Determination of niacin in infant formula by solid-phase extraction and anion-exchange liquid chromatography.

A peer-verified, solid-phase extraction (SPE)/anion exchange liquid chromatographic method is presented for the determination of niacin in milk-based and soy-based infant formula. Analysis is in 3 steps: test sample digestion, extraction/cleanup, and liquid chromatography (LC). Digestion uses a standard AOAC digestion procedure that involves autoclaving at 121 degrees C for 45 min in (1 + 1) H2SO4 to free endogenous niacin from protein and to convert added niacinamide to niacin. The digest solution is adjusted to pH 6.5 with 7.5M NaOH. Acidification to pH <1.0 with (1 + 1) H2SO4 precipitates the protein. The clarified solution is then filtered, and the filtrate is brought to volume. SPE of niacin is accomplished by passing an aliquot of the digest solution through an aromatic sulfonic acid-SPE (ArSCX-SPE) column. After the column is washed with methanol and water to remove extraneous material, the niacin is eluted with 0.25M sodium acetate/acetic acid buffer at pH 5.6. An anion-exchange polystyrene-divinylbenzene column with 0.1 M sodium acetate/acetic acid buffer at pH 4.0 is used for LC. Niacin is determined by UV detection at 260 nm. A standard curve is prepared by passing known amounts of niacin through the ArSCX-SPE columns used for niacin extraction. The following values for x and relative standard deviation (RSD) were obtained for National Institute of Standards and Technology Standard Reference Material (NIST SRM) 1846 Infant Formula with a certified value for niacin of 63.3 +/- 7.6 microg/g: Submitting laboratory.-- x = 59.7 +/- 4.0 microg/g; RSD = >6.7%; confidence interval (CI) = +/- 1.4 microg/g; n = 27. Peer laboratory.--x = 56.6 +/- 6.6 microg/g; RSD = >11.7%; CI =+/- 4.1 microg/g; n = 8.     

Determination of niacin in food materials by liquid chromatography using isotope dilution mass spectrometry

The availability of deuterium-labeled nicotinic acid makes stable isotope dilution mass spectromerty (MS) coupled with liquid chromatography (LC) an attractive option for the determination of the water-soluble B-vitamin niacin in food samples. A method was developed based on acid digestion, solid-phase extraction with a strong cation exchange column, and reversed-phase chromatography with a C18 column. Detection is by positive ion electrospray MS. Analysis in selected ion recording mode is subject to interference problems similar to those found with other LC determinations of niacin, but the additional selectivity of multiple reaction monitoring mode largely eliminates interference problems. The method was applied to 6 different food matrixes and to appropriate reference materials, including milk samples with niacin levels near 1 ppm. The method exhibited good accuracy, based on levels obtained for the reference materials, and relative standard deviations in the range of 0.5-5%.     

Determination of niacin in infant formula by solid-phase extraction/liquid chromatography: peer-verified method performance-interlaboratory validation

This paper reports the results of the interlaboratory peer validation study of AOAC Peer-Verified Method (PVM) 1:2,000 for the determination of niacin in infant formula by solid-phase extraction/liquid chromatography. We have used a Data Quality Objectives (DQO) approach to address not only method variability and robustness but also accuracy of data through the use of an appropriate reference material in conjunction with the interlaboratory validation study. Our DQO included the following: (1) statistical agreement of analytical results and quantitative recovery between 2 collaborating laboratories; (2) the repeatability relative standard deviation (RSDr) values and the HORRAT (Horwitz ratio) obtained (1.07), which satisfied the criteria of the Horwitz "limits of acceptability" at the analyte level present; (3) validation of lack of interference; and (4) accuracy agreement within assigned values for a certified reference material. National Institute of Standards and Technology Standard Reference Material (NIST SRM) 1846 Infant Formula, with a certified value of 63.3 +/- 7.6 microg/g for niacin content, was used as a test material for collaborative study and accuracy assessment. Niacin values obtained by the originating laboratory were 59.7 +/- 4.0 microg/g (95% confidence interval [CI] = 1.4 microg/g with a relative standard deviation [RSD] of 6.7%) and by the peer laboratory were 56.6 +/- 6.6 microg/g (95% CI = 4.1 microg/g, with an RSD of 11.7%). Statistical evaluation using the means equivalence test showed that nicotinic acid values obtained by the peer laboratory were equivalent to those values obtained by the originating laboratory. Linear calibration curves and quantitative recovery were obtained. Integration of the PVM process with a readily available certified reference material gives the user confidence in the accuracy of the data generated by the method through traceability to the reference material used.     

