高效液相色谱法分析牛和猪肌肉组织中残留的安乃近药物的三种代谢物

建立了牛和猪肌肉组织中残留的安乃近药物的3种代谢物 4-甲酰氨基安替比林（FAA）、4-氨基安替比林（AA）和 4-甲基氨基安替比林（MAA）的高效液相色谱测定法。肌肉组织样品均质后采用硫酸钠-亚硫酸钠溶液提取,过滤后经C18固相萃取柱净化,采用Inertsil ODS-3色谱柱分离,甲醇和水梯度洗脱,于265 nm波长下检测,外标法定量FAA,内标法定量AA和MAA（以 4-异丙基氨基安替比林作为内标物）。FAA的检出限为12.5 μg/L,AA的检出限为15.0 μg/L,MAA的检出限为20.0 μg/L;3种代谢物的测定低限均为50 μg/L;在添加水平为50～400 μg/kg范围内,FAA的回收率为81.3%~92.5%,AA的回收率为82.0%~96.0%,MAA的回收率为80.4%~90.6%,相对标准偏差均在7%以内。     

Analysis of toldimfos in porcine muscle and bovine milk using liquid chromatography–triple quadrupole mass spectrometry

An analytical method was developed for the detection of toldimfos sodium residues in porcine muscle and bovine milk using liquid chromatography–triple quadrupole tandem mass spectrometry (LC–MS/MS) analysis. The drug was extracted from muscle and milk using 10 mm ammonium formate in acetonitrile and then purified using n ‐hexane. The drug was well separated on a Luna C₁₈ column using a mixture of 10 mm ammonium formate in ultrapure water (A) and acetonitrile (B) as the mobile phase. Good linearity was achieved over the tested concentration range (0.005–0.03 mg/kg) in matrix‐matched standard calibration. The determination coefficients (R ² ) were 0.9942 and 0.9898 for muscle and milk, respectively. Fortified porcine muscle and bovine milk contained concentrations equivalent to and twice the limit of quantification (0.005 mg/kg) yielded recoveries in the range of 75.58–89.74% and relative standard deviations of ≤8.87%. Samples collected from large markets located in Seoul, Republic of Korea, tested negative for toldimfos sodium residue. In conclusion, ammonium formate in acetonitrile can effectively extract toldimfos sodium from porcine muscle and bovine milk without solid‐phase extraction, which is usually required for cleanup before analysis. This method can be applied for the routine analysis of toldimfos in foods of animal origins.     

Validated method for determination of ultra-trace closantel residues in bovine tissues and milk by solid-phase extraction and liquid chromatography-electrospray ionization-tandem mass spectrometry

A liquid chromatographic-electrospray ionisation-tandem mass spectrometry method (LC-ESI-MS/MS) with solid extraction was developed and validated for the detection and determination of closantel residues in bovine tissues and milk. An acetonitrile-acetone mixture (80:20, v/v) was used for one-stage extraction of closantel residues in bovine tissues and milk samples, and the extract was cleaned by solid phase extraction with Oasis MAX cartridges. The mass spectrometer was operated in multiple reactions monitoring mode with negative electrospray interface. The limits of detection in different matrices were in the range of 0.008-0.009 microg/kg. The overall recoveries for bovine muscle, liver, kidney and milk samples spiked at four levels including MRL were in the range of 76.0-94.3%. The overall relative standard deviations were in the range of 3.57-8.61%. The linearity is satisfactory with a correlation coefficient (r(2)) of 0.9913-0.9987 at both concentration ranges of 0.02-100 microg/kg and 200-5000 microg/kg. The method is capable of identifying closantel residues at > or =0.02 microg/kg levels and was applied in the determination of closantel residues in animal origin foods.     

Multilaboratory trial for determination of ceftiofur residues in bovine and swine kidney and muscle,and bovine milk

