Hypersensitivity reactions due to black henna tattoos and their components: are the clinical pictures related to the immune pathomechanism?

Hypersensitivity to para-phenylenediamine (PPD) and related compounds induced by temporary black henna tattoos has become a serious health problem worldwide. Different patterns of sensitization with various clinical aspects are described in literature due to PPD associated to henna tattoo and these manifestations are likely correlated with the immunological and dermatological pathomechanisms involved. Henna is the Persian name of the plant Lawsonia inermis, Fam. Lythraceae. It is a woody shrub that grow in regions of North Africa, South Asia, India and Sri Lanka. Nowadays it is rather frequent to see temporary “tattoos” performed with henna. To make tattoos darker and long-lasting PPD has been associated to henna in tattoo drawings mixtures, so obtaining “black henna”. In these years there has been a rise of contact sensitization to PPD and in medical literature an increased number of cases have been reported on temporary henna tattoo application. Here we review the various clinical patterns related to PPD and henna tattoo, to investigate the possible link between clinic-morphological pictures and the immunological response to PPD and henna. The literature underlines that different clinical manifestations are related to black henna containing PPD, and its derivative products may cause delayed-type as well as immediate-type reactions. Further studies are needed to investigate the relationship between clinical and morphological aspects of PPD contact dermatitis and the T cell subsets predominance.     

Simultaneous determination of thiamphenicol, florfenicol and florfenicol amine in eggs by reversed-phase high-performance liquid chromatography with fluorescence detection

A specific, sensitive and widely applicable reversed-phase high-performance liquid chromatography with fluorescence detection (RP-HPLC-FLD) method was developed for the simultaneous determination of thiamphenicol (TAP), florfenicol (FF) and florfenicol amine (FFA) in eggs. Samples were extracted with ethyl acetate-acetonitrile-ammonium hydroxide (49:49:2, v/v), defatted with hexane, followed by RP-HPLC-FLD determination. Liquid chromatography was performed on a 5 μm LiChrospher C(18) column using a mobile phase composed of acetonitrile (A), 0.01 M sodium dihydrogen phosphate containing 0.005 M sodium dodecyl sulfate and 0.1% triethylamine, adjusted to pH 4.8 by 85% phosphoric acid (B) (A:B, 35:65 v/v), at a flow rate of 1.0 mL/min. The fluorescence detector of HPLC was set at 224 nm for excitation wavelength and 290 nm for emission wavelength. Limits of detection (LODs) were 1.5 μg/kg for TAP and FF, 0.5 μg/kg for FFA in eggs; limits of quantitation (LOQs) were 5 μg/kg for TAP and FF, 2 μg/kg for FFA in eggs. Linear calibration curves were obtained over concentration ranges of 0.025-5.0 μg/mL for TAP with determination coefficients of 0.9997, 0.01-10.0 μg/mL for FF with determination coefficients of 0.9997 and 0.0025-2.50 μg/mL for FFA with determination coefficients of 0.9998, respectively. The recovery values ranged from 86.4% to 93.8% for TAP, 87.4% to 92.3% for FF and from 89.0% to 95.2% for FFA. The corresponding intra-day and inter-day variation (relative standard deviation, R.S.D.) found to be less than 6.7% and 10.8%, respectively.     

Flame atomic emission spectrometric determination of aluminium in a bovine blood plasma matrix

A flame atomic emission spectrometric method, is described for the determination of aluminium in bovine blood plasma matrices. Plasma samples are wet-digested and solutions are aspirated into a conventional nitrous oxide-acetylene flame. Analyte emission is monitored at 396.15 nm with corrections for background emission being obtained from measurements several tenths nm on both sides of the aluminium line. The mean recovery of 0.3–5 μg/ml aluminium added to model solutions containing 500–5000 μg Na/ml, 50–1000 μg Ca/ml, 2000–5000 μg K/ml, or simulated plasma digests containing Na, K, and Ca was 100,6% (SD = 10.9, df = 60); the mean recovery of 0.3, 0.5, and 1.0 μg/ml aluminium added to blood plasma before digestion was 94.3% (SD = 9.8, df = 33) indicating no serious interferences. For standard solutions, the detection limit (signal: peak-to-peak noise = 1) was 0.02 μg/ml by flame emission, and 0.12 μg/ml by atomic absorption measurements with the same instrument. A sample taken through the analytical procedure, gave a detection limit of 0.05 μg/ml suggesting the submicrogram per milliliter region as the lower practical limit of the method.     