固相萃取法与基质固相分散法在橙子中有机磷农药残留分析中的应用

建立了固相萃取(SPE)-反相高效液相色谱(RP-HPLC)同时测定橙子中痕量辛硫磷、二嗪农有机磷农药残留量的方法。样品经甲醇超声提取、C18固相萃取柱净化后,采用液相色谱柱分离,以乙腈-水(体积比为85∶15)为流动相等度洗脱,于254 nm下紫外检测。结果表明:在0.1～10.0 mg/L和0.4～10.0 mg/L范围内,辛硫磷、二嗪农的质量浓度与峰面积呈良好的线性关系;样品的加标平均回收率为87.3%～102.7%,相对标准偏差(RSD)为0.9%～4.9%。将该分析结果与用基质固相分散法(MSPD)处理样品所得的结果相比较,发现SPE对二嗪农的提取效果较好,而MSPD对辛硫磷的提取效果较好,但两种方法都能较好地净化样品,均能满足残留量的分析要求。     

Fast cleanup method for the analysis of Sudan I-IV and para red in various foods and paprika color (oleoresin) by high-performance liquid chromatography/diode array detection: focus on removal of fat and oil as fatty acid methyl esters prepared by transesterification of acylglycerols

A fast and effective cleanup method was developed for the analysis of Sudan I, II, III, IV, and Para Red (Sudan dyes) in various foods and paprika color (oleoresin) by high-performance liquid chromatography (LC) with a diode array detector (DAD). Removal of fat or oil in fatty sample was a critical point for reducing the volume of the final sample solution in order to obtain a sufficient level of the analytes. Separation of fat or oil from the dyes with a silica gel solid-phase extraction (SPE) column seemed unfeasible, because elution profiles of oil, fat, and the dyes were similar. Finally, fat and oil were separated from the dyes by elution from the SPE column with n-hexane, not as intact compounds but as fatty acid methyl esters prepared by direct transesterification of acylglycerols in fat and oil, leaving the dyes on the column. The dyes were eluted with n-hexane-diethyl ether (9 + 1). Gradient elution with water and tetrahydrofuran was used for separation on a C18 column by LC. Measurement of spectral of 0.5 microg/g of Sudan dyes in foods and 1 microg/g in paprika color (oleoresin) with the DAD was achieved.     

Determination of ochratoxin A in baby food by immunoaffinity column cleanup with liquid chromatography: interlaboratory study

An interlaboratory study funded by the European Commission, Standards, Measurement and Testing Programme (4th Framework Programme) was performed to evaluate the effectiveness of an immunoaffinity column cleanup liquid chromatographic (LC) method for the determination of ochratoxin A in baby food at a possible future European regulatory limit (0.1 ng/g). The test portion is extracted in a blender with tert-butyl methyl ether (chosen to avoid use of chloroform but shown to give equivalent extraction efficiency) after addition of 0.5 mol/L phosphoric acid-2 mol/L sodium chloride solution. The extract is centrifuged and redissolved in a mixture of phosphate buffered saline solution and methanol. After removal of lypophilic substances with hexane, the extract is applied to an immunoaffinity column containing antibodies specific to ochratoxin A. The column is washed with water to remove the interfering compounds and the purified ochratoxin A is eluted with methanol. The separation and determination of ochratoxin A is performed by reversed-phase LC and detected by fluorescence after postcolumn derivatization (PCD) with ammonia. Test materials (baby food infant formulae), both spiked and naturally contaminated with ochratoxin A, were sent to 13 laboratories in 8 different European countries. Test portions were spiked at a level of 0.085 ng/g ochratoxin A. The average recovery for the spiked blank baby food was 108%. Based on results for spiked samples (blind pairs at 0.085 ng/g) as well as naturally contaminated samples (blind pairs at levels between 0.05 and 0.22 ng/g) the relative standard deviation for repeatability (RSDr) ranged from 18-36%. The relative standard deviation for reproducibility (RSDR) ranged from 29-63% and HORRAT values of between 0.4 and 0.9 were obtained.     