A multilaboratory trial for determining ceftiofur-related residues in bovine and swine kidney and muscle, and bovine milk was conducted following regulatory guidelines of the U.S. Food and Drug Administration, Center for Veterinary Medicine. The methods convert all desfuroylceftiofur-related residues containing the intact beta-lactam ring to desfuroylceftiofur acetamide to establish ceftiofur residues in tissues. Four laboratories analyzed 5 sets of samples for each tissue. Each sample set consisted of a control/blank sample and 3 control samples fortified with ceftiofur at 0.5 Rm, Rm, and 2 Rm, respectively, where Rm is the U.S. tolerance assigned for ceftiofur residue in each tissue/matrix: 0.100 microg/mL for milk, 8.0 microg/g for kidney (both species), 1.0 microg/g for bovine muscle, and 2.0 microg/g for swine muscle. Each sample set also contained 2 samples of incurred-residue tissues (one > Rm and one < Rm) from animals treated with ceftiofur hydrochloride. All laboratories completed the method trial after a familiarization phase and test of system suitability in which they demonstrated > 80% recovery in pretrial fortified test samples. Results showed that the methods met all acceptable performance criteria for recovery, accuracy, and precision. Although sample preparation was easy, solid-phase extraction cartridge performance must be carefully evaluated before samples are processed. The liquid chromatography detection system was easily set up; however, the elution profile may require slight modifications. The procedures could clearly differentiate between violative (> Rm) and nonviolative (< Rm) ceftiofur residues. Participating laboratories found the procedures suitable for ceftiofur residue determination.     

Validation of a Biacore method for screening eight sulfonamides in milk and porcine muscle tissues according to European decision 2002/657/EC

Sulfonamides are commonly used for prophylactic or therapeutic purposes in veterinary medicine. A maximum residue limit (MRL) for sulfonamides has been set at 100 microg/kg in milk and muscle. A multisulfonamide antibody was used for the development of 2 different Biacore protocols, one for the screening of milk samples, the other for muscle samples. Two different Biacore systems were used: Biacore X system (milk protocol), which is considered a research and development apparatus, and Biacore 3000 system (muscle protocol), which is a completely automated system used for high-throughput screening. This report describes the validation of semiquantitative immunological methods according to the European Decision 2002/657/EC "concerning the performance of analytical methods." The different performance characteristics (detection capability CCbeta, specificity/selectivity, precision, stability, and applicability) were determined in relation to the European Union MRL of 100 microg/kg for sulfonamides. The applicability of the method to porcine, bovine, and poultry muscle was studied. The detection capabilities CCbeta were calculated to be 40 microg/L in milk and 60 microg/kg in porcine, bovine, and poultry muscles. Eight different sulfonamides, of which 3 (sulfamethazine, sulfamerazine, and sulfadiazine) are authorized in France, were detected simultaneously, at or below the MRL level, with both Biacore systems.     

Validation of a high-performance liquid chromatography method for the determination of oxytetracycline,tetracycline, chlortetracycline and doxycycline in bovine milk and muscle

High-performance liquid chromatography with diode-array detection (HPLC-DAD) was optimised and validated for the determination of tetracyclines in bovine milk and tissues. Milk and tissue samples were extracted and purified using a solid-phase extraction HLB Oasis cartridge and analysed using HPLC-DAD set at 365 nm. The analyses were carried out using the mobile phase of 0.01 M oxalic acid-acetonitrile-methanol (60:25:15, v/v/v) on a C8 column (250 x 4.6 mm I.D., 5 microm). Recoveries of tetracyclines from spiked samples at the three concentrations (0.5, 1 and 1.5) of the maximum residues limits (corresponding to 100 microg/kg for milk and the muscle) were higher than 81.1% in milk and 83.2% in muscle. The method was successfully validated for bovine milk and muscle in compliance with requirements set by draft SANCO/ 1805/ 2000 European Decision. The decision limit (CCalpha) was in the range 113.2-127.2 microg/kg and 107.7-129.9 micro/kg for all compounds in milk and muscle, respectively. The detection capability (CCbeta) was in the range 117.2-131.3 microg/kg and 114.9-133.1 microg/kg for all compounds in milk and muscle, respectively.     

Determination of chloramphenicol residues in meat, seafood, egg, honey, milk, plasma and urine with liquid chromatography-tandem mass spectrometry, and the validation of the method based on 2002/657/EC

A simple and rapid method for the determination and confirmation of chloramphenicol in several food matrices with LC-MS/MS was developed. Following addition of d5-chloramphenicol as internal standard, meat, seafood, egg, honey and milk samples were extracted with acetonitrile. Chloroform was then added to remove water. After evaporation, the residues were reconstituted in methanol/water (3+4) before injection. The urine and plasma samples were after addition of internal standard applied to a Chem Elut extraction cartridge, eluted with ethyl acetate, and hexane washed. Also these samples were reconstituted in methanol/water (3+4) after evaporation. By using an MRM acquisition method in negative ionization mode, the transitions 321-->152, 321-->194 and 326-->157 were used for quantification, confirmation and internal standard, respectively. Quantification of chloramphenicol positive samples regardless of matrix could be achieved with a common water based calibration curve. The validation of the method was based on EU-decision 2002/657 and different ways of calculating CCalpha and CCbeta were evaluated. The common CCalpha and CCbeta for all matrices were 0.02 and 0.04 microg/kg for the 321-->152 ion transition, and 0.02 and 0.03 microg/kg for the 321-->194 ion transition. At fortification level 0.1 microg/kg the within-laboratory reproducibility is below 25%.     

