HPLC determination of caffeine and theophylline in Paullinia cupana Kunth (guarana) and Cola spp. samples

A reversed-phase high-performance liquid-chromatographic method for the determination of caffeine and theophylline in commercial guarana samples (drug obtained from the seeds of Paulinia cupana Kunth, Sapindaceae of the Amazon Region) and in Cola spp. samples is described and discussed. The methodology developed is simple and rapid with a minimum of samples preparation required. A comparison of five different techniques for the extraction of caffeine and theophylline is discussed. Furthermore the quantitative determination of caffeine and theophylline in five samples of Brasilian guarana, in two samples of dietetic products containing guarana, in two samples of Cola extract and in three of Cola seed powder are reported.     

Direct detection and identification of Arcobacter species by multiplex PCR in chicken and wastewater samples from Spain

Twenty-two chicken livers, 10 chicken carcasses, and 15 wastewater samples were processed and analyzed for Arcobacter by PCR and traditional culture methods. Samples were enriched for 24 and 48 h, incubated at 30 degrees C under aerobic conditions, and streaked on blood selective media. To determine the best isolation conditions, 20 samples also were processed under microaerophilic conditions at 37 degrees C. Simple and multiplex PCR assays were used directly with enrichment broths and isolated strains. Seventeen Arcobacter strains were isolated from chicken samples, and A. butzleri was the only Arcobacter species identified. The direct PCR assay revealed that 29 of the 32 chicken samples were contaminated with Arcobacter. A. butzleri was the most frequently detected species, although Arcobacter cryaerophilus also was present in some of the samples and Arcobacter skirrowii occasionally was detected. All the wastewater samples were positive by PCR assay for Arcobacter after 24 h of enrichment. A. butzleri and A. cryaerophilus were detected with the multiplex PCR assay. Fourteen Arcobacter strains were isolated from 10 of the 15 water samples analyzed; 7 were identified as A. butzleri and the remaining 7 were A. cryaerophilus. Both for chicken and water samples, Arcobacter detection rate for PCR amplification was higher than for culture isolation. These results indicate the high prevalence of Arcobacter in chicken and wastewater and the inadequacy of available cultural methods for its detection. The species-specific multiplex PCR assay is a rapid method for assessing Arcobacter contamination in chicken and wastewater samples and is a viable alternative to biochemical identification of isolated strains.     

Occurrence of aflatoxins and ochratoxin A in Indian poultry feeds

From 1998 to 2001, 216 ingredients intended for incorporation into chicken feed, which included groundnut cake, maize, millets, rice bran, sorghum, soybean, sunflower, and mixed feeds, were assayed for aflatoxins and ochratoxin A contamination using an indirect competitive enzyme-linked immunosorbent assay. Thirty-eight percent of the samples were contaminated with aflatoxins and 6% with ochratoxin A. The incidence scores of aflatoxin contamination in excess of 10 microg/kg were 41 of 95 for maize, 18 of 30 for mixed feeds, 10 of 37 for groundnut, 6 of 29 for sorghum, 5 of 10 for sunflower, 3 of 14 for rice bran, and 1 of 8 for millet. Ochratoxin A contamination, in excess of 10 microg/kg, was found in 9 of 29 sorghum samples, 1 of 27 groundnut samples, 1 of 14 rice bran samples, 1 of 10 sunflower samples, and 2 of 8 millet samples. Ochratoxin A was not found in maize and mixed feeds. None of the three soybean samples contained ochratoxin A. This is the first report, to our knowledge, of co-occurrence of aflatoxins and ochratoxin A in Indian poultry feeds. The results confirm the importance of analysis of ingredients before incorporating them into mixed feeds.     

High prevalence of Clostridium botulinum types A and B in honey samples detected by polymerase chain reaction

A test protocol for reliable detection of Clostridium botulinum types A and B spores in honey by polymerase chain reaction (PCR) was developed and used for a prevalence survey of C. botulinum spores in 190 honey samples. The inhibiting effects of honey on microbial growth and PCR analysis were overcome by using a method of supernatant filtration (SF) in the preparation of the samples before enrichment and PCR. By using this method, an inoculum of 0.1 spore of C. botulinum/g honey could be detected. In the prevalence survey, spores of C. botulinum were detected in 8 (7%) of the 114 Finnish and in 12 (16%) of the 76 imported honey samples. The quantity of spores in PCR-positive samples varied from less than 18 to 140 spores/kg. Neurotoxin gene sequences corresponding to C. botulinum type A were detected in 17 samples and proteolytic type B in 12 samples by PCR analysis. Both types A and B were detected in nine samples. Strains of C. botulinum type A were isolated from 14 and type B from 2 of the 20 PCR-positive samples. This is the first report of type A spores of C. botulinum being detected and isolated in Fennoscandia.     