Determination of halofuginone and amprolium in chicken muscle and egg by liquid chromatography

A liquid chromatographic (LC) method was developed for simultaneous measurement of halofuginone (HFN) and amprolium (APL) in chicken muscle and egg. HFN and APL were extracted from chicken muscle and egg with acetonitrile. In chicken egg, they were partially purified by solid-phase extraction (SPE) to separate them from impurities. The LC separation was performed on a 4.6 mm id x 250 mm TSK-gel ODS-80TM column using acetonitrile-McIlvaine buffer, pH 3.4, containing 0.01M sodium lauryl sulfate (42 + 58) as the mobile phase. Ultraviolet detection of HFN and APL was performed at wavelengths of 242 and 265 nm, respectively. Recoveries of HFN and APL from chicken muscle spiked at 0.5 microg/g were 74.8 +/- 17.7 and 94.2 +/- 5.0%, respectively (mean +/- standard deviation [SD], n = 10). In chicken muscle, the lower limit of determination for both APL and HFN was 0.03 microg/g. Recoveries of HFN and APL from chicken egg spiked at 0.5 microg/g by a cleanup procedure using SPE were 54.6 +/- 3.4 and 85.0 +/- 2.4%, respectively (mean +/- SD, n = 5). In chicken egg, the lower limit of determination for both APL and HFN was 0.04 microg/g.     

Determination of aflatoxin B1 in baby food (infant formula) by immunoaffinity column cleanup liquid chromatography with postcolumn bromination: collaborative study

A collaborative study was conducted to evaluate the effectiveness of an immunoaffinity column cleanup liquid chromatography (LC) method for determination of aflatoxin B, in a milk powder based infant formula at a possible future European regulatory limit (0.1 ng/g). The test portion was extracted with methanol-water (8 + 2 [v + v]), filtered, diluted with water, and applied to an immunoaffinity column. The column was washed with water to remove interfering compounds, and the purified aflatoxin B1 was eluted with methanol. The separation and determination of the aflatoxin B1 was performed by reversed-phase LC and detected by fluorescence after postcolumn derivatization (PCD) involving bromination. PCD was achieved with either pyridinum hydrobromide perbromide (PBPB) or an electrochemical (Kobra) cell by addition of bromide to the mobile phase. The baby food (infant formula) test samples, both spiked and naturally contaminated with aflatoxin B1, were sent to 14 laboratories in 13 different European countries. Test portions were spiked at levels of 0.1 and 0.2 ng/g for aflatoxin B1. Recoveries ranged from 101 to 92%. Based on results for spiked test samples (blind pairs at 2 levels) and naturally contaminated test samples (blind pairs at 3 levels), the relative standard deviation for repeatability (RSDr) ranged from 3.5 to 14%. The relative standard deviation for reproducibility (RSDR) ranged from 9 to 23%. Nine participants used PBPB derivatization, and     

Ultraviolet derivatization of steroidal saponin in garlic and commercial garlic products as p-nitrobenzoate for liquid chromatographic determination

A method is described for determination of the steroidal saponin, eruboside B, originating in garlic and garlic products as the p-nitrobenzoyl chloride (PNBC) derivative by reversed-phase liquid chromatography (with ultraviolet detection at 260 nm. Proto-eruboside B was extracted from garlic (Allium sativum L.); subjected to solid-phase extraction (SPE) with a C18 cartridge, Florisil column chromatography, and silica gel column chromatography; and then enzymatically converted to eruboside B, which was applied as an external standard. Steroidal saponins in garlic and commercial garlic products were extracted with methanol and purified by SPE cartridges, followed by enzymatic treatment. A frostanol saponin such as proto-eruboside B is enzymatically transformed to a spirostanol saponin, eruboside B. After the derivatization with PNBC, the saponin derivative was chromatographed on a C8 column with a gradient elution of (A) 80% aqueous acetonitrile and (B) 100% acetonitrile. The detection limit of the developed method was 1 microg/g for the samples. The method was applied to the analysis of garlic and garlic health food products available in Japan.     

