Analysis of patulin in apple products by liquid–liquid extraction,solid phase extraction and matrix solid-phase dispersion methods: a comparative study

Patulin is a marker of quality in the apple and apple juice industry and due to the potential risk for human health, reliable and potential methods for extracting patulin from a sample are therefore needed. In this study, the three methods with liquid–liquid extraction, matrix solid-phase dispersion (MSPD) and with solid-phase extraction (SPE) were studied for extracting patulin from different apple products. Result showed that for AJC and apple sample, MSPD method is most suitable for extracting patulin among the three methods. The recovery rates of AJC and apple sample were 80.35–114.46 and 79.68–94.32%, respectively, the coefficient variations were 3.18–4.90%; For dilute juice, SPE procedure is suitable for analysis of patulin and the recovery rates were and 85.35–90.14%.     

HPLC determination of stevioside in plant material and food samples

 An HPLC method for the determination of sweet-tasting stevioside (STS) in the leaves of the plant Stevia rebaudiana and in some beverages (e.g. tea, orange juice) was developed. The pre-separation procedure consisted of extraction of STS from the plant material using boiling water and a solid-phase extraction (SPE). Recovery rates of the SPE for the analysed matrices ranged from 92.8% to 97.8% (for concentrations of STS of 105, 210 and 300 μg/ml; Relative Standard Deviation (RSD)≤3.3%). The chromatographic separations were realized using a C₁₈ column, the mobile phase consisting of methanol and water, and with UV detection at 210 nm. The limits of determination of STS were 5 μg/ml for the leaf extracts and the tea sample and 8 μg/ml for the juice sample. Received: 7 April 1998     

HPLC determination of stevioside in plant material and food samples

 An HPLC method for the determination of sweet-tasting stevioside (STS) in the leaves of the plant Stevia rebaudiana and in some beverages (e.g. tea, orange juice) was developed. The pre-separation procedure consisted of extraction of STS from the plant material using boiling water and a solid-phase extraction (SPE). Recovery rates of the SPE for the analysed matrices ranged from 92.8% to 97.8% (for concentrations of STS of 105, 210 and 300 μg/ml; Relative Standard Deviation (RSD)≤3.3%). The chromatographic separations were realized using a C₁₈ column, the mobile phase consisting of methanol and water, and with UV detection at 210 nm. The limits of determination of STS were 5 μg/ml for the leaf extracts and the tea sample and 8 μg/ml for the juice sample. Received: 7 April 1998     

Determination of ochratoxin A in wheat after clean-up through a DNA aptamer-based solid phase extraction column

A DNA aptamer with high affinity and specificity to ochratoxin A (OTA) was conjugated to a coupling gel and used as sorbent for the preparation of solid phase extraction (SPE) columns. The SPE columns packed with 300 μl oligosorbent (24 nmol DNA) showed a linear (r = 0.999) behaviour in the range of 0.4–500 ng OTA. After optimisation of the extraction step, SPE columns were used for clean-up of OTA from wheat prior to liquid chromatographic (HPLC) analysis with fluorescence detection (FLD). Average recoveries from wheat samples spiked at levels of 0.5–50 ng/g ranged from 74% to 88% (relative standard deviation <6%) with limits of detection and of quantification of 23 and 77 pg/g, respectively. The comparative HPLC/FLD analyses of 33 naturally contaminated durum wheat samples cleaned-up on both aptamer-SPE and immunoaffinity (IMA) columns showed a good correlation (r = 0.990). Aptamer-SPE columns could be re-used up to five times without any loss of performance.     

Development of an HPLC–UV method for the simultaneous determination of tetracyclines in muscle and liver of porcine,chicken and bovine with accelerated solvent extraction

A simple and especially rapid method-using accelerated solvent extraction (ASE) and HPLC has been developed for the quantitative determination of oxytetracycline, tetracycline, chlortetracycline, minocycline, methacycline, demeclocycline and doxycycline in muscle and liver of porcine, chicken and bovine. Samples of muscle and liver were extracted with trichloracetic acid/acetonitrile using ASE instrument, parameters such as extraction temperature (40–80 °C) and pressure (45–85 bar) were investigated and the selected extraction (60 °C, 65 bar) was most effective. The limits of detection were lower than 10 μg/kg and limits of quantification no more than 15 μg/kg for all compounds in muscle and liver. The recoveries of tetracyclines spiked at levels of muscle 50–150 μg/kg, liver 150–450 μg/kg, averaged from 75.0% to 104.9% with the relative standard deviation values less than 10%. The method was applied to determine 30 real porcine livers. It is demonstrated that the new method is robust for detection and quantification of seven tetracycline residues in muscle and liver of porcine, chicken and bovine.     