Prevalence of Clostridium botulinum in food raw materials used in REPFEDs manufactured in France

Food raw materials used in refrigerated processed foods of extended durability (REPFEDs) manufactured in France were surveyed for Clostridium botulinum types A, B and E using PCR-Enzyme-linked Immunosorbent assay (PCR-ELISA) and mouse bioassay for detection respectively of cells and toxins in enrichment broth. Portions of 25 to 50 g of food were analysed. A total of 8 out of the 102 samples of fish and shellfish, 12 out of the 143 samples of meat and poultry, 1 out of the 62 samples of aroma, sauce and gravy, 4 out of the 25 samples of thickening agents, 3 out of the 26 samples of dehydrated dairy ingredients, and none of the 65 samples of spices, herbs and dehydrated mushroom were positive for C. botulinum in PCR-ELISA, i.e., 6.6% of all the samples tested. The 28 positive samples comprised 10 type A, 10 type B, 4 with both types A and B, and 4 undetermined by PCR typing. No sample positive for type E was detected. Of the 28 samples positive in PCR-ELISA, 15 were also positive in the mouse bioassay. The MPN count was between 1 and 3 C. botulinum/kg of food, which is similar to or in the lower range of values reported in the literature.     

高效液相色谱法测定牛奶中双酚A

建立了测定牛奶中双酚A含量的高效液相色谱方法。样品经乙腈提取,C18固相萃取柱净化,然后采用高效液相色谱测定。双酚A在3.5×10-3～10mg/kg范围内呈良好线性,线性相关系数R2=0.9991。双酚A检测限为2.1×10-3mg/kg,平均添加回收率80.07%～85.53%,相对标准偏差4.42%～9.14%。采用本文所建立的方法对市售牛奶样品进行了测定,未检出双酚A残留。     

Detection of C. Botulinum Types in Honey by mPCR

The aim of this study was to determine the prevalence of Clostridium botulinum in honey samples using conventional methods and multiplex PCR (mPCR). A total number of 150 honey samples were randomly collected from apiaries, retail shops, weekly open bazaars, and supermarkets in Samsun, Turkey. Of 150 honey samples, 4 (2.6%) were positive for the botulinum neurotoxin gene by mPCR analysis. A total of 4 C. botulinum isolates were obtained from the mPCR positive samples, of which 3 were type A and 1 was type B. No samples were positive regarding the type E and type F neurotoxin genes. This is the first report of type A and type B spores of C. botulinum being detected and isolated in Turkey. This study revealed that some honey samples may present a potential hazard for food borne and infant botulism.     

Determination of ochratoxin A in red wine and vinegar by immunoaffinity high-pressure liquid chromatography

A method is described for the determination of ochratoxin A (OTA) in red wine and vinegar using an acidic chloroform extraction, an immmunoaffinity clean-up step, and a high-performance liquid chromatographic determination with fluorescence detection. The detection limit was estimated at 0.002 microg/liter. The mean recovery factors were found at 91.3 and 96.6% for wine and vinegar, respectively. Thirty-one samples of red wine originating from Mediterranean sea countries and 15 samples of vinegar were examined for the presence of OTA. All red wine samples contained OTA. Seventy-two percent of these samples were found to be contaminated over 0.1 microg/liter. Among them, nine samples contained ochratoxin A in the range of 0.5 to 3.4 microg/liter, 12 samples in the range of 0.10 to 0.50 microg/liter (median: 0.176 microg/liter), and 9 samples in the range of 0.010 to 0.100 microg/liter (median: 0.041 microg/liter). All 15 vinegar samples showed the presence of OTA. The most contaminated ones were three balsamic vinegar samples containing 0.156 microg/liter, 0.102 microg/liter, and 0.252 microg/liter of OTA. In the remaining 12 samples, ochratoxin A levels ranged from 0.008 microg/liter to 0.046 microg/liter (median: 0.012 microg/liter). These data are in good agreement with the hypothesis that wine originating from Southern countries might contain significant OTA concentration and showed the possible occurrence of traces of OTA in vinegar.     