Comparison of various dilutions and solid-phase extraction cleanup on the determination of ephedrine-type alkaloids and internal standard recovery in ephedra botanical raw material and powdered extract

A study was conducted to determine the effect of 3 dilution levels on the precision of the ephedra alkaloid method when used in conjunction with a solid-phase extraction (SPE) column. For the dilutions studied, SPE column cleanup is necessary because it promotes a greater recovery of the internal standard. However, overall, target precision values were not obtained on the test materials. It was determined that the SPE column is not the cause of the lower recovery in the more concentrated solutions. Significant signal suppression of the internal standard occurs in more concentrated solutions within the mass spectrometer. It is hypothesized that this lack of performance on the part of the SPE column may be linked to its inability to fully clean contaminants from the higher concentration solutions and/or a mass spectrometer overload, which resulted in the internal standard not fully correcting for signal suppression in more concentrated solutions. An internal standard is necessary, especially for accuracy, for the determination of all alkaloids, and only dilute solutions can be accurately analyzed. Due to the sensitivity of the mass spectrometer, it is recommended that the determination of ephedrine alkaloids in dietary supplements and botanicals should be studied using a standard graph at a level 10 x less than the current method. It is also recommended that the SPE column used should be evaluated on its need for the sample dilutions made to fit the newly recommended standard graph.     

Automated solid-phase extraction method for the determination of piperaquine in plasma by peak compression liquid chromatography

A validated bioanalytical method for the determination of piperaquine (PQ) in plasma by solid-phase extraction (SPE) and liquid chromatography (LC) using peak compression is presented. Protein is precipitated from plasma with acetonitrile-1% aqueous acetic acid (85:15, v/v). An internal standard (IS) is added to the samples before they are loaded onto a strong cation exchanger (Isolute PRS) SPE column. PQ and the IS are analyzed by LC on a Zorbax SB-CN column (250 x 4.0 mm) with the mobile phase acetonitrile-phosphate buffer [I = 0.1, pH 2.5 (12:88, v/v)] and UV detection at 345 nm. Trichloroacetic acid (TCA) is added to the samples prior to injection into the chromatography system. PQ elutes in a gradient of TCA, which enables peak compression of PQ and significantly higher peak efficiency as a result. The intraassay precision for plasma is determined to be 5.4% at 3.00 microM and 5.8% at 0.050 microM. The interassay precision for plasma is 1.3% at 3.00 microM and 10.0% at 0.050 microM. The lower limit of quantitation and the limit of detection are 0.025 and 0.005 microM, respectively.     

Direct determination of free methionine in soy-based infant formula

A method was developed for the direct determination of free methionine in soy-based infant formula, with analyte separation and quantitation by reversed-phase liquid chromatography (LC), and UV absorbance at 214 nm, respectively. Sample preparation required only dilution with mobile phase and syringe filtration. Using a 0.02M KH2PO4 mobile phase (pH adjusted to 2.9 with 85% o-phosphoric acid) and 0.7 mL/min flow rate, methionine eluted at approximately 8 min, and total run time was 14 min after column regeneration with acetonitrile-water. System linearity was demonstrated as peak area versus analyte concentration, ranging from 80 to 120% of the formula specification for free methionine (r > 0.999, and all residuals < 0.45%). Intermediate precision relative standard deviation values were < 1.5% for ready-to-feed and reconstituted powder samples, and recoveries ranged from 98.0 to 103.5% for inter-method comparison with an amino acid analyzer method. The limit of quantitation was 3 mg methionine/L in the "as fed" infant formula. Despite the relatively weak UV absorptivity of methionine, the 214 nm signal was sufficiently intense in the 30-65 mg/L (201-436 microM) range to afford quantitation by peak area proportionation versus a 2-point external standard calibration. This direct UV detection after reversed-phase LC separation provides a simple and accurate method for determining free methionine without derivatization.     

