Development and single-laboratory validation of a reversed-phase liquid chromatography-electrospray-tandem mass spectrometry method for identification and determination of acrylamide in foods

A rapid and accurate method using reversed-phase liquid chromatography-tandem mass spectrometry interfaced with electrospray was developed for determination of acrylamide in cooked food samples. A simplified sample treatment procedure using an extraction step with acidified water without cleanup was developed. A C18 column with an aqueous formic acid-methanol mixture as the mobile phase was used under isocratic conditions. The method was validated in-house for robustness, limits of detection (LOD) and quantitation (LOQ), linearity, recovery, and accuracy both on standard and baked-product and potato flour matrixes. Good results in the low ppb level were obtained for LOD (< 15 microg/kg) and LOQ (< 25 microg/kg) of acrylamide in samples. Excellent linearity (r2 = 0.999-1.000) was established over 2 orders of magnitude by performing statistical tests. The absence of both constant and proportional systematic errors demonstrated good method accuracy. Excellent results were obtained for intraday repeatability (RSD < 1.5%) and between-day precision (RSD < 5%). Extraction recoveries from food products were calculated in the 97 +/- 3-99 +/- 2% (n = 6) range with a labeled internal standard (13C3-acrylamide). The applicability of the method to determination of acrylamide in cooked food products was demonstrated.     

Development and in-house validation of a liquid chromatography-electrospray-tandem mass spectrometry method for the simultaneous determination of Sudan I, Sudan II, Sudan III and Sudan IV in hot chilli products

An accurate method based on the use of reversed-phase (RP) liquid chromatography-tandem mass spectrometry interfaced with electrospray (LC-ESI-MS/MS) was devised for the determination of Sudan I, Sudan II, Sudan III and Sudan IV in hot chilli food samples. A simple sample treatment procedure entailing the use of an extraction step with acetone without clean-up was developed. A C18 column with an aqueous formic acid/methanol mixture as the mobile phase was used under isocratic conditions. Mass spectral acquisition was done in positive ion mode by applying selected reaction monitoring of three fragmentation transitions per compound to provide a high degree of selectivity. The method was in-house validated in terms of detection limits (LOD), quantitation limits (LOQ), linearity, sensitivity, accuracy, recovery, and selectivity on two kinds of hot chilli sauces. Good results in the low ng/g level were obtained for LOD and LOQ of all analytes in matrices. Under both intra-day repeatability (R.S.D. between 1 and 13%) and intermediate precision (about 5-15% R.S.D. for both chilli sauce matrices) conditions, precision proved to be typical of determinations based on electrospray LC-MS and acceptable for routine monitoring purposes. Extraction recoveries for all four azo-dyes in chilli tomato sauce ranged from 92 to 103% at a spiking level of 5 microg/kg, whereas values between 72 and 97% were calculated in chilli tomato and cheese sauce at the same concentration level. The applicability of the method to the determination of Sudan azo-dyes in hot chilli products was demonstrated.     

Development and validation of a solid phase micro-extraction-gas chromatography-mass spectrometry method for the determination of furan in baby-food

An efficient and simple method for the determination of furan in baby-food (vegetables and fruits) by solid phase micro-extraction-gas chromatography-mass spectrometry (SPME-GC-MS) was developed and validated. Experimental design was used to investigate the effects of temperature and time of extraction. The calculated regression model was used to find the experimental conditions providing the optimal SPME extraction yield. Validation was carried out in terms of limit of detection (LOD), limit of quantitation (LOQ), linearity, precision and trueness. LOD and LOQ values in the low ngkg(-1) were achieved, whereas linearity was established over two order of magnitude. Good precision was obtained both in terms of intra-day repeatability and between-day precision on two concentration levels (RSD% lower than 3.6%). Recovery values of 91.5+/-6.2% and of 96.1+/-1.3% (n=3) were calculated at 75 ngkg(-1) and 75 microgkg(-1) level. Finally, the applicability of the method to the determination of furan in a number of commercial and home-made baby-food samples was demonstrated.     

