µm particle size) and a UPLC column (C18 1.7 µm particle size). A comparative study was conducted to illustrate the effect of the change in column particle size and dimensions on the other chromatographic conditions, backpressure and the separation of both ternary mixtures. The methods were validated as per ICH guidelines where accuracy, repeatability, interday precision and robustness were found to be within the acceptable limits. The RRLC column provided shorter run time and better resolution than HPLC, while the UPLC column gave the shortest run time for all columns. The RRLC column resulted in minimum backpressure, so it could be used with any HPLC instrument, which makes the method more practical and economic. The results obtained from the proposed methods were statistically compared with official ones where no significant difference was observed.

Development and validation of stability-indicating UPLC method for the determination of lafutidine and its impurities in bulk and pharmaceutical dosage form

Abstract Background A simple and rapid stability-indicating, reversed phase ultra-performance liquid chromatography (UPLC) method was developed for the quantitative determination of lafutidine and its four potential impurities. Results Separation was achieved on Acquity BEH-shield RP18 UPLC column (3.0 mm × 100 mm, 1.7 μm) under the gradient mode of elution by using mobile phase A (0.02M diammonium hydrogen phosphate/acetonitrile, 80:20 v/v) and mobile phase B (0.02M diammonium hydrogen phosphate/acetonitrile, 30:70 v/v). The flow rate was maintained at 0.5 mL min−1. UV detection was carried out at 276 nm. Conclusions Stability-indicating capability of the developed method is established by analyzing forced degradation samples in which the spectral purity of lafutidine is ascertained along with the separation of degradation products from analyte peak. The developed UPLC method is validated as per International Conference on Harmonization guidelines with respect to system suitability, specificity, precision, sensitivity, accuracy, linearity, and robustness.

Determination of Flunarazine Dihydrchloride in Bulk Drug and Tablets by RP-UPLC: A Stability-Indicating Assay

With the objective of reducing analysis time and maintaining good efficiency, there has been substantial focus on high-speed chromatographic separations. In this work, the development and validation of a rapid, precise and specific stability-indicating ultra performance liquid chromatography (UPLC) method for the determination of flunarizine dihydrochloride (FNH) in bulk drug and tablets is described. The chromatographic separation of the drug was achieved on a Zorbax extend C18, 50 × 4.6 mm, 1.8 μm UPLC column within a short runtime of 5.0 min with mobile phase consisting of a mixture of triethylamine and formic acid buffer of pH 2.5 and acetonitrile (50:50 v/v). The eluted compound was detected at 254 nm with a UV detector. The newly developed method was validated according to the ICH guidelines with respect to specificity, linearity, accuracy, precision and robustness. The standard curve of mean peak area versus concentration showed an excellent linearity over a concentration range 5.0–300 μg ml−1 FNH with regression coefficient (r) value of 0.9999. The limit of detection (S/N = 3) was 1.6 μg ml−1 and the limit of quantification (S/N = 10) was 4.9 μg ml−1. Forced degradation studies were also performed for the drug sample to demonstrate the stability indicating power of the developed UPLC method. Acidic, basic, hydrolytic, oxidative, thermal and photolytic degradation were used to assess the stability indicating power of the method. The drug was found to be stable in acidic, thermal, hydrolytic and photolytic stress conditions and showed slight degradation in oxidative stress condition and extensive degradation in basic medium.

Optimized chromatographic conditions for separation of halogenated acetic acids by ultra-performance liquid chromatography–electrospray ionization-mass spectrometry

Emerging halogenated acetic acids (HAAs), especially mixed halogenated acids such as chlorobromo-, chloroiodo- and bromoiodo-acetic acids, are unregulated disinfection by-products in drinking water. Because these compounds are hydrophilic and strongly acidic, they are difficult to detect at trace levels using approved analytical methods. In the present study, 13 HAAs were effectively separated on three ultra-performance liquid chromatography columns. The effects of changing in the aqueous mobile phase, acidic solutions and cationic volatile ion pair reagents were investigated. The samples were pretreated by filtration, and extraction, while derivatization and concentration procedures were not required. The limits of quantitation for regulated HAAs were between 0.5μg/L and 1.7μg/L and for unregulated HAAs were 1.2 and 5.8μg/L, especially for the iodinated acetic acids were 1.5 and 2.1μg/L. The method was applied to two finished water samples collected in China (Shanghai and Xuzhou) from water treatment plants that use chlorine for disinfection. Multiple unregulated HAAs were found in the two samples, but iodoacids were only detected in the water sample from Shanghai, which could be attributed to the characteristics of the source water. The presence of unregulated HAAs, especially mixed bromo- and iodoacetic acids, in the finished water samples could affect human health, and this warrants further investigation.

Determination of six Alternaria toxins with UPLC-MS/MS and their occurrence in tomatoes and tomato products from the Swiss market

An ultra performance liquid chromatography (UPLC)-tandem mass spectrometry (MS/MS) method was developed for the determination of the Alternaria toxins tenuazonic acid, alternariol, alternariol monomethyl ether, altenuene, altertoxin I and tentoxin. Owing to its instability, altenusin could not be determined. The sample preparation includes an acidic acetonitrile/water/methanol extraction, followed by SPE clean-up step, before injection into the UPLC-MS/MS system. The separation was made on an Acquity UPLC column using a water/acetonitrile gradient with ammonium hydrogen carbonate as a modifier. Matrix compounds of real samples led to enhancement as well as suppression of the target compounds, depending on analyte and matrix. The recoveries were between 58 and 109% at a level of 10μg/kg. Eighty-five tomato products, consisting of peeled and minced tomatoes, soup and sauces, tomato purées and concentrates, ketchup as well as dried and fresh tomatoes, were taken from the Swiss market in 2010. Tenuazonic acid was found most frequently (81 out of 85 samples) and in the highest levels of up to 790μg/kg. Alternariol and alternariol monomethyl ether were found in lower concentrations, ranging from <1 to 33μg/kg for alternariol and <5 to 9μg/kg for alternariol monomethyl ether. Only a few samples were positive for altenuene and tentoxin. Altertoxin I was never detected.

