Performance Liquid Chromatography-diode Array Research Articles

Waterpipe smoke: A considerable source of human exposure against furanic compounds

Smoking of waterpipes became increasingly popular in the Western hemisphere in recent years. Yet, up to now only little is known about the health hazards and on the composition of waterpipe smoke. To obtain more information on the ingredients present in waterpipe smoke we utilized two different approaches. Based on headspace-solid-phase microextraction–gas chromatography–mass spectrometry (HS-SPME–GC–MS) instrumentation we identified new compounds present in the waterpipe smoke. Additional reversed-phase-high performance liquid chromatography–diode array detection (RP-HPLC–DAD) then led us to perform reliable quantification of the newly detected chemical species. Upon identification of a range of different furanic compounds such as 5-(hydroxymethyl)-2-furaldehyde (HMF), 2-furaldehyde, and others, we developed an easy-to-perform and fast RP-HPLC–DAD method to quantify these compounds in the complex matrix of waterpipe smoke. The detection limits range from 0.04 μg for HMF to 7.1 μg for 3-furan methanol per smoking session. Linearity, intra- and inter-day precision and recovery were determined and proved excellent. We analyzed 5 waterpipe tobacco brands and found up to 62.3 ± 11 mg of HMF generated during one waterpipe smoking session. The applied smoking protocol comprised 171 puffs of 530 mL each and 2.6 s duration every 20 s. Our results reveal that waterpipe smoking constitutes a major source of HMF exposure. Furthermore, we found a distinct filter effect of the bowl water for all furanic compounds investigated except HMF.

Application of an ultrasound-assisted surfactant-enhanced emulsification microextraction method for the analysis of diethofencarb and pyrimethanil fungicides in water and fruit juice samples

In this work, a simple, practical and environmentally friendly sample pre-treatment method, ultrasound-assisted surfactant-enhanced emulsification microextraction coupled with high performance liquid chromatography–diode array detector/electrospray ionisation mass spectrometry, was developed to determine diethofencarb and pyrimethanil residues in water and fruit juice samples. Tween 80 was used as an emulsifier and carbon tetrachloride was chosen as the extraction solvent, and no dispersive organic solvent was needed, which is typically required in common dispersive liquid–liquid microextraction methods. Several variables, such as the type and volume of extraction solvent and surfactant, extraction temperature and ultrasound extraction time were investigated and optimised. Under optimal conditions, the enrichment factors were 265 and 253 for diethofencarb and pyrimethanil, respectively. The limits of detection (LODs), calculated as three times the signal-to-noise ratio (S/N), were 0.01 μg L −1 for both diethofencarb and pyrimethanil. The linearity of the method was obtained in the range of 0.05–2000 μg L −1, with correlation coefficients of 0.9994–0.9998. The water (at fortified levels of 0.1 and 1.0 μg L −1) and fruit juice samples (at fortified levels of 0.1 and 1.0 μg L −1) were successfully analysed using the proposed method, and the relative recoveries were in the range of 88–114%, 93–111%, 86–117% and 94–101%, respectively.

Characterization and determination of the major constituents in Belamcandae Rhizoma by HPLC–DAD–ESI-MS n

Belamcandae Rhizoma, derived from the rhizome of Belamcanda chinensis (L.) DC., has been used as traditional Chinese medicine for the treatment of coughing and pharyngitis. However, there have been few studies dealing with the systematic analysis of the bioactive constituents in Belamcandae Rhizoma. In this work, high performance liquid chromatography–diode array detection–electrospray ionization multiple-stage mass spectrometry (HPLC-DAD-ESI-MS n ) combined with liquid chromatography–time of flight-mass spectrometry (HPLC–TOF/MS) was established for profiling and characterization of multi-constituent in Belamcandae Rhizoma. The ESI-MS n fragmentation behaviors of the authentic references were proposed for aiding the structural identification of components in the extract. Thirty-five flavonoids, including 30 isoflavones and five xanthones, were identified or tentatively identified by comparing their retention times, UV and MS spectra with those of authentic compounds or literature data. Twelve of the identified compounds (neomangiferin, mangiferin, tectoridin, iristectorin B, iristectorin A, iridin, tectorigenin, iristectorigenin A, irigenin, irisflorentin, irilone and dichtomitin) were determined by HPLC–DAD using a C 18 column. The results indicated that the developed analysis method could be employed as a rapid, effective technique for structural characterization of chemical constituents in herbal medicine. This work is expected to provide comprehensive information for the quality evaluation of Belamcandae Rhizoma, which would be a valuable reference for the further study and development of this herb and related medicinal products.