Confirmatory method for the determination of volatile congeners and methanol in Turkish raki according to European Union Regulation (EEC) No. 2000R2870: single-laboratory validation

A method described by European Union Regulation (EEC) No. 2000R2870 was validated and supported by GC/MS analysis for the determination of volatile congeners and methanol in Turkish raki. The method was validated in terms of specificity, accuracy, precision, LOD, LOQ, linearity, and robustness. The specificity of the method was demonstrated, and the method showed excellent accuracy (97.5-100.1%). Linearity was checked in the ranges of 0.200-26.390 mg/100 mL for more volatile compounds and 1.155-48.00 mg/100 mL for less volatile compounds, after concentrations found in Turkish raki were taken into account. The calibration curves of all analytes showed good linearity (R2 > 0.998). The within- and between-day precision (RSD) values of 11 analytes were in the range of 0.18-4.50%. The LOD and LOQ values were in the range of 0.014-0.362 and 0.045-1.085 mg/100 mL, respectively. The method can be used as an absolute quantification method for the determination of volatile congeners and methanol in Turkish raki and for QC.     

Rapid method for the determination of vitamins A and E in foods using ultra-high-performance liquid chromatography

A rapid and novel ultra-HPLC (u-HPLC) method for the determination of vitamins A (retinol) and E (alpha-, gamma-, and delta-tocopherol) in foods was validated in terms of its precision, accuracy, and linearity. The u-HPLC separation was performed on an RP C18 column (particle size 2 microm, id 2 mm, and length 75 mm), followed by fluorescence detection. The recovery of retinol was more than 84.58%; the LOD and LOQ of the u-HPLC analysis were 0.015 and 0.045 mg/kg, respectively. The intraday and interday precision was less than 9.12%. The recoveries of alpha-, gamma-, and delta-tocopherol were more than 81.37%; the LOD and the LOQ were 0.014, 0.002, and 0.001 mg/kg and 0.042, 0.005, and 0.004 mg/kg, respectively. All calibration curves had good linearity (r2 = 0.99) within the test ranges. The novel, rapid method coupled to u-HPLC can provide significant improvements in the speed, sensitivity, and resolution compared with a conventional HPLC method.     

Use of experimental design for the purge-and-trap-gas chromatography-mass spectrometry determination of methyl tert.-butyl ether,tert.-butyl alcohol and BTEX in groundwater at trace level

An efficient method for the simultaneous determination of methyl tert.-butyl ether, tert.-butyl alcohol, benzene, toluene, ethylbenzene and xylene isomers in groundwater by purge-and-trap-gas chromatography-mass spectrometry was developed and validated. Experimental design was used to investigate the effects of temperature of extraction, time of extraction and percentage of salt added to the water samples. Regression models and desirability functions were applied to find the experimental conditions providing the highest global extraction yield. Validation was carried out in terms of limits of detection (LOD), limits of quantitation (LOQ), linearity and precision. LOD values ranging from 2.6 to 23 ng l(-1) were achieved, whereas linearity was statistically verified over two orders of magnitude for each compound. Precision was evaluated testing two concentration levels. Good results were obtained both in terms of intra-day repeatability and intermediate precision: RSD% lower than 4.5% at the highest concentration and lower than 13% at the lowest one were calculated for intra-day repeatability. A groundwater sample suspected of contamination by leaking underground petroleum storage tanks was analysed and some of the analytes were detected and quantitated.     

Development and validation of a solid-phase extraction and gas chromatography-tandem mass spectrometry method for the determination of isopropyl-9H-thioxanthen-9-one in carton packaged milk

A simple and efficient method for the determination of isopropyl-9H-thioxanten-9-one (ITX) in different fat content milk samples and baby milk samples stored in packaged cartons was developed and validated. Samples were extracted using solid-phase extraction (SPE) and analysed by gas chromatography-tandem mass spectrometry operated in selected reaction monitoring mode (SRM). Validation was carried out in terms of limit of detection (LOD), limit of quantitation (LOQ), linearity, precision and trueness. LOD and LOQ values in the low microg/L were achieved, whereas linearity was established within 0.5-500 microg/L range. Good precision was obtained both in terms of intra-day repeatability and inter-day precision on two concentration levels (RSD% lower than 2%). Good percentage recoveries were obtained (92.0-102.0%) even in the presence of high amount of fat. Finally, the developed method was successfully applied to analyse a number of commercial milk samples with different fat content and baby milk samples.     

