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.     

A new method based on electrospray ionisation ion mobility spectrometry (ESI-IMS) for simultaneous determination of caffeine and theophylline

In this work for the first time, simultaneous analysis of caffeine and theophylline was done by ion mobility spectrometry (IMS) only, without a powerful separation technique (e.g., HPLC). Ion mobility spectrometry with low cost, inexpensive maintenance and very fast analysis makes an attractive technique for the simultaneous determination of the caffeine and theophylline in foodstuff samples and biological matrices. In this study, the extraction protocol using molecular imprinted polymer-solid phase extraction (MIP-SPE) was successfully used to directly extract caffeine and theophylline from real samples. The results obtained provided the detection limits of 0.2 and 0.3 μg mL⁻¹ for caffeine and theophylline, respectively. The linear dynamic range of about two orders of magnitude was obtained for these compounds. Also, the proposed method was used to analyse various real samples of green tea and spiked human plasma, and the obtained results confirmed the capability of ESI-IMS for simultaneous detection of caffeine and theophylline.     

高效液相色谱法测定能力验证样品(饮料)中咖啡因含量不确定度评定

目的 对高效液相色谱法测定能力验证样品(饮料)中咖啡因含量的不确定度进行评定。方法 根据JJF1059.1-2012标准要求, 建立不确定度评估的数学模型, 通过对不确定度的各主要分量进行分析计算, 得出合成不确定度以及扩展不确定度, 最终进行测定结果的不确定度评定。结果 当能力验证样品中咖啡因的测定结果为156.9 mg/kg时, 在95%的置信区间下, 其扩展不确定度为5.8 mg/kg (k=2)。结论 采用高效液相色谱法测定饮料中咖啡因含量, 其不确定度的主要来源为标准溶液配制和标准曲线拟合, 该评估模型为检测饮料中咖啡因的不确定度评估提供了参考依据。     

HPLC determination of caffeine in tea,chocolate products and carbonated beverages

Different brands of mate and black tea, chocolate products and carbonated beverages available on the Brazilian market were analysed for caffeine by high-performance liquid chromatography with a UV-vis detector at 254 nm. The column was a reverse phase C18 and the mobile phase consisted of methanol–water (30:70, v/v), acetonitrile–water (10:90, v/v) and methanol–water (25:75, v/v) for tea, chocolate products and soft drinks respectively. Caffeine content ranged from 1.05 to 15.83 mg per cup in mate tea, from 32.21 to 36.23 mg per cup in black tea, from 0.14 to 0.95 mg g⁻¹ in chocolate products from 2.73 to 7.49 mg per can in guaraná-type soft drinks and from 19.81 to 45.89 mg per can in cola soft drinks. These data indicate that the levels of caffeine in Brazilian teas, chocolates and soft drinks are within the ranges reported for similar products in other countries. © 1999 Society of Chemical Industry     

Identification of barramundi (Lates calcarifer) and tilapia (Oreochromis spp.) fillets by DNA- and protein-analytical methods

Fish and fillet of barramundi (Lates calcarifer) and tilapia (Oreochromis species) obtained from wholesale and retail trade were assigned to species by sequencing of PCR products. Two segments (358 and 464 bp) of the cytochrome b gene (cytb) were amplified using universal primers. The amplicons gave characteristic patterns in SSCP-analysis (single strand conformation polymorphism) suitable for differentiation of Lates calcarifer from Lates niloticus and Lateolabrax japonicus. Intra-specific variation of sequences and SSCP patterns were observed for barramundi. In case of tilapia species, it was found to be difficult to identify samples by BLAST due to the high similarity of cytb sequences of O. niloticus, O. mossambicus, O. aureus and Sarotherodon galileus. Four different patterns of single strand DNA (ssDNA) were obtained by SSCP analysis of the 464 bp amplicon of tilapia. Different patterns of ssDNA matched to variations in sequences. Protein profiles obtained by IEF (isoelectric focusing) of water-soluble proteins from raw fillet were found to be suitable for rapid differentiation of Lates calcarifer from Lateolabrax japonicus, but the three different Oreochromis species expressed only minor differences in protein patterns. The patterns of the tilapia and barramundi species showed a number of acidic, heat-stable proteins, presumably representing parvalbumin.     