Application of ultra-performance columns in high-performance liquid chromatography for determination of albendazole and its metabolites in turkeys

Methods for determination of albendazole (ALB), albendazole sulfoxide (SOX) and albendazole sulfone (SON) in turkey blood plasma, using high-performance liquid chromatography (HPLC) with fluorescence detection, were developed. Moreover, comparison of HPLC columns with ultra-performance liquid chromatography (UPLC) columns was performed. Albendazol was administered orally in 5-week-old birds (n = 18) at a dose of 25 mg/kg b.w. Accuracy and precision of the developed method were satisfactory and stability studies showed acceptable variation (below 15%) in ALB, SOX and SON concentrations when the samples were stored at -75°C for 15 days. UPLC(®) columns gave higher peaks from typical HPLC columns retaining high quality of analysis. Pharmacokinetic analysis indicated quick elimination of ALB from turkey blood plasma. The mean residence time of SON was at least two times longer than that of SOX and four times longer than that of ALB. The elimination half-lives for ALB, SOX and SON were 0.7 ± 0.27, 5.37 ± 6.03, 9.17 ± 5.12 h, respectively. The obtained results indicate that the described method allows for precise determination of albendazole and its metabolites in turkey plasma. Moreover, using UPLC columns in HPLC apparatus results in higher sensitivity as compared with the classical HPLC columns.

Comparison of UPLC and HPLC for Analysis of Dietary Folates

Ultra performance liquid chromatography (UPLC) using small sub-2 μm particles and high performance liquid chromatography (HPLC) were compared for separation and determination of the most common dietary folates; 5-methyltetrahydrofolate, tetrahydrofolate, 5-formyltetrahydrofolate, 10-formylfolic acid and folic acid. Two UPLC columns—Acquity BEH C18 and Acquity HSS T3, and two HPLC columns with similar surface chemistry—Xbridge C18 and Atlantis d18 were tested. When using UPLC, the signal-to-noise ratio could be improved by a factor of 2–50 for different folate derivatives and the run time could be reduced fourfold without sacrificing separation efficiency. The applicability of UPLC to real food samples was demonstrated.

Quantification of Lansoprazole in Oral Suspension by Ultra-High-Performance Liquid Chromatography Hybrid Ion-Trap Time-of-Flight Mass Spectrometry.

An LC-MS/MS method was developed and validated to be used as a stability indicating assay for the study of a 3 mg/mL lansoprazole oral suspension. The method utilizes a UPLC (ultra-performance liquid chromatography) column and unique mass spectrometric detection (ion-trap time-of-flight (IT-TOF)) to achieve a sensitive (LOD 2 ng/mL), accurate, and reproducible quantification of lansoprazole. This method reports an intraday and interday coefficient of variation of 2.98 ± 2.17% (n = 5 for each concentration for each day) and 3.07 ± 0.89% (n = 20 for each concentration), respectively. Calibration curves (5–25 μg/mL) were found to be linear with an R2 value ranging from 0.9972 to 0.9991 on 4 different days. Accuracy of the assay, expressed as % error, ranged from 0.30 to 5.22%. This method is useful for monitoring the stability of lansoprazole in oral suspension.

A high‐sensitivity UPLC‐MS/MS method for simultaneous determination and confirmation of triptolide in zebrafish embryos

A high-sensitivity ultra-performance liquid-chromatography (UPLC) coupled with tandem mass spectrometric method was developed for simultaneous quantification and confirmation of triptolide in both zebrafish embryos and the aqueous-exposure solution on a tandem quadrupole mass spectrometer (TQ-MS). This was achieved by performing quantification using the multiple reaction monitoring (MRM) acquisition with simultaneous characterization of the MRM peak using product ion confirmation (PIC) acquisition as it elutes from the chromatographic system. Separation was achieved on a 1.7 µm C(18) UPLC column using 0.1% formic acid water-acetonitrile mobile phase with a cycle time of 6 min. The linear range of 0.115-360 ng/mL, and lower limits of detection of 0.02 ng/mL and quantification of 0.064 ng/mL were established. This method was successfully applied to determine the time course of triptolide absorption by zebrafish embryos and the amount of triptolide remaining in the culture medium after administration of two triptolide dosages at three time points. This coupled MRM with PIC approach could provide both qualitative and quantitative results without the need for repetitive analyses. This resulted in the reduction of further confirmative experiments and analytical time, and ultimately increased laboratory productivity.

High-Throughput log P Determination by Ultraperformance Liquid Chromatography: A Convenient Tool for Medicinal Chemists

Accurate determinations of lipophilicity indices benefit from recent advances in chromatographic sciences such as the launch of ultraperformance liquid chromatography (UPLC). The fast strategy presented here emerges as a powerful method suitable for high-throughput log P measurements of therapeutic compounds in isocratic and gradient modes. Because UPLC columns are highly stable in basic pH conditions, this approach allows a direct lipophilicity estimation of basic compounds in their neutral forms.