Development and validation of a HPLC method for 4,7-phenanthroline-5,6-dione I and identification of its major impurity by HPLC-MS-APCI

In this study, the development and validation of an analytical method for the assay of 4,7-phenanthroline-5,6-dione I (dione I) using high-performance liquid chromatography (HPLC) and the determination of its synthetic impurities by employing the method in HPLC-mass spectrometry with atmospheric-pressure chemical ionization and photodiode-array UV detection is reported. The results show that dione I is eluted as a spectrally pure peak resolved from its impurities. 5-Bromo,4-7-phenanthroline is identified as the main impurity. This is supported by elemental analysis of the dione I, which demonstrated the presence of bromine. Validation parameters such as specificity and selectivity, linearity, accuracy, precision, limit of detection (LOD), limit of quantitation (LOQ), ruggedness, stability, and system suitability, which are evaluated for this method. The LOD and LOQ are 2.0 microg/mL and 50 microg/mL with a 0.50% relative standard deviation (%RSD), respectively. The calibration curves showed good linearity over the concentration range of 0.05-1.50 mg/mL. The correlation coefficient is > 0.9991 in each case. The %RSD values for intra- and interday precision studies are < 0.40%.     

Validation of HPLC‐UV method for determination of minor glycosides contained in Stevia rebaudiana Bertoni leaves

Leaves of Stevia rebaudiana contain glycosides with sweetness and biological activity. However besides the major glycosides, there are other glycosides within extracts that may contribute to its activity, and therefore it is important to quantify them. In this work, an isocratic HPLC method was validated for determination of dulcoside A, steviolbioside, rebaudioside C and rebaudioside B. An HPLC method was performed using a C₁₈ column (250 × 4.6 mm, particle size 5 µm) and a UV detector set at 210 nm. The mobile phase consisted of a 32:68 (v/v) mixture of acetonitrile and sodium phosphate buffer (10 mmol/L, pH 2.6), set to a flow rate of 1.0 mL/min. The calculated parameters were: sensitivity, linearity, limit of detection (LOD), limit of quantification (LOQ), accuracy and precision. The calibration curves were linear over the working range 25–150 µg/mL, with coefficient of correlation of ≥0.99 and coefficient of determination of ≥0.98. The LOD was 5.68–8.81 µg/mL, while the LOQ was 17.21–26.69 µg/mL. The percentage recoveries of fortified samples were 100 ± 10% and precision, relative standard deviation, was <10%. The method validation showed accuracy, linearity and precision; therefore this method can be applied for quantitative analysis of minor steviol glycosides in S. rebaudiana leaves. Copyright © 2014 John Wiley & Sons, Ltd.     

Use of experimental design for optimisation of the cold plasma ICP-MS determination of lithium,aluminum and iron in soft drinks and alcoholic beverages

A sensitive method for the simultaneous determination of (7)Li, (27)Al and (56)Fe by cold plasma ICP-MS was developed and validated. Experimental design was used to investigate the effects of torch position, torch power, lens 2 voltage, and coolant flow. Regression models and desirability functions were applied to find the experimental conditions providing the highest global sensitivity in a multi-elemental analysis. Validation was performed in terms of limits of detection (LOD), limits of quantitation (LOQ), linearity and precision. LODs were 1.4 and 159 ng L(-1) for (7)Li and (56)Fe, respectively; the highest LOD found being that for (27)Al (425 ng L(-1)). Linear ranges of 5 orders of magnitude for Li and 3 orders for Fe were statistically verified for each compound. Precision was evaluated by testing two concentration levels, and good results in terms of both intra-day repeatability and intermediate precision were obtained. RSD values lower than 4.8% at the lowest concentration level were calculated for intra-day repeatability. Commercially available soft drinks and alcoholic beverages contained in different packaging materials (TetraPack, polyethylene terephthalate (PET), commercial cans and glass) were analysed, and all the analytes were detected and quantitated.     