Characterization of flavone glycosides and aglycones in Cephalotaxus sinensis by HPLC-DAD-MS

High performance liquid chromatography diode array detector electrospray ionization mass spectrometry (HPLC-DAD-ESI-MS) was utilized to separate and identify flavone glycosides and aglycones in Cephalotaxus sinensis, a natural plant with an anti-hyperglycemic effect. The HPLC-DAD-ESI-MS system could provide a great amount of information on the structure of flavone aglycones and glycosides in C. sinensis without time-consuming pre-separation or purification procedures. A total of seven flavone glycosides and two flavone aglycones were unambiguously identified based on their retention times, ultraviolet spectra, molecular weights and MS/MS spectra. It's interesting that five 5-O-flavone glycosides were found in C. sinensis since 5-O-flavone glycosides are rare and difficult to form in the natural plant. Among them, quercitrin and apigenin-5-O-β-D-glucopyranoside were isolated from this genus for the first time. A novel flavone glycoside, chryoseriol-5-O-[α-L-rhamnopyranosyl-(1→4)-β-D-6-O-acetylglucopyranoside], was isolated for the first time and identified by the ultraviolet spectrum, infrared spectrum, mass spectrum and nuclear magnetic resonance spectrum. In addition, the proposed fragment pathway of the novel flavone glycoside was concluded in the negative ionization mode by HPLC-ESI-MS/MS.

Authentication of organic and conventional eggs by carotenoid profiling

Organic production benefits from fair competition and sustained consumer confidence. The latter can only be assured by paper trailing and verification assessments. Traditional analytical strategies for guaranteeing quality and uncovering adulteration have relied on the determination of the amount of a marker compound or compounds in a material and a subsequent comparison of the value(s) obtained with those established for equivalent material. Since it is unlikely to find a unique marker that allows discrimination between organic and conventional produce, selective fingerprinting (profiling) combined with chemometrics is a more promising approach. In this study, carotenoid High Performance Liquid Chromatography–Diode Array Detection profiling combined with k-nearest neighbour classification chemometrics was used to predict the production system (organic vs. non-organic) of chicken eggs. A proof-of-concept set (training set for the model) was composed of eggs from 24 organic farms, 12 free range, and 12 barn farms in the Netherlands. The identities of organic, free range, and barn eggs were predicted with success rates of 100%, 100%, and 84%, respectively. The identity of eggs determined using an additional market test set with eggs from 12 organic, 12 free range, and 12 barn farms originating from the Netherlands and New Zealand resulted in correct classifications for 35 of the 36 samples. The results of this study indicate that this fingerprint approach is a promising tool for analytical verification of the production system of organic eggs.

Interference-free simultaneous determination of Sudan dyes in chili foods using solid phase extraction coupled with HPLC–DAD

A simple, rapid, inexpensive and robust solid phase extraction (SPE) cleanup method coupled with high performance liquid chromatography–diode array detector (HPLC–DAD) for the determination of four Sudan dyes in chili products was developed. The interference substances were removed completely by normal SPE-Alumina N, resulting in a clean and interference-free baseline. Without the use of expensive LCMS/MS instrument the low levels of Sudan dyes were identified and quantified in chili foods with good precision and recovery. Depending on the Sudan dyes involved, the limit of determination (LOD) and limit of quantification (LOQ) were in the range of 4.1–5.8 and 13.2–19.1 μg/kg, respectively. The recovery, repeatability and reproducibility varied from 93.2% to 103%, from 0.3% to 4.4%, and from 0.6% to 4.9%, respectively. This method is suitable for the routine analysis of Sudan dyes for the common factory laboratory due to its sensitivity, simplicity, convenience and low-cost.