Analysis of arabic gum: Study of degradation and water desorption processes

Arabic gum is very much used in the agroalimentary industry, mainly as an emulsifier. It is a natural origin polysaccharide and therefore subject to more or less important variabilities of its properties. This work deals with two similar polysaccharides exuded by Acacia senegal of different history and an Acacia seyal sample. To characterize these samples, wide angle X-ray scattering (WAXS) and thermogravimetric analysis in dynamic and isothermal mode are used.The initial structure checked by X-ray diffraction allows to affirm that samples are equivalent in their crystalline structure. The activation energy used to characterize the main degradation process is the same for A. senegal samples, but higher for the A. seyal gum sample. Differences are furthermore observed for the kinetics of water desorption. We conclude that the samples from the same type of gum (A. senegal) are very similar and that A. seyal exudates can be detected by this technique. The water desorption of the two exudates is different.     

Planing of Norway spruce with very varied ring width

A way of defining a “machining ability” is proposed. Cutting forces: The samples from the slow grown spruce give well known results. In this case the cutting forces are linked with specific gravity by a linear relation. For the samples coming from the fast grown spruces (A), there is no significant relationship between the cutting forces and the specific gravity. These different behaviours are illustrated by the aspect of the chips produced by both batches of spruce. Dimensions scattering. Analysis shows clearly that distributions of dimension quality are similar for spruces A and B. That means there is no compression in the thickness of the samples (z axis). Surface, roughness: More than one roughness criterion is needed to describe surface quality. The 8 criteria used in this work describe the vertical distribution of the roughness measurements. For all the samples, best results are measured on A sample.     

Survey for co-occurrence of ochratoxin A and aflatoxin B1 in dried figs in Turkey by using a single laboratory-validated alkaline extraction method for ochratoxin A

A survey was carried out to determine the co-occurrence of ochratoxin A and aflatoxin B1 in dried figs from Turkey. Samples from two seasons of crops (2003 and 2004) intended for export to the European Union and the 2004 crop obtained from the domestic Turkish market were analyzed. Affinity column cleanup methods were employed for determining separately ochratoxin A and aflatoxin B1, but for ochratoxin A an alkaline extraction procedure was employed (in contrast to the conventionally employed acidic extraction), which gave consistently higher toxin recovery. In-house validation of the ochratoxin A method gave a limit of detection of 0.15 ng/g and a limit of quantification of 0.5 ng/g with a repeatability of 5.8% in the range 5 to 10 ng/g (with a mean recovery of 94% for spiked samples). Positive results for ochratoxin A were confirmed by liquid chromatography-mass spectrometry. For the 2003 export figs (58 samples), 7 samples contained only aflatoxin B1, 2 samples contained only ochratoxin A, and 2 samples contained both toxins (with maximum concentrations of 35.1 ng/g for aflatoxin B1 and 13.0 ng/g for ochratoxin A). Similarly for the 2004 export figs (41 samples), 16 samples contained only aflatoxin B1, 4 samples contained only ochratoxin A, and 2 samples contained both toxins (with maximum concentrations of 20.6 ng/g for aflatoxin B1 and 26.3 ng/g for ochratoxin A). Of 20 retail samples of dried figs from Turkey, only one sample contained ochratoxin A (2.0 ng/g) and none were contaminated with aflatoxin B1. This survey revealed a 14 to 15% incidence of occurrence of ochratoxin A for 2 years, which is higher than previously reported.     

Occurrence of ochratoxin A in Danish wheat and rye, 1992-99

Ochratoxin A concentrations in rye and wheat in Denmark for 1992-99 are reported. The results show that the concentration of ochratoxin A is higher in rye than in wheat for both conventionally and organically grown rye and wheat. The levels in organically grown rye are higher than in conventionally grown based on multiyear mean contents. However, the difference between the two groups of cereals has decreased since the Danish food-monitoring system for ochratoxin A was started in 1986; 2.0% of all samples exceeded the Danish maximum limit of 5 µg kg -1 introduced in 1995. For rye samples, 3.2% exceeded the maximum limit, and for wheat samples, 0.5% exceeded the maximum limit.     