基于分子印迹整体柱的在线固相萃取-液相色谱联用测定奶粉中四环素类兽药残留

目的建立高交联结构的分子印迹整体柱(molecularly imprinted polymer monolithic column,MIP-MC)制备方法,利用在线固相萃取与液相色谱联用技术检测奶粉样品中四环素类兽药残留。方法在不锈钢色谱柱中,以土霉素为模板,甲基丙烯酸和甲基丙烯酸羟乙酯为功能单体,乙二醇二甲基丙烯酸酯和双季戊四醇六丙烯酸酯为交联剂,偶氮二异丁腈为引发剂,在丙酮-甲醇-十二醇混合溶剂中,制备了分子印迹整体柱。将整体柱与液相色谱联用,在线固相萃取奶粉中的四环素类兽药残留。结果在最佳在线固相萃取条件下,获得了较高的富集因子(19.3)和净化效果。四环素类在0.05、0.25和0.5 mg/kg 3个加标水平下,回收率为84.2%~103.4%,相对标准偏差为1.37%~4.87%,方法检出限(S/N=3)和定量限(S/N=10)分别为8.48~11.74?g/kg和28.24~39.09?g/kg。结论该在线固相萃取方法简单快速、灵敏性高、选择性好,适用于奶粉中四环素类抗生素残留的测定。     

HPLC determination of trimethoprim in meat and milk with an SPE preseparation procedure

 An HPLC method in combination with solid-phase extraction and dual wavelength detection has been developed in order to monitor trimethoprim residues in meat and milk. The extraction recoveries for the maximal residual limit concentration in food (50 ng/g) were 86±5% for meat and 73±6 % for milk samples. The limits of determination of trimethoprim were 5 ng/g at 229 nm and about 15 ng/g at 280 nm. The method has been developed in response to specific needs of food control laboratories. This publication introduces the assay for milk and meat samples. Received: 27 March 1996/Revised version: 23 July 1996     

Development of a liquid chromatography-tandem mass spectrometric method for the simultaneous determination of tropane alkaloids and glycoalkaloids in crops

A new, rapid and sensitive multiresidue method is reported for the simultaneous determination of tropane alkaloids (tropine, atropine, scopolamine, homatropine, anisodamine) and glycoalkaloids (α-solanine, α-chaconine) in grains and seeds (wheat, rye, maize, soybean, linseed). Dispersive solid phase extraction (DSPE) was performed with 0.5% formic acid in acetonitrile/water and a mixture of magnesium sulphate, sodium chloride and sodium citrate. For a fast and effective clean-up procedure for oily matrices such as soybean and linseed, matrix solid phase dispersion (MSPD) C₁₈ material was used to remove co-extracted non-polar components. No clean-up was necessary for less oily matrices following extraction. The analytes were separated by isocratic HPLC on a Chirobiotic V column and detected using a triple quadrupole mass spectrometer with electrospray ionization (ESI). All analytes were monitored in the positive ion mode. The method performance is presented in terms of linearity in the range 5–80?ng/g (r ^2?=?0.998), specifity, selectivity, accuracy (recoveries from 61–111%), precision (CV?<?5%) and ruggedness. The limits of quantitation (LOQ) were in the range 2.2–4.9?ng/g.     