Thin-layer chromatography-densitometric analysis of alpha-mangostin content in Garcinia mangostana fruit rind extracts

The fruit rinds of mangosteen (Garcinia mangostana Linn.) have long been used as traditional medicines for treatment of skin infections, wounds, and diarrhea. A simple thin-layer chromatography (TLC)-densitometric method has been developed for the simultaneous quantification of alpha-mangostin in the extracts from unripe and ripe fruit rinds of G. mangostana. It was found in the ranges of 10.48 +/- 0.83 and 16.65 +/- 0.38% (w/w) in the dried unripe and ripe fruit rinds, respectively. The method was validated for linearity, precision, accuracy, limit of detection (LOD), and limit of quantitation (LOQ). The linearity was found over the range of 100-500 ng/spot with regression coefficient 0.999. Intraday and interday precision studies showed the relative standard deviation was <2%. Accuracy of the method was determined by a recovery study conducted at 3 different levels, and the average recovery was 99.49%. The LOD and LOQ were 40 and 100 ng, respectively. The proposed TLC-densitometric method was found to be simple, precise, specific, sensitive, and accurate. This method can be used for routine quality control of raw material of G. mangostana fruit rind, extract, and its products. It also can be applied in quantifying this marker compound in other drugs.     

Determination of ciprofloxacin and its impurities by capillary zone electrophoresis

Capillary zone electrophoresis (CZE) has been elaborated for separation, identification and determination of ciprofloxacin and its impurities. The separation, phosphate buffer pH 6.0 was supplemented with 0.075 M pentane-1-sulfonic acid sodium salt. The elaborated method was validated. The selectivity, linearity, limits of detection (LOD) and quantification (LOQ), precision, and accuracy of capillary zone electrophoresis were evaluated. The results obtained by CZE were also compared with those obtained by liquid chromatography. Regarding the validation results the CE method fulfils the current European Pharmacopoeia (Eur. Ph.) requirements. The evaluated CE method could be applicable to the analysis of different medicinal products containing ciprofloxacin.     

Optimization and validation of arsenic determination in foods by hydride generation flame atomic absorption spectrometry

A hydride generation flame atomic absorption spectrometric method was developed and optimized to quantitate arsenic (As) in foods. A wet digestion of the samples with HNO3 + H2O2 was performed and excess oxidants were eliminated by addition of hydrochloric acid and urea. As5+ in As3+ was then reduced by potassium iodide. The As3+ solution was analyzed by generation of arsine with sodium tetrahydroborate. As determination ranged from 2.5 to 20 microg/L, with a determination coefficient of 0.997. The limits of detection (LOD) and quantitation (LOQ) were 0.6 and 2.1 microg/L, respectively. The method was validated and good results were obtained for recovery, precision, accuracy, LOD, and LOQ. This method is now used to analyze foods from Rwanda.     

UV spectrophotometric determination of bioflavonoids from a semisolid pharmaceutical dosage form containing Trichilia catigua Adr. Juss and Ptychopetalum olacoides Bentham standardized extract: analytical method validation and statistical procedures

A precise, accurate, and sensitive UV spectrophotometric method was developed and validated for routine quantification of total bioflavonoids, expressed as rutin, from a topical oil-in-water pharmaceutical emulsion containing the extract of Trichilia catigua Adr. Juss and Ptychopetalum olacoides Bentham. The method was validated experimentally, and the data were treated rigorously by statistical analysis. The following analytical parameters were assessed: linearity, specificity, intra- and interrun precision measured as relative standard deviation (RSD, %), intra- and interrun accuracy (E, %), recovery (Rec., %), limit of detection (LOD, microg/mL), and limit of quantification (LOQ, microg/mL). The UV spectrophotometric method was linear (r = 0.9995) for standard rutin over the concentration range of 5.0-15.0 microg/mL with specificity for total bioflavonoids (expressed as rutin) at 361.0 nm with an absence of interferents from the complex matrix; RSD of < or = 1.79%, intrarun (E = 97.88 +/- 1.75 to 99.0 +/- 0.33%) and interrun (E = 98.38 +/- 1.12 to 100.79 +/- 1.30%) accuracy; Rec. = 98.64 +/- 0.42 to 100.74 +/- 0.41%; LOD = 0.20 microg/mL; and LOQ = 0.30 microg/mL.     