A simplified protein precipitation/mixed-mode cation-exchange solid-phase extraction, followed by high-speed liquid chromatography/mass spectrometry, for the determination of a basic drug in human plasma

A simplified protein precipitation/mixed-mode cation-exchange solid-phase extraction (PPT/SPE) procedure has been investigated. A mixture of acetonitrile and methanol along with formic acid was used to precipitate plasma proteins prior to selectively extracting the basic drug. After vortexing and centrifugation, the supernatants were directly loaded onto an unconditioned Oasis MCX microElution 96-well extraction plate, where the protonated drug was retained on the negatively charged sorbent while interfering neutral lipids, steroids or other endogenous materials were washed away. Normal wash steps were deemed unnecessary and not used before sample elution. The sample extracts were analyzed under both conventional and high-speed liquid chromatography/tandem mass spectrometry (LC/MS/MS) conditions to examine the feasibility of the PPT/SPE procedure for human plasma sample clean-up. For the conventional LC/MS/MS method, chromatographic separation was achieved on a C18, 2.1 x 50 mm column with gradient elution (k' = 5.5). The mobile phase contained 0.1% formic acid in water and 0.1% formic acid in acetonitrile. For the high-speed LC/MS/MS method, chromatographic separation was achieved on a C18, 2.1 x 10 mm guard column with gradient elution (k' = 2.2, Rt = 0.26 min). The mobile phase contained 0.1% formic acid in water and 0.001% trifluoroacetic acid in acetonitrile. Detection for both conventional and high-speed LC/MS/MS methods was by positive ion electrospray tandem mass spectrometry on a ThermoElectron Finnigan TSQ Quantum Ultra, where enhanced resolution (RP 2000; 0.2 amu) was used for high-speed LC/MS/MS. The standard curve, ranging from 0.5 to 100 ng/mL, was fitted to a 1/x weighted quadratic regression model.This combined PPT/SPE procedure effectively eliminated time-consuming sorbent conditioning and wash steps, which are essential for a conventional mixed-mode SPE procedure, but retained the advantages of both PPT (removal of plasma proteins) and mixed-mode SPE (analyte selectivity). The validation results demonstrated that this PPT/SPE procedure was well suited for both conventional and high-speed LC/MS/MS analyses. In comparison with a conventional mixed-mode SPE procedure, the simplified PPT/SPE process provided comparable sample extract purity. This simple sample clean-up procedure can be applied to other basic compounds with minor modifications of PPT solvents.     

Determination of zearalenone in barley, maize and wheat flour, polenta, and maize-based baby food by immunoaffinity column cleanup with liquid chromatography: interlaboratory study

An interlaboratory study was performed on behalf of the UK Food Standards Agency to evaluate the effectiveness of an affinity column cleanup liquid chromatography (LC) method for the determination of zearalenone (ZON) in a variety of cereals and cereal products at proposed European regulatory limits. The test portion is extracted with acetonitrile:water. The sample extract is filtered, diluted, and applied to an affinity column. The column is washed, and ZON is eluted with acetonitrile. ZON is quantified by reversed-phase LC with fluorescence detection. Barley, wheat and maize flours, polenta, and a maize-based baby food naturally contaminated, spiked, and blank (very low level) were sent to 28 collaborators in 9 European countries and 1 collaborator in New Zealand. Participants were asked to spike test portions of all samples at a ZON concentration equivalent to 100 microg/kg. Average recoveries ranged from 91-111%. Based on results for 4 artificially contaminated samples (blind duplicates) and 1 naturally contaminated sample (blind duplicate), the relative standard deviation for repeatability (RSDr) ranged from 6.9-35.8%, and the relative standard deviation for reproducibility (RSDR) ranged from 16.4-38.2%. The method showed acceptable within- and between-laboratory precision for all 5 matrixes, as evidenced by HorRat values <1.7.     