Consistency of FMEA used in the validation of analytical procedures

In order to explore the consistency of the outcome of a Failure Mode and Effects Analysis (FMEA) in the validation of analytical procedures, an FMEA was carried out by two different teams. The two teams applied two separate FMEAs to a High Performance Liquid Chromatography–Diode Array Detection–Mass Spectrometry (HPLC–DAD–MS) analytical procedure used in the quality control of medicines. Each team was free to define their own ranking scales for the probability of severity ( S), occurrence ( O), and detection ( D) of failure modes. We calculated Risk Priority Numbers (RPNs) and we identified the failure modes above the 90th percentile of RPN values as failure modes needing urgent corrective action; failure modes falling between the 75th and 90th percentile of RPN values were identified as failure modes needing necessary corrective action, respectively. Team 1 and Team 2 identified five and six failure modes needing urgent corrective action respectively, with two being commonly identified. Of the failure modes needing necessary corrective actions, about a third were commonly identified by both teams. These results show inconsistency in the outcome of the FMEA. To improve consistency, we recommend that FMEA is always carried out under the supervision of an experienced FMEA-facilitator and that the FMEA team has at least two members with competence in the analytical method to be validated. However, the FMEAs of both teams contained valuable information that was not identified by the other team, indicating that this inconsistency is not always a drawback.

Acute interstitial nephritis due to administration of multiple antibiotics:a case report

Objective:To identify the chemical components in traditional Chinese herbal medicine Ginseng by high performance liquid chromatography diode array detector (HPLC-DAD) and High performance liquid chromatography-time of flight mass spectrometry (HPLC-TOF/MS).Methods: An Agilent Zorbax XDB-Cl8 column (250 mm×4.6 mm,5 μm) was used for isolation and identification of chemical components in Ginseng with a mobile phase of acetonitrile and water in gradient elution.A gradient program was used as follows:1-35 min,19% A; 35-55 min,19%-29% A; 55-70 min,29% A; 70-110 min,29%-40% A; 110-150 min,95% A.The flow rate was set at 1.0 ml/min,the injection volume was 20 μl,and the column temperature was set at 20℃.The time-of-flight mass spectrometer was equipped with an EIS ion source.Scanning mass range was between m/z 100-1 350.Results: Thirty-nine chemical compounds in Ginseng were identified unequivocally.Conclusion: Chromatographic demonstration of 39 chemical compounds of Ginseng in one run is achieved by HPLC-TOF/MS,which provides a foundation for further study on the metabolism and action mechanism of traditional Chinese herbal medicine.

MCRC software: A tool for chemometric analysis of two-way chromatographic data

This paper describes the development and implementation of MCRC software, chemometric software for Multivariate Curve Resolution of two-way Chromatographic data. MCRC software is developed for chemometric analysis of chromatographic data; however, it may also be used for other types of multivariate data. It consists of five groups of techniques for preprocessing, chemical rank determination, local rank analysis, multivariate resolution and peak integration. This software has the ability of the analysis of complex multi-component chromatographic signals of gas chromatography–mass spectrometry (GC–MS) and high performance liquid chromatography–diode array detection (HPLC–DAD). The software allows a user to apply the implemented methods in an easy way and it gives a straightforward possibility to visualize the obtained results. The main features of the presented software are providing a number of preprocessing techniques, implementation of different chemical rank determination methods, usage of iterative and non-iterative resolution techniques and a user-friendly environment with a variety of graphical outputs. The implementation of the MCRC software is demonstrated by the analysis of an overlapped peak cluster of simulated GC–MS data.