QuEChERS-同位素内标-高效液相色谱-串联质谱法测定动物源性食品中植物生长调节剂类农药残留

建立了高效液相色谱-串联质谱法测定动物源性食品中植物生长调节剂类农药残留量的方法。选取猪肉、牛肉、鸡肉、猪肝、鸡蛋和牛奶作为样品,样品经乙腈提取,4 g无水硫酸镁(MgSO₄)和1 g氯化钠(NaCl)盐析脱水后,取上清液经50 mg N-丙基乙二胺(PSA)+50 mg十八烷基硅烷(C18)粉末净化(含150 mg MgSO₄)。采用Agilent ZORBAX Eclipse Plus C18柱分离待测物,电喷雾电离,正负离子切换多反应检测模式检测,以乙腈和5 mmol/L乙酸铵水溶液作为流动相进行梯度洗脱,基质匹配内标法定量。在猪肝、鸡蛋基质中,矮壮素、噻苯隆和多效唑在0.1~100 μg/L范围内线性关系良好;在猪肉、牛肉和鸡肉中3种植物生长调节剂在0.1~50 μg/L范围内线性关系良好;在牛奶基质中,噻苯隆和多效唑的线性范围为0.05~10 μg/L,矮壮素的线性范围为0.05~5 μg/L,相关系数(r²)均大于0.990。以信噪比(S/N)≥3对应的添加水平作为检出限(LOD), S/N≥10对应的添加水平作为定量限(LOQ),矮壮素、噻苯隆和多效唑在不同基质下的LOD为0.01~0.1 μg/kg, LOQ为0.5~5 μg/kg。分别添加LOQ、2倍LOQ和10倍LOQ 3个水平的目标化合物,平均回收率为70.0%~117.4%, RSD为0.8%~16.1%。该方法操作简单、灵敏度高,采用基质匹配内标法定量,能最大限度地消除基质干扰,使检测结果更加精确,可满足动物源性食品中矮壮素、噻苯隆和多效唑残留的定量检测工作。     

Residues of chlormequat and mepiquat in grain--results from the Danish National Pesticide Survey

The objective of the present work was to establish information on chlormequat and mepiquat residues in grain for human consumption. Chlormequat (2-chloro-N,N,N-trimethylethylammonium, CAS RN 7003-89-6) and mepiquat (1,1-dimethylpiperidinium, CAS RN 15302-91-7) are plant growth regulators used to stabilize stalks in cereals. The study was part of the Danish National Pesticide Survey, managed by the Danish Veterinary and Food Administration. Samples were collected in autumn 1997. Residue contents were determined with a newly developed liquid chromatographic-tandem mass spectrometric (LC-MS/MS) method for chlormequat analysis. The method was extended to include mepiquat in the present study. Quantitation was done by the internal standards method, using mass chromatograms of the most intense daughter ions of mepiquat (m/z 98), chlormequat (m/z 58), and [13C]-chlormequat (m/z 61, internal standard). For chlormequat, the overall limit of detection (LD) was 6 micrograms/kg and the limit of determination (LOD) was 10 micrograms/kg. For mepiquat, LD was 2 micrograms/kg and LOD was 3 micrograms/kg. Of 77 samples analyzed, 51 contained chlormequat and 11 contained mepiquat. The highest levels of chlormequat were found in samples of oatmeal (3.76 mg/kg) and rye (1.08 mg/kg). In 9 rye grain samples containing chlormequat, 5 also contained mepiquat. However, in all samples analyzed, the residues of chlormequat and mepiquat were below maximum residue limits.     

Development of a simple and sensitive high-performance liquid chromatography method for determination of glucosamine in pharmaceutical formulations

A column high-performance liquid chromatography (HPLC) method was developed for the determination of glucosamine in dosage forms. Glucosamine was derivatized by addition of a solution containing orthophthaldialdehyde. The HPLC separation was achieved on a Spherimage 80 ODS2 column (250 x 4 mm id, 5 microm particle size) using an isocratic mobile phase containing phosphate buffer-methanol (90 + 10, v/v, pH 6.50) and methanol-tetrahydrofuran (97 + 3, v/v) in proportions of 85 + 15 at a flow rate of 1 mL/min, followed by fluorescence detection. The method was validated for specificity, linearity, accuracy, precision, limit of detection (LOD), and limit of quantitation (LOQ). The detector response for glucosamine HCI was linear over the concentration range of 0.1-20 microg/mL with a correlation coefficient of 0.9980. The accuracy was between 99.4 and 100.8%. The LOD and the LOQ were 0.009 and 0.027 microg/mL, respectively. The method was applied to determination of glucosamine in solid dosage forms.     