Determination of abamectin, doramectin, emamectin, eprinomectin, ivermectin, and moxidectin in milk by liquid chromatography electrospray tandem mass specrometry

The avermectin and milbemycin families of compounds are derived from naturally occurring yeasts. They have proven to be potent preventatives against a variety of pests such as insects and parasites. Only eprinomectin and moxidectin are currently approved for use on lactating cattle with tolerances in milk of 12 microg/kg for eprinomectin and 40 microg/kg for moxidectin. Detection of misuse or inadvertent contamination in milk requires a sensitive and definitive analytical method. A method has been developed for the determination of 5 avermectins and 1 milbemycin in milk using a simple liquid-liquid extraction and liquid chromatography/tandem mass spectrometry (LC/MS/MS) analysis. Ivermectin (IVR), doramectin (DOR), abamectin (ABA), eprinomectin (EPR), emamectin (EMA), and moxidectin (MOX) were extracted from whole milk by partitioning into acetonitrile with a subsequent solvent exchange into methanol-water. Simultaneous confirmation and quantification were achieved with LC separation, positive electrospray ionization (ESI+), and MS/MS. The limits of detection ranged from 16 pg/g (ppt) for EMA to 1.7 microg/g (ppb) for MOX.     

Multiresidue determination of zeranol and related compounds in bovine muscle by gas chromatography/mass spectrometry with immunoaffinity cleanup

A gas chromatography/mass spectrometry (GC/MS) method with immunoaffinity cleanup was developed for the determination of zeranol and related compounds, taleranol, zearalanone, and alpha-zearalenol in bovine muscle. Muscle samples were extracted with methanol and cleaned up with immunoaffinity chromatography (IAC) columns containing monoclonal antibodies raised against zeranol coupled to CNBr-activated Sepharose 4B. After derivatization, the compounds were analyzed by GC/MS. The dynamic column capacities for zeranol, taleranol, zearalanone, and alpha-zearalenol were 2639.7, 2840.3, 2731.5, and 2736.3 ng/mL Sepharose gel, respectively. The limits of detection and quantification were 0.5 and 1.0 ng/g, respectively, for all 4 compounds. Mean recoveries were 79.6-110.7% with coefficients of variation of 3.2-11.4% at spiked levels of 1.0-5.0 ng/g. This IAC-GC/MS method may be used for the determination of zeranol, taleranol, zearalanone, and alpha-zearalenol residues in bovine muscle, and possibly other tissues.     

Determination of four nitroimidazoles in poultry and swine muscle and eggs by liquid chromatography/tandem mass spectrometry

A rapid method is presented for the determination of 4 nitroimidazoles in poultry and swine muscle and eggs by liquid chromatography/electrospray ionization-tandem mass spectrometry (LC/ESI-MS/MS). Samples were extracted with acetonitrile, then evaporated the organic phase. After filtration, the extract was directly injected into the LC/ESI-MS/MS system. The LC separation was made on a C8 column by applying a gradient composed of water and acetonitrile. Overall average recoveries ranged from 50-86% for egg, 66-115% for poultry muscle, and 79-111% for swine muscle. The decision limits ranged from 0.05-0.25 microg/kg for egg, 0.07-0.27 microg/kg for poultry muscle, and 0.07-0.26 microg/kg for swine muscle; and detection capabilities ranged from 0.51-0.68 microg/kg for egg, 0.41-0.75 microg/kg for poultry muscle, and 0.53-0.78 microg/kg for swine muscle.     