HPLC-integrated solid-phase extraction with photochemical post-column derivatization for the determination of oxacillin, cloxacillin and dicloxacillin in raw milk

 An automated reversed-phase high-performance liquid chromatography (HPLC) method with UV detection at 300 nm after on-line coupled solid-phase extraction (SPE) and photochemical post-column derivatization was developed for the residue analysis of oxacillin, cloxacillin and dicloxacillin in raw milk. After a centrifugation step for defatting and ultrafiltration, a HPLC-integrated SPE using a restricted access sorbent, is performed. By operating the system in parallel mode, SPE and HPLC run are performed simultaneously. The procedure allows the analysis of about 30 samples within 24 h and was used to investigate the milk of cows which had been treated with oxacillin. The detection limit was 3 μg/kg for oxacillin and cloxacillin and 5 μg/kg for dicloxacillin. The recoveries were between 66% and 81% with coefficients of variation between 3.3% and 7.7%. Using an acetonitrile extraction procedure better recoveries of ≥90% were obtained with fortified samples, but more or less identical results were obtained for the samples with incurred residues after correction for recovery. Received: 16 February 1998     

固相萃取净化-高效液相色谱法测定肉制品中的9 种N-亚硝基化合物

建立采用固相萃取-高效液相色谱法同时测定肉制品中9 种N-亚硝基化合物的方法。以甲醇为提取剂,60 ℃条件下超声提取,经CNWBOND HC-C18固相萃取柱净化后,采用AQ-C18色谱柱分析。甲醇和超纯水为流动相实现了待测物的良好分离,紫外检测器检测,结合保留时间和光谱图定性,外标法定量。结果表明：9 种N-亚硝基化合物线性关系良好,相关系数均大于0.99;方法定量限（RSN = 10）为0.047～0.097 μg/g;回收率为72.6%～97.6%;相对标准偏差（n＝6）为3.1%～7.9%。此方法前处理简单、回收率高、精密度好,适用于肉制品中9 种N-亚硝基化合物的同时测定。     

Development of a liquid chromatography-tandem mass spectrometric method for the simultaneous determination of tropane alkaloids and glycoalkaloids in crops

A new, rapid and sensitive multiresidue method is reported for the simultaneous determination of tropane alkaloids (tropine, atropine, scopolamine, homatropine, anisodamine) and glycoalkaloids (α-solanine, α-chaconine) in grains and seeds (wheat, rye, maize, soybean, linseed). Dispersive solid phase extraction (DSPE) was performed with 0.5% formic acid in acetonitrile/water and a mixture of magnesium sulphate, sodium chloride and sodium citrate. For a fast and effective clean-up procedure for oily matrices such as soybean and linseed, matrix solid phase dispersion (MSPD) C(18) material was used to remove co-extracted non-polar components. No clean-up was necessary for less oily matrices following extraction. The analytes were separated by isocratic HPLC on a Chirobiotic V column and detected using a triple quadrupole mass spectrometer with electrospray ionization (ESI). All analytes were monitored in the positive ion mode. The method performance is presented in terms of linearity in the range 5-80 ng/g (r(2)=0.998), specifity, selectivity, accuracy (recoveries from 61-111%), precision (CV<5%) and ruggedness. The limits of quantitation (LOQ) were in the range 2.2-4.9 ng/g.     

Determination of ochratoxin A in organic and non-organic cereals and cereal products from Spain and Portugal

The objective of this work was to know the occurrence of OTA in organic and non-organic cereals and cereal products from Spain and Portugal. A method based on extraction with matrix solid phase dispersion (MSPD) using octylsilica (C₈) followed by liquid chromatography coupled with fluorescence detection (LC–FD) was used to determine OTA from the selected samples. Recoveries of OTA from the studied samples spiked at 10 ng/g level ranged from 78% to 89% with a standard deviation of 3.66. The limits of detection and quantification of this method were 0.05 and 0.19 ng/g, respectively. Furthermore, LC–FD after OTA methylation was used to confirm the identity of OTA in all positive samples. This procedure was applied to 83 organic and non-organic samples including rice, wheat, barley, rye, oats and maize from Spain and Portugal. OTA was detected in 22% of the samples, with concentrations ranging from 0.20 to 27.10 ng/g. From the total OTA contaminated samples (n = 18), 72% were organic cereal and 28% were non-organic cereal samples, with mean concentrations of 1.64 and 0.05 ng/g, respectively. The 66% and 34% of contaminated samples were from Spain and Portugal, respectively, with mean concentrations of 0.93 and 0.64 ng/g for each country. Six contaminated samples exceeded the maximum limits (ML) for OTA fixed by European Commission Regulation (5 μg/kg), among them three were from Spain and three from Portugal.     