Comprehensive Comparisons between 1-Phenyl-3-methyl-5-pyrazolones, 1-(4-Methoxyphenyl)-3-methyl-5-pyrazolones and 1-(2-Naphthyl)-3-methyl-5-pyrazolones as Labeling Reagents Used in LC-DAD-ESI-MS-MS Analysis of Neutral Aldoses and Uronic Acids

Comprehensive comparisons between three 1-aryl-3-methyl-5-pyrazolone (AMP) labeling reagents were carried out for the analysis of reductive monosaccharides using reversed phase high performance liquid chromatography diode array detection coupled to electrospray ionization mass spectrometry. AMP derivatives included 1-phenyl-3-methyl-5-pyrazolone (PMP), 1-(4-methoxyphenyl)-3-methyl-5-pyrazolone (PMPMP) and 1-(2-naphthyl)-3-methyl-5-pyrazolone (NMP). The separation of AMP-monosaccharides was found to be pH-dependent under reversed phase conditions and acceptable separations were obtained at pH < 4.5. The elution orders of AMP-aldoses were rationalized by geometric factors involved in the presence of hydroxyl groups at C2 and C3 positions of the saccharide moiety. When PMP or PMPMP were used as labeling agents Glucose and galactose were completely separated, while arabinose and xylose were observed to co-elute. The use of NMP revealed that arabinose and xylose could be separated while glucose and galactose were co-eluted. MS-MS data of AMP-monosaccharides gave characteristic fragment ions resulting from cleavage between the C2-C3 bond (m/z 373 for PMP derivatives, m/z 433 for PMPMP derivatives, and m/z 473 for NMP derivatives). Bond breakage between C1-C2 (m/z 359 for PMP derivatives, m/z 419 for PMPMP derivatives, and m/z 459 for NMP derivatives) of the saccharide moiety was also found to be characteristic for AMP-saccharides. Application to the analysis of hydrolyzed rape pollen polysaccharides revealed the presence of four unexpected monosaccharides, namely ribose, erythrose, threose and glyceraldehyde.

Microbial degradation of 17β -estradiol and 17α -ethinylestradiol followed by a validated HPLC-DAD method

This work aimed at studying the biodegradation of two estrogens, 17α -estradiol (E2) and 17β -ethinylestradiol (EE2), and their potential metabolism to estrone (E1) by microbial consortia. The biodegradation studies were followed by High Performance Liquid Chromatography–Diode Array Detector (HPLC–DAD) using a specifically developed and validated method. Biodegradation studies of the estrogens (E2 and EE2) were carried out with activated sludge (consortium A, CA) obtained from a Wastewater Treatment Plant (WWTP) and with a microbial consortium able to degrade recalcitrant compounds, namely fluorobenzene (consortium B, CB). E2 was more extensively degraded than EE2 by CA whereas CB was only able to degrade E2. The addition of acetate as a supplementary carbon source led to a faster biodegradation of E2 and EE2. E1 was detected as a metabolite only during the degradation of E2. The 16S rRNA gene sequence analyses of strains recovered from the degrading cultures revealed the presence of the genera Pseudomonas, Chryseobacterium and Alcaligenes. The genera Pseudomonas and Chryseobacterium were retrieved from cultures supplied with E2 and EE2, while the genus Alcaligenes was found in the presence of E2, suggesting that they might be involved in the degradation of these compounds.

Comparison of two different isolation methods of benzimidazoles and their metabolites in the bovine liver by solid-phase extraction and liquid chromatography–diode array detection

A new analytical method that uses high performance liquid chromatography–diode array detector (HPLC–DAD) was developed for the analysis of 14 benzimidazoles residues, including metabolites, in bovine liver. Samples were extracted using two different extraction procedures: with phosphate buffer after enzymatic hydrolysis (method A) or using organic solvent, i.e. acetonitrile (method B). Then, samples were purified on a strong cation exchange (SCX) cartridge and analyzed in HPLC/DAD. The recovery percentages, obtained spiking the matrix (liver) at concentrations of 500 and 100 μg kg −1 with a standard mixture of benzimidazoles, were in the range 6–101% and 80–102% for methods A and B, respectively. The repeatability of the methods was assessed in all cases by the % of correlation value (CV) that was lower than 19%. The limits of quantification (LOQs) in the matrix for methods A and B were in the range 40–60 and 20–50 μg kg −1, respectively. The best of the two methods, method B, was used for the analysis of 10 bovine liver samples.