Determination of tetracycline antibiotics in cow's milk by liquid chromatography/tandem mass spectrometry

A liquid chromatographic/tandem mass spectrometric (LC/MS/MS) multiresidue method for the simultaneous quantitative determination of oxytetracycline, 4-epi-oxytetracycline, tetracycline, 4-epi-tetracycline, chlortetracycline, 4-epi-chlortetracycline and doxycycline in milk has been developed. An extraction procedure consisting of a liquid extraction of the milk samples with trichloroacetic acid was performed. The extract was centrifuged and the supernatant was filtered. Solid-phase extraction (SPE) with an OASIS HLB SPE column was used to clean up the sample extracts. The samples were analysed by LC/MS/MS. The LC separation was performed on a reversed-phase C18 column using gradient elution with a mobile phase consisting of water and a mixture of methanol/acetonitrile. The tetracycline analytes were detected with a quadrupole mass spectrometer using positive ion electrospray ionisation. The confirmatory method has acceptable detection limits and the different tetracyclines can be detected at a residue concentration between 5 and 20 microg/L. The method is validated according to the European requirements for veterinary drug residues and all determined parameters were found to conform to the criteria. The recovery values ranged from 90.4 to 101.2% with relative standard deviations (RSDs) no larger than 9.7%. The overall or between-day precision of the analytical assay determined as repeatability at several residue concentrations and expressed as RSD ranged from 3.3 to 10%. This analytical assay is a useful tool within the Belgian monitoring programme for confirmation of samples which have been positively screened for residues of tetracyclines in raw farm cow's milk.     

Determination and confirmation of tulathromycin residues in bovine liver and porcine kidney via their common hydrolytic fragment using high-performance liquid chromatography/tandem mass spectrometry

An accurate, reliable, and reproducible analytical method using HPLC/MS/MS for the determination of tulathromycin residues in bovine liver and porcine kidney via their common hydrolytic fragment (CP-60,300) was developed and validated. Briefly, the method involved an initial acid treatment of intact tissues, which yielded the common fragment (CP-60,300). A portion of the acid hydrolyzate was purified by SPE using a strong cation exchange cartridge. Evaporation of the purified extract was followed by reconstitution in aqueous buffer and analysis by HPLC/MS/MS under isocratic conditions. The developed method provided acceptable sensitivity for determinative surveillance of tulathromycin in porcine kidney and bovine liver with an LOQ of 7.50 and 2.75 microg/g, respectively. The overall recovery and precision of 45 determinations of each tissue were 97.8% (5.3%) for porcine kidney and 96.9% (7.9%) for bovine liver. Accuracy, precision, linearity, specificity, and ruggedness were demonstrated. An HPLC/MS/MS method was also developed for use in these tissues as a confirmatory assay following modifications to the MS detection parameters. The confirmatory method demonstrated acceptable sensitivity for confirmatory evaluation of tulathromycin in porcine kidney and bovine liver at tolerances of 15 and 5.5 microg/g, respectively.     

Confirmation of sulfamethazine, sulfathiazole, and sulfadimethoxine residues in condensed milk and soft-cheese products by liquid chromatography/tandem mass spectrometry

A liquid chromatography/tandem mass spectrometry method (LC/MS/MS) is described for the simultaneous detection of 3 sulfonamide drug residues at 1.25 ppb in condensed milk and soft-cheese products. The 3 sulfonamide drugs of interest are sulfathiazole (STZ), sulfamethazine (SMZ), and sulfadimethoxine (SDM). The method includes extraction of the product with phosphate buffer, centrifugation of the diluted product, and application of a portion of the extract onto a polymeric solid-phase extraction cartridge. The cartridge is washed with water, and the sulfonamides are eluted with methanol. After evaporation, the residue is dissolved in 0.1% formic acid solution, and the solution is filtered before analysis by LC/MS/MS. The LC/MS/MS program involved a series of time-scheduled selected-reaction monitoring transitions. The transitions of MH+ to the common product ions at m/z 156, 108, and 92 were monitored for each residue. In addition, SMZ and SDM had a fourth significant and unique product ion transition that could be measured. Validation was performed with control and fortified-control condensed bovine milk with 2.5, 5, and 10 ppb sulfonamides. This method was applied to imported flavored and unflavored condensed milk and cream cheese bars. The presence of STZ and SMZ residues was confirmed in 3 out of 6 products.     

Development and validation of a liquid chromatographic/ tandem mass spectrometric method for determination of chlortetracycline, oxytetracycline, tetracycline, and doxycycline in animal feeds

A selective and accurate LC/MS/MS method for the simultaneous determination of chlortetracycline (CTC), oxytetracycline (OTC), tetracycline (TC), and doxycycline (DC) in animal feeds was developed. Samples were extracted with Na2EDTA-McIlvaine buffer and further purified with Oasis HLB SPE columns. The purified extract was separated on an Xbridge C18 column and detected by LC/MS/MS with positive electrospray ionization in the multiple reaction monitoring mode. This method provided average recoveries of 80.9 to 119.5%, with CVs of 1.7 to 9.8% in the range of 0.5 to 50 mg/kg CTC, OTC, TC, and DC in feeds, except the average recovery of CTC was 76.0%, with a CV of 14.6% in pig feed spiked with 0.5 mg/kg CTC. The linear ranges for the four TCs determined by LC/MS/MS ranged from 0.005 to 2.5 microg/mL with a linear correlation coefficient (R2) >0.99. The LOD and LOQ for CTC, OTC, TC, and DC in pig and poultry feeds ranged from 0.003 to 0.02 and 0.01 to 0.05 microg/g, respectively. The method was successfully applied for the analysis of 30 real feed samples, and no illegal use was detected.     