Ochratoxin A in Brazilian roasted and instant coffees

Thirty-four samples of roast and ground coffee, 14 samples of instant coffee and two samples of decaffeinated instant coffee were collected in markets and supermarkets in the city of Campinas, Brazil, and analysed for ochratoxin A using immunoaffinity columns for clean-up and HPLC with fluorescence detection for quantification. The limit of detection was 0.2 ng/g ochratoxin A. Twenty-three samples of ground and roast coffee were found to be contaminated with the toxin at levels ranging between 0.3 and 6.5 ng/g. The average concentration in all 34 samples was 0.9 ng/g. All samples of instant coffee contained ochratoxin A at levels ranging from 0.5 to 5.1 ng/g, with an average figure of 2.2 ng/g. Roast and ground coffee is the type of coffee most used by Brazilians for the preparation of the beverage. Considering that an average Brazilian adult takes five cups of coffee per day, which corresponds to 30 g of roast and ground coffee, the probable daily intake of ochratoxin A by a 70 kg adult would be 0.4 ng/kg bw, which is far below the current Provisional Tolerable Daily Intake of 14 ng/kg bw for ochratoxin A as set by the Codex Alimentarius. To study the transfer of ochratoxin A into coffee brew, the beverage was prepared by two methods: (a) the drip method and (b) the Brazilian country style method. No significant difference was observed between the two methods in terms of extraction of the toxin using five contaminated samples containing between 0.8 and 6.5 ng/g ochratoxin A. The drip method extracted 86 +/- 15% and the Brazilian country style 74 +/- 20% of the ochratoxin A initially present in the roast and ground coffee.     

Some virulence properties and characterization of motile Aeromonas species from milk and white cheese

Three hundred and thirty-eight samples of milk and milk products in Ankara were screened for the presence of motile Aeromonas species. The overall frequency of aeromonads in these samples was 27%. As expected, raw milk samples were more contaminated with aeromonads than were other products. Sixty-five (49.2%) of the 132 bulk raw milk samples were contaminated with aeromonads. In 35 of these samples, populations of aeromonads from 1.5 × 10² to 3.0 × 10³ cfu/mL were counted in ampicillin dextrin agar (ADA). Ten (40%) of the 25 raw milk samples sold in the street were contaminated with aeromonads. In eight of these samples, 1.2 × 10²−3.0 × 10³ cfu/mL were counted. Five (16%) of the 31 pasteurized milk and 12 (8%) of the 150 white cheese samples were contaminated with Aeromonas spp., but no countable aeromonads population was noted in ADA. The incidence of A. hydrophila, A. sobria and A. caviae in all samples was found to be 90.2%, 4.3% and 5.4%, respectively. The majority of the strains identified as A. hydrophila and A. sobria were able to produce haemolysin, protease and DNase, while strains identified as A. caviae were only positive for DNase. All isolated Aeromonas species ( A. hydrophila, A. sobria and A. caviae ) were positive for uptake of Congo red dye. Nevertheless, only strains identified as A. hydrophila and A. sobria showed a high rate of positive results when tested for the production of the Voges–Proskauer reaction and lysine decarboxylase. The results of this work show that motile Aeromonas spp., especially A. hydrophila , are frequently found in these samples. As these products are usually commonly consumed in Ankara, they can pose a risk especially for children, the elderly and immunocompromised individuals.     

Occurrence of enniatins and beauvericin in 60 Chinese medicinal herbs

A total of 60 Chinese medicinal herbs were examined for contamination of the emerging Fusarium mycotoxins enniatins (ENNs) A, A1, B, B1 and beauvericin (BEA). The herbs under study are commonly used in China as both medicines and food. The dried samples of herbs were randomly collected from traditional Chinese medicine stores in Zhejiang province, China. Sample preparation was achieved by methanol extraction, followed by a simple membrane filtration step; no tedious clean-ups were involved. ENNs A, A1, B, B1 and BEA were analysed by the recently developed stable isotope dilution assays, using liquid chromatography-tandem mass spectrometry (LC-MS/MS). With limits of detection ranging between 0.8 and 1.2 µg kg^–1 for the analytes under study, 25% of all analysed samples were contaminated with at least one of the ENNs and BEA. BEA was the most frequently detected toxin with a 20% incidence in all samples. The percentages of ENN-positive samples were lower: each single ENN was detected in 6.7–11.7% of all samples. Considering the total amounts of the five mycotoxins in single samples, values between 2.5 and 751 µg kg^–1 were found. The mean total amount in positive samples was 126 µg kg^–1. Regarding ginger, the frequent occurrence of ENNs and BEA in dried ginger could be confirmed in samples from Germany. However, in fresh ginger root the toxins were not detectable. This is the first report on the presence of ENNs and BEA in Chinese medicinal herbs.     