2D LC Separation and Determination of Bradykinin in Rat Muscle Tissue Dialysate with On-Line SPE-HILIC-SPE-RP-MS

A 2D liquid chromatography (LC) system using hydrophilic interaction chromatography (HILIC) and reversed phase columns has been employed for comprehensive (LC × LC) separation of rat muscle tissue micro-dialysate. Incorporation of an on-line reverse-phase solid phase extraction (SPE) enrichment column in front of the first dimension enabled aqueous samples with high salt concentrations to be injected directly without compromising the chromatographic performance of the HILIC column. Since the SPE enrichment column allowed injection of large sample volumes (e.g. 450 μL), a capillary HILIC column (inner diameter 0.3 mm) could be employed instead of a larger column which is often used in the first dimension to load sufficient amounts of sample. The two chromatographic dimensions were connected using a column selector system with 18, 1.0 mm I.D. C₁₈ “transition” SPE columns. A PLRP C₁₈ column was used in the second dimension. The 2D LC system’s performance was evaluated with a tryptic digest mixture of three model proteins. Good trapping accuracy (HILIC→transition SPE→RP recovery >95%) and repeatability (within-and between day retention time RSDs of first and second dimension chromatography >1%) was achieved. A dialysis sample of rat muscle tissue was separated with the 2D system, revealing complexity and large differences in concentrations of the various compounds present, factors which could potentially interfere with the quantification and monitoring of two target analytes, arg-bradykinin and bradykinin. Subsequently, “Heart-cut” 2D LC-electrospray–mass spectrometry (ESI–MS) with post-column on-line standard injection was employed to monitor arg-bradykinin and bradykinin levels as a function of various muscle conditions. The method’s quantification precision was RSD = 3.4% for bradykinin.     

Matrix effects in the determination of β‐receptor agonists in animal‐derived foodstuffs by ultra‐performance liquid chromatography tandem mass spectrometry with immunoaffinity solid‐phase extraction

Matrix effects in determination of three β‐receptor agonists including salbutamol (SAL), clenbuterol, and terbutaline in animal‐derived foodstuffs were studied by ultra‐performance LC‐MS/MS with cleanup of immunoaffinity SPE column (IAC). Some animal tissue samples including pig liver, swine muscle, and fish muscle were hydrolyzed by the mixed enzyme solution or HCl solution, and the cleanup efficiencies with SAL IAC, MCX SPE column, and C₁₈‐SCX tandem columns were examined and compared by using spiked experiments. The results showed that the matrix effects in the determination of SAL and terbutaline can be eliminated with SAL IAC cleanup, and the average recoveries of SAL were 77.4～81.5%, 79.0～80.3%, and 85.0～87.2% in pig liver, swine muscle, and fish muscle, respectively. The decision limit (ccα) and detection capability (ccβ) for SAL in pig liver were 0.02 and 0.05 μg/kg, respectively.     

Fast sample preparation involving MASE and coupled column normal phase liquid chromatography for the rapid trace analysis of dioxins in air-dust samples from fire catastrophe emissions

A new approach has been developed and tested for the urgent analysis of dioxins in samples of air-dust filters originating from catastrophe emissions. The procedure consists of a fast extraction of the sample with microwave solvent extraction (MASE) and acetone as solvent followed by a fast cleanup of the extract with normal phase coupled column liquid chromatography (LC/LC).

The multi-dimensional LC/LC system employs a 50 mm×4.6 mm i.d. column packed with 3 μm silica and a 150 mm×4.6 mm i.d. column packed with 5 μm PYE as the first and second analytical column, respectively. Iso-hexane is used on both columns to perform cleanup and dichloromethane to perform efficient back-flush elution of the compounds from the second column. The obtained polarity-based separation in the first dimension and molecular-structure based separation in the second dimension provides a fast and powerful cleanup.

Validation was done by analysing samples of homemade RIVM air-dust with aged residues (n=8, spiking level about 15 pg mg⁻¹ per compound) of dioxins/furans and samples of reference Urban Dust SRM 1649a (n=4) with both the new approach and the existing conventional procedure and were instrumentally analyzed with capillary gas chromatography and high resolution mass spectrometric detection (GC/HRMS).

In comparison to the existing conventional procedure, the new approach reduces sample processing from several days to several hours per sample.

As regards the aged-residue air-dust samples, the new method shows a good accuracy, precision and high selectivity providing a performance in good agreement with the existing procedure. In SRM air-dust, the concentration of a few compounds obtained by the new method was below (10–50%) the certified value.     