An analytical method to screen for six thyreostatic drug residues in the thyroid gland and muscle tissues of food producing animals by liquid chromatography with ultraviolet absorption detection and liquid chromatography/mass spectrometry

A method was developed and validated to screen for residues of the thyreostatic drugs, tapazole (TAP), mercaptobenzimidazole (MBI), thiouracil (TU), methylthiouracil (MTU), propylthiouracil (PrTU), and phenylthiouracil (PhTU) in bovine, equine, ovine, and porcine thyroid and muscle tissues at concentrations > or = 5 ng/g using 2-methoxy-mercaptobenzimidazole (MeMBI) and dimethylthiouracil (DMTU) as internal standards. In this method, the drugs were solvent extracted from thyroid and muscle tissue and cleaned up on an amino-propyl solid-phase extraction (SPE) cartridge. The unretained fraction containing TAP and MBI and the internal standard, MeMBI, was collected as Fraction 1. The retained fraction containing TU, MTU, PrTU, PhTU, and the internal standard, DMTU, was eluted with 3% acetic acid-isopropanol as Fraction 2. Fraction 1 was further cleaned up on an alumina B SPE cartridge and analyzed by gradient elution on a C18 high-performance liquid chromatography (HPLC) column with ultraviolet detection at wavelengths of 255 and 300 nm. Fraction 2 was taken to dryness, derivatized with 4-chloro-7-nitrobenzo-2-furazan at pH 8, and analyzed by gradient elution on a C18 LC column with mass spectrometry (MS) detection. Any "presumptive positive" test results were submitted for further analysis by LC/MS/MS. The validated method was applied to the analysis of over 300 thyroid tissue samples.     

Alternative methodology for the analysis of progesterone, testosterone, and epi-testosterone in bovine liver and veal muscle

Research has shown that traditional solvent extraction procedures used for the analysis of endogenous steroids often give inconsistent recoveries and results. However, a single-laboratory validation of a liquid chromatography/tandem mass specrometry method using 2 product ions per transition for progesterone, testosterone, and epi-testosterone in bovine liver and veal muscle showed accuracy and precision to within 23% at concentrations ranging from 0.5 to 2.0 microg/kg. Homogenized samples were pretreated with methanol to denature endogenous enzymes. Following removal of methanol, samples were treated overnight with Helix pomatia beta-glucuronidase to deconjugate glucuronide conjugates. Alkali digestion of the samples in KOH solutions was done under shaking at 37 degrees C for 30 min. The digestate was extracted with methyl tert-butyl ether, and the extracts were cleaned by partitioning between acetonitrile-hexane, followed by solid-phase extraction cleanup on silica cartridges. In bovine liver, average recoveries exceeded 54% for all analytes, and the within-run assay coefficients of variations were < 6 and 13% for high (2.0 microg/kg) and low (0.3 microg/kg) analyte concentrations, respectively. In veal muscle, average recoveries exceeded 60%, and the analysis of blind spikes gave accuracy estimates of over 85%, with coefficients of variation (CVs) < 15% for all analytes. The CVs for the multiple reaction monitoring ion ratios for all compounds were < 22% for all validation data. The method meets the requirements for confirmatory methods as outlined in 2002/657/EC. An analyst is capable of processing up to 20 samples within 5 days.     