Determination of caffeic acid in rabbit plasma by high-performance liquid chromatography

A simple and sensitive high-performance liquid chromatographic method involving UV detection was developed for determination of caffeic acid in rabbit plasma. A Lichrosphere CN column (250 mm × 4 mm I.D., 5 μm) was used as the stationary phase and the mobile phase consisted of 2% acetic acid solution at a flow-rate of 1.0 ml/min. The UV absorbance was monitored at 320 nm. The plasma sample was acidified by the addition of 0.01 parts of concentrated phosphoric acid (85%) to maintain caffeic acid stability. After a simple clean-up procedure, the limit of quantitation achieved was 0.1 μg/ml, and the standard curve was found to be linear over the concentration ranges of 0.1–2.0 μg/ml and 0.1–40 μg/ml. The coefficient of variation for within- and between-run precision and accuracy was less than 10%, and the recovery was 82.3%.     

HPLC-ELSD法测定家桑野桑植株中1-脱氧野尻霉素的含量

采用HPLC-ELSD法对桑植株各部位所含的1-脱氧野尻霉素(DNJ)进行了含量测定,使用TSKgel Am-ide-80(4.6 mm×250 mm,5μm)色谱柱,流动相为:乙腈-水(84∶16,6.5 mM醋酸铵),流速为1.00 mL/min,柱温:30℃,线性范围为0.505μg～20.2μg(r=0.9991,n=6),平均回收率为94.95%(n=5),RSD=1.98。测定结果表明,野桑叶、嫩枝和根皮部位的DNJ含量较高,而家桑的叶、枝皮和根皮中含量较高。本方法准确度高、重现性好,能快速检测桑树资源及类似植物和相关产品中1-脱氧野尻霉素的含量。     

Flow-injection-electrochemical oxidation fluorimetry for determination of methotrexate.

A novel fluorimetric model of on-line electrochemical oxidation for the determination of methotrexate (MTX) is described in this paper. The method was based on the oxidation of MTX to a highly fluorescent product, 2,4-diaminopteridine-6-carboxylic acid, by constant potential in the electrochemical oxidation flow cell. Stopped-flow techniques were employed. Under optimal conditions, the fluorescence intensity of 2,4-diaminopteridine-6-carboxylic acid was measured at excitation and emission wavelengths of 380 and 465 nm, respectively. The calibration graph was linear over concentrations of methotrexate in the range 2.0 x 10(-7)-1.0 x 10(-5) g/mL, with a detection limit of 5.2 x 10(-8) g/mL (3sigma). The method has been successfully applied to the determination of methotrexate in human urine samples and showed a percentage recovery in the range 94.3-102.5%.     

Flow injection analysis of tetracyclines using inhibited Ru(bpy)3(2+)/tripropylamine electrochemiluminescence system.

Tetracyclines (TCs) were found to strongly inhibit the electrochemiluminescence (ECL) from the Ru(bpy)3(2+)-tripropylamine system when a working Pt electrode was maintained at 1.05 V (vs. Ag/AgCl) in pH 8.0 carbonate buffer solution. On this basis, a flow injection (FI) procedure with inhibited electrochemiluminescence detection has been developed for the determination of tetracycline (TC) and oxytetracycline (OTC). Under the optimized condition, the linear ranges of 2.0 x 10(-8)-1.0 x 10(-5) and 1.0 x 10(-8)-1.0 x 10(-5) g/mL and the detection limits of 4.0 x 10(-9) and 3.8 x 10(-9) g/mL were obtained for TC and OTC, respectively. The relative standard deviations (RSD) were 0.68% and 1.18% for 5.0 x 10(-7) g/mL TC and OTC (n = 13), respectively. The method showed higher sensitivity than most of the reported methods. It was successfully applied to the determination of tetracycline in a Chinese proprietary medicine, Tetracyclini and Cortisone Eye Ointment, and the residues of tetracycline in honey products. The inhibition mechanism has been proposed due to an energy transfer between electrogenerated Ru(bpy)3(2+)* and benzoquinone derivatives at the electrode surface.     