Interlaboratory study of a multiresidue gas chromatographic method for determination of organochlorine and pyrethroid pesticides and polychlorobiphenyls in milk,fish, eggs,and beef fat

An interlaboratory study was conducted to validate a gas chromatographic (GC) method for determination of 21 organochlorine pesticides, 6 pyrethroid pesticides, and 7 polychlorobiphenyl (PCB) congeners in milk, beef fat, fish, and eggs. The method was performed at low contamination levels, which represent relevant contents in food, and is an extension of the European standard (method NF-EN-1528, Parts 1-4). It enlarges the applicable scope of the reference EN method to pyrethroid pesticides and proposes the use of solid-phase extraction (SPE) as a cleanup procedure. Cryogenic extraction was made, and SPE cleanup was performed with 2 successive SPE cartridges: C18 and Florisil. After injection of the purified extract onto a GC column, residues were measured by electron capture detection. Food samples (liquid milk, beef fat, mixed fish, and mixed eggs) were prepared, tested for homogeneity, and sent to 17 laboratories in France. Test portions were spiked with 27 pesticides and 7 PCBs at levels from 26 to 45, 4 to 27, 31 to 67, and 19 to 127 ng/g into milk, eggs, fish, and fat, respectively. Based on results for spiked samples, the relative standard deviation for repeatability ranged from 1.5 to 6.8% in milk, 3 to 39% in eggs, 4.5 to 12.2% in fish, and 7 to 13% in fat. The relative standard deviation for reproducibility ranged from 33 to 50% in milk, 29 to 59% in eggs, 31 to 57% in fish, and 30 to 62% in fat. This method showed acceptable intra- and interlaboratory precision data, as corroborated by HORRAT values at low levels of pesticide and PCB contamination. The statistical evaluation of the results was performed according to the International Organization for Standardization (ISO; ISO 3534 standard) and 5725-2 Guideline.     

Determination of seventeen polar/thermolabile pesticides in apples and apricots by liquid chromatography/mass spectrometry

A simple liquid chromatography/mass spectrometry (LC/MS) approach for the determination of widely used representatives of polar/thermolabile pesticides in fruits was developed and validated. The group of pesticides comprised benzimidazoles and azoles (carbendazim, thiabendazole, imazalil, propiconazole, prochloraz, epoxiconazole, flusilazole, tebuconazole, bitertanol); N-methylcarbamates (carbaryl, carbofuran, methiocarb); and phenylureas and benzoylphenylureas (linuron, diflubenzuron, triflumuron, teflubenzuron, flufenoxuron). Matrixes (apple, apricot) were extracted with acetonitrile and crude extracts were cleaned up by solid-phase extraction (SPE) using either mixed cation exchange or hydrophilic lipophilic balance cartridges. LC separation of pesticides was performed on a reversed-phase column, Discovery C18. Electrospray ionization and ion trap MS/MS detection were applied. For most pesticides, overall recoveries ranged from 75 to 122%, and repeatability (as relative standard deviation) from 5 repetitive determinations of recovery ranged from 3 to 21%. Carbofuran was the only compound for which recovery was not satisfactory due to its loss in the SPE cleanup step. Limits of detection were 0.1-3 microg/kg for benzimidazole and azole fungicides and carbamate insecticides. For urea insecticides, detection limits were slightly higher (3-10 microg/kg).     

Determination of residues of azamethiphos in salmon tissue by liquid chromatography with fluorescence detection.

A liquid chromatographic (LC) method with fluorescence detection (FLD) is described for determining residues of the pesticide azamethiphos (AZA) in salmon tissue. The sample is extracted with ethyl acetate, centrifuged, dehydrated with anhydrous sodium sulfate, evaporated, reconstituted in water, and defatted with hexane. The aqueous phase is passed through a C18 solid-phase extraction (SPE) column. The SPE column is eluted with methanol, and the eluate is evaporated to dryness and then taken up in 10% acetonitrile (ACN) in water. The analyte is determined by LC using a C18 column, ACN-H2O (32 + 68) mobile phase, and FLD with excitation at 230 nm and emission at 345 nm. Composited salmon tissues were fortified with AZA at 5, 10, 21, 42, and 83 ng/g or ppb (target level, X = 10 ng/g). Overall recoveries were 86%, with between-day variability of 5.3%. The method detection limit was calculated as 1.2 ppb AZA based on a 5 g sample. The limit of quantitation as determined empirically by this method is the lower limit of the standard curve, approximately 5 ppb.