Study on the radical scavengers in the traditional Chinese medicine formula Shengmai San by HPLC–DAD coupled with chemiluminescence (CL) and ESI–MS/MS

Shengmai San (SMS) has been used for the treatment of cardiovascular disease in Asia for long-time. It has been reported that SMS can protect against oxidative stress and tune immune response in patients. In this paper, a high performance liquid chromatography–diode array detection–chemiluminescence (HPLC–DAD–CL) method for on-line detection was successfully applied to screen antioxidants in complex SMS extracts. The H 2O 2 scavenging activity of every fraction of SMS extracts was measured to evaluate each contribution to the total anti-oxidative activities of the prescription, followed by structures identification of the main active components by LC/MS/MS. HPLC–DAD–CL method was also applied for comparing the scavenging rates of main active components between the individual herbs and SMS. Interestingly, we found that the scavenging activities of main components detected in the individual herbs were different from those in SMS. The experimental data indicated that drug interactions during decocting process could result in the changes of the antioxidants’ solubility. This study demonstrated that HPLC–DAD–CL on-line assay was a useful technique for rapidly screening and identifying bioactive components from complex multi-herbal prescription.

Fast chromatographic method for the determination of dyes in beverages by using high performance liquid chromatography—Diode array detection data and second order algorithms

A fast chromatographic methodology is presented for the analysis of three synthetic dyes in non-alcoholic beverages: amaranth (E123), sunset yellow FCF (E110) and tartrazine (E102). Seven soft drinks (purchased from a local supermarket) were homogenized, filtered and injected into the chromatographic system. Second order data were obtained by a rapid LC separation and DAD detection. A comparative study of the performance of two second order algorithms (MCR-ALS and U-PLS/RBL) applied to model the data, is presented. Interestingly, the data present time shift between different chromatograms and cannot be conveniently corrected to determine the above-mentioned dyes in beverage samples. This fact originates the lack of trilinearity that cannot be conveniently pre-processed and can hardly be modelled by using U-PLS/RBL algorithm. On the contrary, MCR-ALS has shown to be an excellent tool for modelling this kind of data allowing to reach acceptable figures of merit. Recovery values ranged between 97% and 105% when analyzing artificial and real samples were indicative of the good performance of the method. In contrast with the complete separation, which consumes 10 mL of methanol and 3 mL of 0.08 mol L −1 ammonium acetate, the proposed fast chromatography method requires only 0.46 mL of methanol and 1.54 mL of 0.08 mol L −1 ammonium acetate. Consequently, analysis time could be reduced up to 14.2% of the necessary time to perform the complete separation allowing saving both solvents and time, which are related to a reduction of both the costs per analysis and environmental impact.

Efficient Method for Extraction and Simultaneous Determination of Active Constituents in Cornus officinalis by Reflux Extraction and High Performance Liquid Chromatography with Diode Array Detection

A high performance liquid chromatography diode array detector (HPLC-DAD) based method was developed for the simultaneous determination of the three active constituents, morroniside, loganin, and gallic acid in Cornus officinalis fruits. The separation was achieved on a C18 column with a gradient solvent system of acetonitrile, methanol, and 0.1% acetic acid at 260 nm. The method was validated for linearity, repeatability, limits of detection, and limits of quantification. The analytes were confirmed by electrospray ionization mass spectrometry (ESI-MS). In addition, the effect of various parameters including solvent, extraction technique, and time on the extraction of the three active constituents from C. officinalis fruits was examined. The highest extraction efficiency was obtained with 100% methanol using reflux extraction. The developed method could be used to effectively and comprehensively evaluate the quality of C. officinalis.