RP-HPLC法测定青海产四种秦艽中獐牙菜苦苷的含量

建立了秦艽中獐牙菜苦苷含量测定的RP-HPLC方法。色谱条件：采用ZorbaxEclipseXDB-C18柱（150mm×4.6mm,5μm）,流动相：甲醇-0.03%磷酸水溶液梯度洗脱（0～25min：15%～23%;25～30min：23%～30%）,流速：1mL/min,柱温：25℃,检测波长240nm,獐牙菜苦苷在0.20～1.8μg范围内成良好的线性关系,r=0.9999,回收率RSD=1.32%。对青海产4种秦艽中獐牙菜苦苷的含量进行了定量分析,测定结果：小秦艽中獐牙菜苦苷的含量最高为：0.62%,麻花秦艽与管花秦艽的含量没有显著差异,分别为：0.43%,0.45%,簧管秦艽的含量最低为：0.32%。     

Development of a high-performance liquid chromatography method for the simultaneous quantification of four organoarsenic compounds in the feeds of swine and chicken

A high-performance liquid chromatography (HPLC) method with UV detection was developed for the simultaneous determination of arsanilic acid, roxarsone, nitarsone, and carbarsone in the feeds of swine and chicken. Feed samples were extracted with methanol/1% acetic acid (90:10, v/v) in an ultrasonic bath and the protein was precipitated with 2% Cu(2)SO(4). The samples were further purified by solid phase extraction (SPE) on SAX cartridges. Separation was performed on a Zorbax SB-Aq C18 HPLC column using an isocratic procedure with methanol and 1% acetic acid (3:97, v/v) at a flow-rate of 0.7 mL min(-1), and the UV detector was set at a wavelength of 260 nm. The recoveries of organoarsenic compounds spiked at levels of 2, 20 and 200 μg g(-1) ranged from 81.2% to 91.3%; the inter-day relative standard deviation values were less than 7.0%. The limits of quantification for four organoarsenic compounds were 1.0-2.0 μg g(-1). This simple and fast method could be applied to the determination of multi-residues of organic arsenic compounds in animal feeds.     

Simultaneous analysis of amoxicillin and sulbactam in human plasma by HPLC-DAD for assessment of bioequivalence

A simple and accurate high-performance liquid chromatography with diode array detection-based (HPLC-DAD) method has been developed and validated for simultaneous determination of amoxicillin and sulbactam in human plasma. Sample preparation was involved in protein precipitation with acetonitrile followed by one-step extraction procedure. Chromatographic separation was achieved on a C18 column with an isocratic mobile phase consisting of water (containing 30 mM potassium dihydrogen phosphate, pH 2.8) and acetonitrile. The detection wavelengths of a diode array detector were set at 210 nm for amoxicillin and sulbactam, and 263 nm for the internal standard (cefadroxil). The method was validated for linearity, accuracy, precision, and stability. The calibration curve was linear from 0.163 to 14.7 μg/mL with correlation coefficient squared of 0.9991 for amoxicillin and 0.250-15.0 μg/mL with correlation coefficient squared of 0.9988 for sulbactam using 500 μL plasma samples. The lower limit of quantification was 0.163 and 0.250 μg/mL for amoxicillin and sulbactam, respectively. The imprecisions of intra- and inter-day validations for amoxicillin and sulbactam were <11% and their accuracies (%) were within the range of 95.4-105.7%. Mean recoveries were 75.9, 72.8, and 70.0% for amoxicillin, sulbactam, and cefadroxil, respectively. The established method was successfully applied to a bioequivalence study of two combination formulations of amoxicillin and sulbactam pivoxil in healthy male volunteers.     

UPLC-UV法测定不同产地青蒿中青蒿素的含量

目的:建立快速简便检测青蒿素的超高效液相-紫外(UPLC-UV)法,并对不同产地青蒿中青蒿素的含量进行检测。方法:色谱柱Agilent Eclipse Plus C18(2.1 mm×50 mm,1.8μm),流动相乙腈/水(45/55),流速1.0mL/min,柱温28℃,波长200 nm。结果:该方法对青蒿素的分离度较好,保留时间缩短为1.5 min。并且,整个分析过程可以在2 min内完成。线性范围0.101 17～10.117μg,进样量与峰面积线性相关,A=109.4C+6.7026,R2=0.9 993(n=9),加样回收率为99.3%(RSD=2.6%,n=6)。结论:UPLC-UV法分析时间短、样品前处理简单、精密度、稳定性、加样回收率等符合分析检测要求,对于青蒿中青蒿素的含量能进行快速准确的分析。     

Rapid and selective liquid chromatographic/tandem mass spectrometric method for the determination of fosfomycin in human plasma