Validation of a gas chromatography/triple quadrupole mass spectrometry based method for the quantification of pesticides in food commodities

A new multiresidue method has been validated in cucumber matrix for the routine analysis of 130 multiclass pesticide residues by gas chromatography/triple quadrupole mass spectrometry. The pesticides were extracted with ethyl acetate. A first identification of the pesticides was based on a tandem mass spectrometric (MS/MS) screening method, which monitors a single transition for each target compound, in less than 12 min. After that, potentially non-negative samples were analyzed again by the MS/MS confirmation/quantification method, which monitors two or three MS/MS transitions for each compound, also in less than 12 min. Performance characteristics, such as trueness, precision, linear range, detection limit (LOD) and quantification limit (LOQ), for each pesticide were calculated. The average recoveries obtained ranged between 70 and 120% at three different fortification levels (25, 200 and 500 microg/kg) with precision, expressed as relative standard deviation (RSD), values lower than 15%. The calculated LOD and LOQ were typically <3.2 and 9.6 microg/kg, respectively. Such limits were much lower than the maximum residue levels (MRLs) established by European legislation. The proposed methodology was applied to the determination of pesticides in real vegetable samples from Almería (Spain).     

Development and validation of a new HPLC-UV method for the simultaneous determination of triclabendazole and ivermectin B1a in a pharmaceutical formulation

A rapid, simple, and sensitive RP-HPLC analytical method was developed for the simultaneous determination of triclabendazole and ivermectin in combination using a C18 RP column. The mobile phase was acetonitrile-methanol-water-acetic acid (56 + 36 + 7.5 + 0.5, v/v/v/v) at a pH of 4.35 and flow rate of 1.0 mL/min. A 245 nm UV detection wavelength was used. Complete validation, including linearity, accuracy, recovery, LOD, LOQ, precision, robustness, stability, and peak purity, was performed. The calibration curve was linear over the range 50.09-150.26 microg/mL for triclabendazole with r = 0.9999 and 27.01-81.02 microg/mL for ivermectin with r = 0.9999. Calculated LOD and LOQ for triclabendazole were 0.03 and 0.08 microg/mL, respectively, and for ivermectin 0.07 and 0.20 microg/mL, respectively. The intraday precision obtained was 98.71% with RSD of 0.87% for triclabendazole and 100.79% with RSD 0.73% for ivermectin. The interday precision obtained was 99.51% with RSD of 0.35% for triclabendazole and 100.55% with RSD of 0.59% for ivermectin. Robustness was also studied, and there was no significant variation of the system suitability of the analytical method with small changes in experimental parameters.     

Detection of melamine in milk by surface-enhanced Raman spectroscopy coupled with magnetic and Raman-labeled nanoparticles

A new method based on surface-enhanced Raman spectroscopy (SERS) has been developed for sensitive and rapid detection of melamine. Spherical magnetic-core gold-shell nanoparticles (AuNPs) and rod-shaped gold nanoparticles (nanorods) labeled with a Raman-active compound were used to form a complex with the melamine molecules. 5,5'-Dithiobis(2-nitrobenzoic acid) was used as Raman-active compound because it is readily adsorbed by a gold nanoparticle surface forming a self-assembled monolayer (SAM) and has strong Raman scattering at 1330 cm(-1), because of the symmetric NO(2) stretch. The calibration curve was obtained by plotting Raman band area at 1330 cm(-1) against melamine concentration. A linear relationship was obtained with a high determination coefficient (R(2)=0.997). The method was validated for linearity, sensitivity, precision (intra-day and inter-day repeatability), and recovery. In the model system, the limits of detection (LOD) and quantification (LOQ) were 0.38 and 1.27 mg L(-1), respectively. For melamine-spiked milk samples, LOD and LOQ values were 0.39 mg L(-1) and 1.30 mg L(-1), respectively. Intra and inter-day precision were 3.73 and 4.94 %, respectively. This method was applied to samples of skimmed milk that had been spiked with melamine at different concentrations. The recovery of the method was 95-109 % in the concentration range 2-15 mg L(-1), and average RSD was 1.71 %. Total analysis time was less than 15 min.