Diuron biodegradation in activated sludge batch reactors under aerobic and anoxic conditions

Diuron biodegradation was studied in activated sludge reactors and the impacts of aerobic and anoxic conditions, presence of supplemental substrate and biomass acclimatization on its removal were investigated. Diuron and three known metabolites, namely DCPMU (1-(3,4-dichlorophenyl)-3-methylurea), DCPU (1-3,4-dichlorophenylurea) and DCA (3,4-dichloroaniline), were extracted by solid-phase extraction (dissolved phase) or sonication (particulate phase) and determined using High Performance Liquid Chromatography–Diode Array Detector (HPLC–DAD). During the experiments only a minor part of these compounds was associated with the suspended solids. Under aerobic conditions, almost 60% of Diuron was biodegraded, while its major metabolite was DCA. The existence of anoxic conditions increased Diuron biodegradation to more than 95%, while the major metabolite was DCPU. Mass balance calculation showed that a significant fraction of Diuron is mineralized or biotransformed to other unknown metabolites. The presence of low concentrations of supplemental substrate did not affect Diuron biodegradation, whereas the acclimatization of biomass slightly accelerated its elimination under anoxic conditions. Calculation of half-lives showed that under aerobic conditions DCPMU, DCPU and DCA are biodegraded much faster than the parent compound. In the future, the sequential use of anoxic and aerobic conditions could provide sufficient removal of Diuron and its metabolites from runoff waters.

Photodegradation of E110 and E122 dyes in a commercial aperitif: A high performance liquid chromatography–diode array–tandem mass spectrometry study

A commercial aperitif containing E110 and E122 dyes is exposed to photo-irradiation in solar box under conditions chosen to simulate sun light irradiation. After 16 days of irradiation the red-coloured aperitif assumes a very pale yellow colour. HPLC–diode array–MS/MS analysis evidences the presence in the commercial aperitif of some impurities with naphthalene-based structures. The structures for the photodegradation products are proposed.

Chromatographic fingerprint analysis and characterization of furocoumarins in the roots of Angelica dahurica by HPLC/DAD/ESI-MS n technique

A high performance liquid chromatography–diode array detection–electrospray ionization tandem mass spectrometry (HPLC/DAD/ESI-MS n ) method was used for the chromatographic fingerprint analysis and characterization of furocoumarins in the roots of Angelica dahurica for the first time. Nine “common peaks” were identified by comparing with the retention time, UV and MS spectra of reference furocoumarins. The software “Similarity Evaluation System for Chromatographic Fingerprint of TCM” was used to evaluate the similarities of 13 batches of Baizhi samples collected from Henan, Zhejiang, Sichuan and Anhui provinces of China. The results indicated that the samples from different batches had similar HPLC fingerprints, and the method could be applied for the quality control of the roots of Angelica dahurica. In addition, a total of 20 furocoumarins were identified or tentatively characterized by HPLC/DAD/ESI-MS n technique, and their fragmentation patterns in an electrospray ion trap mass spectrometer were also summarized.

HPLC Determination of Pesticides in Soybeans using Matrix Solid Phase Dispersion

A method to determine six pesticides applied in soy cultivation (imazethapyr, imazaquin, metsulfuron‐methyl, carboxin, chlorimuron‐ethyl, and tebuconazole) using matrix solid phase dispersion (MSPD) as the extraction technique followed by a clean‐up step, using a C8 co‐column, and subsequent chromatographic analysis by high performance liquid chromatography diode array detection (HPLC‐DAD) was developed. The validated method showed good recuperation for all pesticides (60–120%), except for metsulfuron‐methyl and tebuconazole at their lowest concentration levels, and the quantification limits of the method (0.04–0.08 µg g−1) were below the maximum residue limits imposed by the principal regulatory agencies. This article not subject to U.S. copyright law.

Determination of imipenem and rifampicin in mouse plasma by high performance liquid chromatography–diode array detection

A high performance liquid chromatographic (HPLC) method for the determination of imipenem and rifampicin was developed and validated. The method involves plasma deproteinisation with methanol, gradient elution on a RP-18 column and diode array detection. Separation was carried out in 8 min using a mobile phase composed of methanol and 0.2 M borate buffer (pH 7.2). Imipenem and rifampicin were detected at 300 nm and 255 nm, respectively. A linear response was observed at plasma levels ranged between 0.3 and 30 μg mL −1 for imipenem and 1.5 and 20 μg mL −1 for rifampicin. The detection limits were 0.07 μg mL −1 and 0.47 μg mL −1 for imipenem and rifampicin, respectively. The method was applied to the determination of both compounds in mouse plasma samples.