A rapid and selective liquid chromatographic/tandem mass spectrometric method for determination of fosfomycin was developed and validated. Following protein-precipitation, the analyte and internal standard (fudosteine) were separated from human plasma using an isocratic mobile phase on an Ultimate XB-CN column. An API 4000 tandem mass spectrometer equipped with Turbo IonSpray ionization source was used as detector and was operated in the negative ion mode. Multiple reaction monitoring using the precursor to product ion combinations of m/z 137-->79 and m/z 178-->91 was performed to quantify fosfomycin and fudosteine, respectively. The method was linear in the concentration range of 0.10-12.0 microg/mL using 50 microL of plasma. The lower limit of quantification was 0.10 microg/mL. The intra- and inter-day relative standard deviation over the entire concentration range was less than 10.6%. Accuracy determined at three concentrations (0.25, 1.00 and 8.00 microg/mL for fosfomycin) ranged from -1.0% to -4.2% in terms of relative error. Each plasma sample was chromatographed within 5.0 min. The method was successfully used in a bioequivalence study of fosfomycin in human plasma after an oral administration of capsules containing 1.0 g fosfomycin (approximately 1.3g calcium fosfomycin).     

Studies on decolorization process for rhEGF as cosmetic ingredient

A decolorization process using ion exchange chromatography was developed to refine rhEGF as a cosmetic ingredient. A macroreticular resin (D314) was selected, with respect to its high decolorization rate and recovery yield of rhEGF, and the operational conditions of the decolorization process optimized. The optimum conditions were as follows: the rhEGF effluent was ion exchanged at a flow rate of 60.0 mL/h, with an effluent pH 5.0, using a chromatographic column (i.d. 16mm) packed with D314, with a 7.5 cm in bed height. The decolorization process was carried out under the optimum conditions, and halted when the effluent volume reached 350 mL, giving a decolorization rate and recovery yield of rhEGF higher than 67 and 80%, respectively. When the decolorization rate exceeded 67%, the final product turned out to be white or light yellowish, which was to the satisfaction of the cosmetic standard.     

Chemiluminescence of synephrine based on the cerium(IV)-rhodamine B system.

A new chemiluminescence (CL) method is described for the determination of synephrine. It is based on the reaction between synephrine and Ce(IV) in a nitric acid medium and measurement of the CL intensity produced by rhodamine B used as a luminophore, similar to luminol or lucigenin in basic media, instead of as a sensitizer. In the optimum conditions, the increase of CL intensity was correlated with synephrine concentration over the range 5.0 x 10(-9)-1.0 x 10(-6) g/mL with a detection limit of 1.0 x 10(-9) g/mL. The relative standard deviation (RSD) was 2.9% for 1.0 x 10(-7) g/mL synephrine (n = 11). The method was applied to the determination of a drug in herbal products, citrus fruit and biological samples, with satisfactory results. The results given by the proposed method are in good agreement with those given by HPLC-UV and UV spectrophotometry.     

高效液相色谱法测定重组人干扰素α-2b注射液中乙二胺四乙酸二钠含量

目的:利用高效液相色谱(HPLC)法测定重组人干扰素α-2b注射液中EDTA二钠(乙二胺四乙酸二钠)的含量。方法:将EDTA二钠与氯化铁溶液于70℃水浴中反应20 min左右;色谱柱为SunFire C18(250 mm×4.6 mm,5μm,Waters),流动相为5%甲醇+95%0.64 g/L四丁基溴化铵和4.1 g/L三水合乙酸钠混合液,用冰乙酸调pH值至4.0,流速1 mL/min,检测波长254 nm。结果:该测定方法线性范围为0.025~0.5 g/L,线性关系良好(r=0.9997),加样回收率为98.27%(n=9,RSD=2.55%)。结论:本方法准确、快速、可靠,可用于重组人干扰素α-2b注射液中EDTA二钠含量的测定。     

高效液相色谱法测定川射干鸢尾苷元磺酸钠及其制剂泰克吉宁注射液含量和有关物质

建立高效液相色谱法测定鸢尾苷元磺酸钠(4′,5,7-三羟基-6-甲氧基异黄酮-5′-磺酸钠)及其制剂泰克吉宁注射液的含量及有关物质。采用C18色谱柱(150×4.6mm,5μm),流动相为甲醇-5%醋酸水(80:20),流速为1.0mL/min,检测波长为263nm。鸢尾苷元磺酸钠在11～100μg/mL范围内线性关系良好,回归方程Y=43.3609X-1.5973(r=0.99998),平均回收率为99.77%(n=9)。最低检测限为0.052μg/mL,与有关物质分离良好。本法快速、简便、准确,专属性强。     