Characterization of new sources of mungbean (Vigna radiata (L.) Wilczek) resistance to bruchids, Callosobruchus spp. (Coleoptera: Bruchidae)

The bruchid beetles Callosobruchus chinensis (L.) and C. maculatus (F.) are destructive pests of stored mungbean (Vigna radiata (L.) Wilczek). The development of resistant mungbean varieties to manage bruchids is a major breeding objective. In the present study, we investigated the characteristics of resistance to C. chinensis and C. maculatus in two new resistant mungbean accessions V1128 and V2817, and two previously reported resistant accessions V2709 and V2802, compared to the susceptible mungbean cultivar, KPS1. Evaluation for resistance to both bruchid species using whole and decorticated seeds revealed that V1128 and V2817 were free from damage, V2709 and V2802 showed partial damage with low or moderate number of bruchids emerging from seeds, and KPS1 showed complete damage with the highest number of bruchids emerged. Comparison of the seeds harvested at seed filling, early maturity and full maturity stages revealed that the percentage of damaged seeds from resistant accessions was lower at all stages compared with KPS1. V1128 and V2817 showed complete resistance against both bruchids regardless of when their seeds were harvested, while resistance in V2709 and V2802 were most pronounced at full maturity, and KPS1 was totally damaged at all times tested. These results suggest that the chemical factor(s) conferring resistance is synthesized as early as the seed filling stage. Evaluation of resistance using artificial seeds showed that increasing the percentage of resistant seed powder adversely affected bruchid growth and development. The number of adults emerging from seeds and number of damaged seeds decreased while adult developmental period increased as the proportion of resistant seed powder increased. The weight of emerging male and female adults of C. maculatus was lighter than those from the seeds containing susceptible seed powder alone. However, C. chinensis adults were not affected by the same test. The results suggest that biochemical(s) in cotyledon tissue are responsible for the resistance and the seed coat had no protective role against the bruchids. Although all four resistant accessions evaluated are useful for mungbean breeding, V1128 and V2817 show complete resistance to both C. chinensis and C. maculatus. Thus, these two new resistant sources may be the most effective for breeding purposes.     

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     

Voltammetric determination of caffeine in beverage samples on bare boron-doped diamond electrode

A sensitive and selective electrochemical method for the caffeine determination using bare boron-doped diamond electrode was developed. It was found that caffeine provided highly reproducible and well-defined irreversible oxidation peak at very positive potential. The effects of supporting electrolyte, pH and scan rate on the voltammetric response of caffeine oxidation were studied to select the optimum experimental conditions. Linear response of peak current on the concentration in the range from 4×10(-7) to 2.5×10(-5)M, good repeatability (RSD of 2.1%) and detection limit of 1.5×10(-7)M without any chemical modifications and electrochemical surface pretreatment were evaluated. The effect of possible interferents appeared to be negligible which evidently proved very good selectivity. The proposed method was successfully applied for the caffeine determination in commercially available beverage samples, with results in a close statistical agreement to those declared by manufacturer and HPLC used as independent method.     

Yersinia spp. in foods and related environments

Food samples, porcine swabs and environmental swabs were examined for the presence of Yersinia spp. over a 3-year period. A total of 118 strains were isolated from 3717 samples. Most isolates belonged to Y. enterocolitica biotype 1 or Y. intermedia. The most commonly detected source of Yersinia organisms in foods was pasteurized milk. Investigations at one local dairy indicated a need for improved hygiene at the processing plant. Enrichment at 4°C in 1% buffered peptone water followed by subculture to Cefsulodin-Irgasan-Novobiocin (CIN) agar appears suitable for the isolation of Yersinia spp. from a variety of food commodities and environments.     

Oviposition site selection by Tineola bisselliella, Tinea spp (Lepidoptera: Tineidae) and Anthrenus flavipes (Coleoptera: Dermestidae)

Odours from dried yeast localized oviposition on fabric by Tineola bisselliella (Hummel), Tinea translucens Meyrick and Tinea pellionella L. in choice experiments in the laboratory. In Tineola bisselliella this effect was augmented by odours from the faeces from conspecific larvae that had eaten a favourable diet containing dried yeast. In contrast with these Lepidoptera, oviposition by Anthrenus flavipes LeConte was unaffected by yeast odours and deterred by odours from conspecific cultures. Blood-stains did not influence ovipositional choice. Some plant phenolics and flavonoids and quinones incompletely deterred oviposition. Both vegetable oils and fatty acids from wool wax deterred oviposition equally well when present in large amounts. Treatment of wool to control micro-organisms might reduce insect damage indirectly.     