Application of principal component analysis to chemical characteristics of Mahon cheese

 This paper describes a method suitable for the analysis of diglycerides (DG) in cheese. The method, also suitable for other foods, is essentially based upon the solid-phase extraction (SPE) of DG followed by capillary gas chromatographic analysis with a polar stationary phase (TAP). SPE is applied to avoid the column overloading caused by triglycerides; however, SPE is able to catalyse isomerization from the 1,2-form to the 1,3-form. This introduces an error into the results, as the two forms have different origins and are produced at different stages of ripening. A comparison between the use of silica and diol phases was made and it was seen that no isomerization occurred when the diol phase was used. Repeatability and recovery trials were carried out and the coefficient of variation (relative standard error) of the former trial was approximately 10%. Finally, it is demonstrated that it is possible to re-use the SPE cartridge at least four times. Received: 15 July 1996/Revised version: 19 September 1996     

虚拟模板分子印迹固相萃取-高效液相色谱法检测食品中联苯三唑醇与烯唑醇

利用虚拟模板分子印迹固相萃取技术对食品中的联苯三唑醇与烯唑醇进行分离富集,并建立高效液相色谱检测方法。以腈菌唑为模板分子,丙烯酰胺（AM）为功能单体,EDMA为交联剂,乙腈为致孔剂,采用本体聚合法制备得到了腈菌唑分子印迹聚合物（MMIP）,以此聚合物为固相萃取剂,制备固相萃取柱,对食品中联苯三唑醇和烯唑醇进行分离富集,并采用高效液相色谱法测定其在食品中的残留。采用C18色谱柱分离,以甲醇-水（体积比为90：10）溶液为流动相,紫外检测波长为210 nm,外标法定量。结果表明,联苯三唑醇和烯唑醇的平均回收率分别为89.40~90.88%和81.37~83.26%,RSD分别为1.08~2.04%和1.29~2.52%,检测限均为0.20 μg/g。此聚合物对两种杀菌剂具有良好的吸附能力,拟模板分子印迹固相萃取技术可以用于这两种杀菌剂的分离检测。     

RP-HPLC快速检测牛奶中7中四环素类药物残留量的研究

目的：建立快速检测牛奶中7 种四环素类药物残留的反相高效液相色谱法。方法：用0.1mol/L McIlvaine-EDTA 溶液和50% 三氯乙酸(TCA)溶液共同处理牛奶样品,通过Waters Oasis HLB 固相萃取柱进行净化处理。仪器条件中所用色谱柱为SunfireTM C18(250mm × 4.6mm,5μm)柱,以甲醇- 乙腈-0.01mol/L 草酸(pH2)为流动相,采用梯度洗脱模式。结果：7 种四环素类药物线性范围宽,峰面积和样品浓度在0.05～1μg/ml 范围内呈很好的线性关系,加标平均回收率为78.66%～106%,最低检测限为0.02μg/ml。结论：该方法操作简单、重现性好、检测限低、灵敏可靠,可在14min 内同时将四环素等7 种四环素类药物达到基线分离。     

Evolution of free fatty acid profile during ripening in cheeses manufactured from bovine, ovine and caprine milks with extracts of Cynara cardunculus as coagulant

 Changes in the concentrations of free fatty acids (FFA) in bovine, ovine and caprine milk cheeses manufactured with a plant rennet (flowers of Cynara cardunculus) were studied throughout a 68-day ripening period. The long-chain saturated (C16 : 0 and C18 : 0) and unsaturated (C18 : 1, C18 : 2, and C18 : 3) FFA were the most abundant at all stages of ripening. The overall concentration of FFA in fresh cheese was 3598, 3538 and 3868 mg/kg cheese for bovine, ovine and caprine milk cheeses, respectively; these values increased to 5047, 6517 and 5257 mg/kg cheese, respectively, by 68 days, of which 1171, 1734 and 1791 mg/kg cheese, respectively, were accounted for by C4 : 0 – C12 : 0. The FFA that showed the highest fractional increase by 68 days of ripening in bovine milk cheese were C4 : 0, C6 : 0, C8 : 0, C12 : 0, C18 : 1 and C18 : 2; in ovine milk cheese they were C4 : 0, C6 : 0, C8 : 0, C10 : 0, C14 : 0, and C18 : 1; and in caprine milk cheese they were C4 : 0, C8 : 0, C10 : 0, C14 : 0 and C18 : 1. Received: 2 August 1996     