Liquid chromatography/tandem mass spectrometry assay for the simultaneous determination of cefoperazone and sulbactam in plasma and its application to a pharmacokinetic study

A rapid and highly sensitive liquid chromatography-tandem mass spectrometric (LC-MS/MS) method for simultaneous determination of cefoperazone sodium and sulbactam sodium in human plasma was developed. The analytes and internal standard (IS), cefuroxime sodium, were extracted from human plasma via liquid-liquid extraction with ethyl acetate and separated on a Waters Xterra C18 column within 3.5 min. Quantitation was performed on a triple quadrupole mass spectrometer employing electrospray ionization technique, operating in selected reaction monitoring (SRM) and negative ion mode. The precursor to product ion transitions monitored for cefoperazone, sulbactam and IS were m/z 644.1→528.0, 232.1→140.0, and 423.0→362.0, respectively. The assay was validated in the linear range of 0.1-20 μg/mL for cefoperazone and 0.02-4 μg/mL for sulbactam. The intra- and inter-day precisions (CV%) were within 8.39% for each analyte. The recoveries were greater than 87.3% for cefoperazone and 87.2% for sulbactam. Each analyte was found to be stable during all sample storage, preparation and analytical procedures. The method was successfully applied in a pharmacokinetic study of Sulperazon injection in six hospital-acquired pneumonia (HAP) patients.     

Development and validation of a selective and sensitive LC-MS/MS method for determination of cycloserine in human plasma: application to bioequivalence study

A selective and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay for the determination of cycloserine in human plasma is developed using niacin as internal standard (IS). The analyte and IS were extracted from 500 μL of human plasma via solid phase extraction on Waters Oasis MCX cartridges. Chromatographic separation was achieved on a Peerless Basic C18 (100 mm × 4.6mm, 3 μm) column under isocratic conditions. Detection of analyte and IS was done by tandem mass spectrometry, operating in positive ion and multiple reaction monitoring (MRM) acquisition mode. The protonated precursor to product ion transitions monitored for cycloserine and niacin were at m/z 103.1 → 75.0 and 124.1 → 80.1 respectively. The method was fully validated for its selectivity, interference check, sensitivity, carryover check, linearity, precision and accuracy, reinjection reproducibility, recovery, matrix effect, ion suppression/enhancement, stability and dilution integrity. The limit of detection (LOD) and lower limit of quantitation of the method were 0.0013 and 0.20 μg/mL respectively with a linear dynamic range of 0.20-30.00 μg/mL for cycloserine. The intra-batch and inter-batch precision (%CV) across six quality control levels was less than 8.0% for cycloserine. The method was successfully applied to a bioequivalence study of 250 mg cycloserine capsule formulation in 24 healthy Indian male subjects under fasting condition.     

Validated Method for the Quantification of Atractylenolide III in Different Processed Products of Rhizoma Atractylodes Macrocephalae

Introduction – Rhizoma Atractylodes Macrocephalae (RAM) contains several sesquiterpene compounds including atractylenolide III (AO‐III). This bioactive compound may be used as a chemical marker for the quality control of different processed RAM products. Objective – To develop and validate an RP‐HPLC method for the quantitative determination of AO‐III in RAM and in a variety of processed RAM products. Methodology – HPLC was carried out using a Kromssil C₁₈ RP‐column eluted with methanol–water (70:30) at a flow rate of 1.0 mL/min and with UV detection at 220 nm. Full validation was performed using standard methods. Results – The linear range of AO‐III was 5–50 µg/mL; the regression equation was y = 10210x + 11194 (r = 0.9994) and the average recovery was 101.08% (RSD = 0.98%). The detection and quantification limits for AO‐III were determined to be 0.005 and 0.018 µg/mL at signal‐to‐noise ratios of approximately 3:1 and 10:1, respectively. Conclusion – The described HPLC method is appropriate for quality assurance and differentiation of AO‐III in RAM and different processed products.     

RP-HPLC法测定泽兰中桦木酸的含量

目的：建立反相高效液相色潽法测定泽兰中桦木酸的含量.方法：采用AlltimaTM-C18（250 mm×4.6 mm,5μm）;以甲醇-0.2％磷酸水溶液=85：15进行等度洗脱,流速为1.0 mL/min,检测波长为202 nm,柱温为35℃.结果：桦木酸的线性范围为3.968～99.200 μg（r=0.997 3）,平均回收率为100.87％.结论：该方法结果准确、简便可行、重复性好,可为泽兰的质量控制提供依据.