Derivatization procedure and HPLC determination of 2-ethoxyethanol in cosmetic samples

Ethylene glycol monoethyl ether or 2-ethoxyethanol finds a wide industrial application as a solvent for lacquers, inks, dyes, household products and as a surfactant. It is also found in cosmetics such as nail products, face cleansers, liquid soaps, oral care products, hair colours and fixatives. The potential hazard to human health of 2-ethoxyethanol following inhalation and dermal exposure has been recently reviewed and the European Cosmetic, Toiletry and Perfumery Association (COLIPA) has issued recommendations suggesting its non-use as a cosmetic ingredient. Therefore a simple and fast monitoring method is necessary for routine control to identify and quantify 2-ethoxyethanol in raw materials and finished cosmetics. We have developed a sensitive and selective method to determine 2-ethoxyethanol in complex matrices by precolumn derivatization with 1-naphthyl isocyanate and RP-HPLC analysis. Four laboratory-made cosmetic formulations (a nail lacquer remover, a baby oil, a skin lotion and an emollient O/W emulsion) containing three known amounts of 2-ethoxyethanol (0.1%, 2.0%, 5.0%) have been studied. The obtained results show that this chromatographic procedure provides a good estimate of the true concentration of 2-ethoxyethanol in complex matrices and it is reliable for routine analyses in quality control.     

克罗诺杆菌环境耐受性的评价

婴儿配方奶粉(powdered infant formulas,PIF)是克罗诺杆菌(Cronobacter spp.)的主要污染来源,其原辅料的添加和加工环境是最为主要的传播途径。为彻底有效地对克罗诺杆菌进行预防和控制,保障乳制品的安全,本实验以8株分离自PIF及其加工环境中的克罗诺杆菌为研究对象,对其耐热性、耐酸碱性、耐干燥性及耐高渗性进行研究,分析不同菌株环境耐受性之间的差异。结果表明：8株克罗诺杆菌对热的耐受性存在差异性,其中分离自终产品的菌株耐热性相对较强;克罗诺杆菌普遍具有耐酸不耐碱的特性,在pH为1.5的酸性条件下,所有菌株在处理30 s后仍可存活;克罗诺杆菌具有较强的抗干燥能力,在室温条件下,空气相对湿度为20.7%的条件下,所有菌株在7个月的试验过程中菌数均从初始浓度10⁸ CFU/mL下降到10⁵ CFU/mL,这种缓慢的下降趋势说明克罗诺杆菌具有很强的抵抗干燥环境的能力;克罗诺杆菌具有较强的耐高渗性,当NaCl的浓度大于8%时,所有菌株生长缓慢,且在等渗条件下,克罗诺杆菌对山梨醇的耐受性强于NaCl。     

A quantitative micromethod for the caffeine determination (author's transl)

A combined procedure with thin-layer-chromatography and densitometry is described for the quantitative estimation of caffeine in biological material. This method is applicable in the nanogram range. Test samples of less than 100 microliter may be used. The samples (capillary-blood) are extracted with the same volume of chloroform. Caffeine is separated from interfering compounds by thinlayer-chromatography. Commercial silica-60-plates with chloroform/acetone (9+1; v/v) as solvent are used. The running time is about 30 min. The quantitative densitometric determinations are performed in the remission mode at 273 nm. In the range from 10 to 60 ng/spot the calibration curve is linear. Accurate quantitative data will be obtained even at concentrations of 1 mg/l caffeine. The detection limit is at about 0.1 mg/l.     