欧马距离法在香料指纹图谱模式识别中的应用

提出了用欧马距离评价混合体系指纹图谱的方法,并结合主成分分析计算了6种香精标准样品和2种掺兑样品、6种香精标准样品和8种无水乙醇稀释样品GC/MS指纹图谱的欧马距离.结果表明,欧马距离能较好区分6种香精标准样品与2种香精的混合物、6种香精标准样品和8种乙醇稀释样品.欧马距离算法结合主成分分析可用于香精样品指纹图谱识别.     

Prevalence of arcobacter species in drinking water, spring water, and raw milk as determined by multiplex PCR

The aim of this study was to investigate the incidence of Arcobacter species in water sources and raw milk from healthy animals in Kayseri, Turkey. A total of 175 samples of drinking water (n = 100), spring water (n = 25), and raw milk (n = 50) were examined. Arcobacter species were isolated using the membrane filtration technique. Overall, 7 (4%) of the 175 samples yielded Arcobacter spp.: 3 (3%) drinking water samples, 1 (4%) spring water sample, and 3 (6%) raw milk samples. Two species of Arcobacter were recovered from the seven positive samples: Arcobacter butzleri, Arcobacter skirrowii, and A. butzleri plus A. skirrowii found in 3 (1.7%), 2 (1.1%), and 2 (1.1%) samples, respectively. Our study is the first to report the isolation of both A. butzleri and A. skirrowii together from drinking water and is the first report of Arcobacter in milk from healthy cows in Turkey. Based on these findings, the presence of Arcobacter species in environmental waters and raw milk may pose a potential hazard for human health.     

Comparison of the mouse bioassay and enzyme-linked immunosorbent assay procedures for the detection of type A botulinal toxin in food

Samples of chili linked to a foodborne illness outbreak of type A botulism were examined for preformed type A botulinal toxin using two enzyme-linked immunosorbent assay (ELISA) procedures and the mouse bioassay. One of the samples was positive for type A botulinal toxin and three of the samples were negative for type A, B, E, and F botulinal toxins using the three methods. The mouse bioassay indicated that type A toxin was present at the 10,000 minimal lethal dose per gram (MLD per g) of product. The ELISA tests indicated a toxicity of 7,650 MLD per g with one method and 8,350 MLD per g with the other method. The sample toxicity determined by the ELISA was estimated by comparing samples to a standard curve generated with standard type A neurotoxin in casein buffer. The ELISA methods are more rapid than the mouse bioassay, since the toxin type can be determined in 1 day. The mouse bioassay is more sensitive than the ELISA but usually requires multiple assays to obtain the toxin type and toxicity. Type A culture isolates from the sample were also verified using one ELISA method.     

加热方式对大蒜挥发性物质的影响

为探究加热方式对大蒜在色差、气味维度上的变化,实验采用色差仪、电子鼻、气质联用仪分析未加热大蒜(A)、炒制大蒜(B)和蒸制大蒜(C),结合主成分、维恩图等方法分析其差异.色差仪分析表明样品B颜色深,饱和度高,样品A、C颜色浅,饱和度低.电子鼻主成分分析显示样品B、C在风味轮廓上更为相似,与样品A差异大.GC-MS分析显...     

On ochratoxin A and fungal flora in Polish cereals from conventional and ecological farms - Part 1: occurrence of ochratoxin A and fungi in cereals in 1997.

Over 200 samples of Polish cereal grain from the 1997 harvest obtained from conventional and ecological farms were tested for the presence of ochratoxin A as well as for contamination by microscopic fungi. Ochratoxin A contamination of rye from ecological farms was over six times more frequent than that from conventional cultivation. The ochratoxin A content in wheat and barley samples from ecological farms was also higher. No wheat sample from conventional farms contained the mycotoxin. In the group of ecological farms, there were differences in the percentage of cereal samples containing ochratoxin A. The ochratoxin A levels ranged from 0.2 to 57 microg kg(-1). The mean concentration of ochratoxin A in investigated cereal grain was 5.7 microg kg(-1). From samples containing detectable amounts of ochratoxin A, fungi producing ochratoxin A under laboratory conditions were isolated. They were classified as belonging to the species Penicillium cyclopium, P. viridicatum, P. chrysogenum and also Aspergillus alliaceus, A. versicolor, A. glaucus and A. flavus. Penicillium strains - producers of ochratoxin A - were isolated from 93% of the samples; in 7% of samples, only Aspergillus strains producing this mycotoxin were noted. Rye samples mainly from one farm with an ecological type of cultivation and from one conventional farm were contaminated with both Aspergillus and Penicillium mycotoxigenic strains.