A new quick solid-phase extraction method for the quantification of heterocyclic aromatic amines

A new solid-phase extraction (SPE) method using only one SPE cartridge is described as a clean-up procedure for the determination of heterocyclic aromatic amines (HAAs). In particular, the polar HAAs imidazoquinolines and imidazoquinoxalines, which are well-known toxic compounds in thermally treated food, can be determined by this quick and simple method. For validation of the method meat extracts were analyzed. For determination of the percentage recovery and standard deviation the beef extract was spiked (40 ng/g of each HAA) and analyzed 10 times. The polar HAAs were determined in this meat extract with a recoveries of 62 % to 95% and standard deviations of 3% to 5%. The recovery rates of the less polar HAAs Harman and Norharman were much lower (25±5% to 32±5%). The limit of detection for polar and less polar HAAs was between 3 ng/g and 9 ng/g in the meat extract matrix. The method comprises extraction with methanolic NaOH, centrifugation and SPE using a commercially available polystyrene copolymer cartridge. After different washing steps the eluate was analyzed by high-performance liquid chromatography with diode-array detection. The advantages of this new method are the reduced amounts of time and organic solvents required. Received: 29 November 1999 / Revised version: 21 February 2000     

The evolution of free fatty acids during the ripening of Idiazábal cheese: influence of rennet type

 The object of the present study was to determine the influence of the rennet type on the free fatty acid (FFA) content of Idiazábal cheese. Varying this parameter during cheese-making resulted in significant differences in FFA content in the cheeses during ripening. The main FFAs in both cheese batches were caproic acid (C6 : 0) and capric acid (C10 : 0). The differences between the extreme values for the lipolysis rate were around 45%, which emphasizes the importance of the cheese-making procedure employed on lipolysis in this type of cheese. The use of locally made rennet in the preparation of Idiazábal cheese increased the level of lipolysis in the cheese. Received: 4 December 1998 / Revised version: 18 March 1999     

Application of High-Performance Liquid Chromatography with Diode Array Detector for Simultaneous Determination of 11 Synthetic Dyes in Selected Beverages and Foodstuffs

A simple, inexpensive and robust high-performance liquid chromatography diode array detector (HPC-DAD) procedures are proposed to analyse food dyes in beverages, hard candy and fish roe samples. An ether-linked phenyl stationary phase provides sufficient selectivity and chromatographic performance for separation of 11 sulfonated azo dyes. Beverage samples were only diluted (and degassed when needed) before analysis. Solid-phase extraction (SPE) or matrix solid-phase dispersion (MSPD) procedures are proposed for efficient extraction of the analytes from candies or fish roe samples, respectively. Limits of detection (LODs) were from 0.005 to 0.013 μg mL^?1 and limits of quantification (LOQs) between 0.014 and 0.038 μg mL^?1. HPLC-DAD method was validated in terms of intra- and inter-day accuracy and precision at three concentration levels 2, 1, and 0.1 μg mL^?1. Validation was also performed for SPE and MSPD extraction procedures including intra- and inter-day accuracy (Recovery %) and precision (RSD%), as well as intra-laboratory reproducibility. Application to analysis of beverages and food samples available to consumers proved that described methods are suitable for the routine analysis of dyes in food products.     

亲水亲脂平衡柱萃取高效液相色谱法快速测定干酪中的黄曲霉毒素M1

研究了利用RP-HPLC对干酪中黄曲霉毒素M1进行测定,分别采用乙腈、甲醇、丙酮作为提取溶剂对干酪样品中的黄曲霉毒素M1进行提取,利用一种亲水亲脂平衡柱萃取黄曲霉毒素M1,然后上机检测,采用质量分数为25％的乙腈水溶液为流动相,流速1 mL/min,利用荧光检测器进行检测.结果表明,采用乙腈作为干酪中黄曲霉毒素M1的提取试剂比其他2种溶剂更为适宜;OASIS HLB固相萃取净化柱对于黄曲霉毒素M1的分离纯化效果很好;干酪中的黄曲霉毒素在8mm内被快速检出,平均同收率较高,相对标准偏差(RSD)为1.4％～5.7％,最低检出限为0.037μg/kg.该方法是一种针对干酪中黄曲霉毒素M1的快速高效的检测方法.