Simultaneous determination of spinosad,temephos, and piperonyl butoxide in animal‐derived foods using LC–MS/MS

Pesticides, which are used as plant protection products, can enter the food chain, and exposure to these xenobiotics can cause a wide array of health problems in humans. Therefore, the objective of the present study was to develop an analytical method for the simultaneous determination of residual spinosad (sum of spinosyn A and D), temephos and piperonyl butoxide in porcine muscle, egg, milk, eel, flatfish and shrimp (sampling period: February to June 2018) using liquid chromatography–triple quadrupole tandem mass spectrometry (LC–MS/MS). The target analytes were extracted with a combination of acidified acetonitrile and ethyl acetate and subsequently purified with original QuEChERS kits (composed of magnesium sulfate and sodium chloride) as well as n‐hexane. All analytes were separated on a reversed‐phase analytical column using a mobile phase of (A) 0.1% formic acid containing 10 mm ammonium formate in distilled water and (B) methanol. Good linearity (R² ≥ 0.980) was achieved over the tested concentration range (3.5–35 μg/kg for spinosyn A; 1.5–15 μg/kg for spinosyn D; 5–50 μg/kg for temephos and piperonyl butoxide) in matrix‐matched standard calibrations. Fortified samples at three spiking levels yielded recoveries in the range of 71–105% with relative standard deviations ≤9.2%. The applicability of the method was evaluated via evaluating samples collected from a large wholesale market located in Seoul, and none of the samples contained any of the target analytes. In conclusion, the current approach is simple, efficient and reliable and can successfully determine the residual levels of spinosad, temephos and piperonyl butoxide in complex animal‐derived food products.     

Determination of chloramphenicol residues in milk, eggs, and tissues by liquid chromatography/mass spectrometry

A new liquid chromatography/mass spectrometry (LC/MS) method is presented for the determination of chloramphenicol (CAP) residues in milk, eggs, chicken muscle and liver, and beef muscle and kidney. CAP is extracted from the samples with acetonitrile and defatted with hexane. The acetonitrile extracts are then evaporated, and residues are reconstituted in 10mM ammonium acetate--acetonitrile mobile phase and injected into the LC system. CAP is determined by reversed-phase chromatography using an Inertsil ODS-2 column and MS detection with negative ion electrospray ionization. Calibration curves were linear between 0.5-5.0 ng/g for all matrixes studied. The relative standard deviations for measurements by this method were generally <12%, and average recoveries ranged from 80 to 120%, depending on the matrix involved. The method detection limits of CAP ranged from 0.2 to 0.6 ng/g, which are comparable to previously reported results. The proposed method is rapid, simple, and specific, allowing a single analyst to easily prepare over 40 samples in a regular working day.     

Screening and quantitative confirmatory method for the analysis of glucocorticoids in bovine milk using liquid chromatography-tandem mass spectrometry

An LC/MS/MS method was developed and validated for the simultaneous identification, confirmation, and quantification of 12 glucocorticoids in bovine milk. The method was validated in accordance with the criteria defined in Commission Decision 2002/657/EC. The developed method can detect and confirm the presence of dexamethasone, betamethasone, prednisolone, flumethasone, 6alpha-methylprednisolone, fluorometholone, triamcinolone acetonide, prednisone, cortisone, hydrocortisone, clobetasol propionate, and clobetasol butyrate in bovine milk. Milk samples are extracted with acetonitrile; sodium chloride is subsequently added to aid partition of the milk and acetonitrile mixture. The acetonitrile extract is then subjected to liquid-liquid purification by the addition of hexane. The purified extract is evaporated to dryness and reconstituted in a water-acetonitrile mixture, and determination is carried out by LC/MS/MS. The method permits analysis of up to 30 samples in 1 day.     

A quantitative method for residues of macrolide antibiotics in porcine kidney by liquid chromatography/tandem mass spectrometry

An LC/MS/MS-based multiresidue quantitative method was developed for the macrolides erythromycin A, neospiramycin I, oleandomycin, spiramycin I, tilmicosin, and tylosin A in porcine kidney tissues. The Canadian Food Inspection Agency (CFIA) had as part of its analytical scope an LC/UV method for quantification of residues of two macrolide antibiotics, tilmicosin and tylosin A, in the kidney, liver, and muscle of cattle, swine, and poultry. The method could not reliably detect concentrations below 10 microg/kg. To increase the scope of the CFIA's analytical capabilities, a sensitive multiresidue quantitative method for macrolide residues in food animal tissues was required. Porcine kidney samples were extracted with acetonitrile and alkaline buffer and cleaned-up using silica-based C18 SPE cartridges. Sample extracts were analyzed using LC/MS/MS with positive electrospray ionization. Fitness for purpose was verified in a single-laboratory validation study using a second analyst. The working analytical range was 5 to 50 microg/kg. LOD and LOQ were 0.5 to 0.6 microg/kg and 1.5 to 3.0 microg/kg, respectively. Limits of identification were 0.5 to 2.0 microg/kg. Relative intermediate precisions were 8 to 17%. Average absolute recoveries were 68 to 76%.