HPLC/PDA determination of carminic acid and 4-aminocarminic acid using relative molar sensitivities with respect to caffeine

To accurately determine carminic acid (CA) and its derivative 4-aminocarminic acid (4-ACA), a novel, high-performance liquid chromatography with photodiode array detector (HPLC/PDA) method using relative molar sensitivity (RMS) was developed. The method requires no analytical standards of CA and 4-ACA; instead it uses the RMS values with respect to caffeine (CAF), which is used as an internal standard. An off-line combination of ¹H-quantitative nuclear magnetic resonance spectroscopy (¹H-qNMR) and HPLC/PDA was able to precisely determine the RMSs of CA^274nm/CAF^274nm and 4-ACA^274nm/CAF^274nm. To confirm the performance of the HPLC/PDA method using RMSs, the CA and 4-ACA contents in test samples were tested using four different HPLC-PDA instruments and one HPLC-UV. The relative standard deviations of the results obtained from five chromatographs and two columns were less than 2.7% for CA^274nm/CAF^274nm and 1.1% for 4-ACA^274nm/CAF^274nm. The ¹H-qNMR method was directly employed to analyse the CA and 4-ACA contents in test samples. The differences between the quantitative values obtained from both methods were less than 5% for CA and 3% for 4-ACA. These results demonstrate that the HPLC/PDA method using RMSs to CAF is a simple and reliable quantification method that does not require CA and 4-ACA certified reference materials.     

Dermestes leechi Kalik in stored products and new diagnostic characters for Dermestes spp (Coleoptera: Dermestidae)

The Indian species Dermestes leechi Kalik has been found in bone grist imported into Scotland from Pakistan. This is the first record from stored products. An illustrated key is provided to distinguish this species from Dermestes peruvianus Laporte de Castelnau, Dermestes haemorrhoidalis Küster and Dermestes ater Degeer. New taxonomic characters, including structures associated with the female genitalia, are described. D. leechi is recorded from Africa for the first time.     

Use of high pressure liquid chromatography (HPLC) for the determination of cda-tocopherol levels in forage (silage/grass) samples collected from different regions in Ireland

First, second and third cut grass silage samples were collected from eight regions around Ireland (184 samples in total) and analysed for vitamin E content. Fresh grass samples were also collected at one site in Co. Cork and analysed for α-tocopherol content. The concentration of α-tocopherol [μg/g dry matter (DM)] was determined using high performance liquid chromatography analysis. A wide variation in α-tocopherol levels in silage samples was found. α-Tocopherol levels in first, second and third cut silage samples ranged from 4.9 to 20.8, 4.4 to 13.0 and 1.3 to 3.9 μg/g DM, respectively. The mean values of α-tocopherol in first, second and third cut silage samples were 11.3±0.9, 9.7±0.7 and 2.3±0.5 μg/g DM, respectively. There was no significant difference in α-tocopherol levels between first and second cut silage samples. However, third cut silage samples had significantly (P<0.05) less α-tocopherol compared to first and second cut samples. There was no decrease in α-tocopherol levels in silage samples following storage at −20°C for 2 months. Fresh grass samples were classified as pasture grass, meadow grass (consisting of 20% Bromus spp and 80% Holcus spp, hedgerow grasses (predominately Dactylis glomerata L.) and white or red clover. Pasture grass had significantly (P<0.05) higher α-tocopherol levels than other grass types analysed. Meadow grass had significantly (P<0.05) higher α-tocopherol levels than clover or hedgerow grasses. α-Tocopherol concentration in the different grass types decreased in the order: pasture grass>meadow grass>hedgerow>white clover>red clover.     

Attractancy of diethyl ether extracts of Arachis hypogaea (Linnaeus) (Papilionaceae), Citrullus lanatus (Thunberg) (Cucurbitaceae) and Irvingia gabonensis var. excelsa (Baillon) (Irvingiaceae) to Oryzaephilus mercator (Fauvel) (Coleoptera: Silvanidae)

Volatiles of the diethyl ether extracts of I. gabonensis, C. lanatus and A. hypogaea attracted 1–7 and 28–42-day-old adult O. mercator when tested in a two-choice pitfall Petri dish bioassay. For both age groups, a significant difference was observed in response between fed and starved adults to the different extracts. Differences in starvation time did not affect responses of individuals in the 1–7-day-old group; a significant effect was observed in the 28–42-day-old group.

Adults, 28–42-day-old were significantly more responsive to virtually all extracts than 1–7-day-old adults. Responses of adults in each age group to the different extracts were not significantly different. The time of trial within the period 0900–1900 h did not significantly affect the response of adults in both age groups, starved for 2 days, to any extract.