Analytical procedure to simultaneously measure trace amounts of trenbolone acetate and beta-trenbolone residues in porcine muscle using HPLC-UVD and MS

The current study was undertaken to validate the performance for the determination of both TBA and beta-trenbolone (beta-TB) residues in porcine muscle at concentrations required to monitor compliance with the maximum residue limit (MRL). The method involves a one phase liquid-liquid extraction, cleanup with low-temperature fat precipitation, separation of the respective compounds by HPLC on a Capcell pak C(18) column, use of a methanol-water isocratic system as an eluent, and measurement by UV absorbance detection at 340 nm. Both compounds were confirmed using LC-MS/MS with electrospray interface (ESI) and a triple quadrupole (QqQ) analyzer. The method was found to be precise and accurate, with a linearity range of 1-10 microg/kg (r(2) >0.973). The intra- and interday precision showed good reproducibility with RSDs < or =13.25%. The LODs were 0.12 and 0.22 microg/kg, and the LOQs were 0.37 and 0.66 microg/kg, for TBA and beta-TB, respectively. The applicability of the method was demonstrated by analyzing real samples collected from major cities in the Republic of Korea. No residues of the selected compounds were detected in any of the samples. The advantages of our method are that it is: selective, sensitive, requires a short time for analysis (13 min), and performs simple sample extraction and clean-up procedure with low-temperature fat precipitation as compared to the previously published methods.     

Structure-activity relationships in cholapod anion carriers: enhanced transmembrane chloride transport through substituent tuning

Chloride transport by a series of steroid-based "cholapod" receptors/carriers was studied in vesicles. The principal method involved preincorporation of the cholapods in the vesicle membranes, and the use of lucigenin fluorescence quenching to detect inward-transported Cl-. The results showed a partial correlation between anion affinity and transport activity, in that changes at the steroidal 7 and 12 positions affected both properties in concert. However, changes at the steroidal 3-position yielded irregular effects. Among the new steroids investigated the bis-p-nitrophenylthiourea 3 showed unprecedented activity, giving measurable transport through membranes with a transporter/lipid ratio of 1:250 000 (an average of <2 transporter molecules per vesicle). Increasing transporter lipophilicity had no effect, and positively charged steroids had low activity. The p-nitrophenyl monourea 25 showed modest but significant activity. Measurements using a second method, requiring the addition of transporters to preformed vesicle suspensions, implied that transporter delivery was problematic in some cases. A series of measurements employing membranes of different thicknesses provided further evidence that the cholapods act as mobile anion carriers.     

25‐Hydroxyvitamin D3 Synthesis by Enzymatic Steroid Side‐Chain Hydroxylation with Water

The hydroxylation of vitamin D₃ (VD₃, cholecalciferol) side chains to give 25‐hydroxyvitamin D₃ (25OHVD₃) is a crucial reaction in the formation of the circulating and biologically active forms of VD₃. It is usually catalyzed by cytochrome P450 monooxygenases that depend on complex electron donor systems. Cell‐free extracts and a purified Mo enzyme from a bacterium anaerobically grown with cholesterol were employed for the regioselective, ferricyanide‐dependent hydroxylation of VD₃ and proVD₃ (7‐dehydrocholesterol) into the corresponding tertiary alcohols with greater than 99 % yield. Hydroxylation of VD₃ strictly depends on a cyclodextrin‐assisted isomerization of VD₃ into preVD₃, the actual enzymatic substrate. This facile and robust method developed for 25OHVD₃ synthesis is a novel example for the concept of substrate‐engineered catalysis and offers an attractive alternative to chemical or O₂ /electron‐donor‐dependent enzymatic procedures.     

Catalytic Transformation of Oxygenated Organic Compounds into Pure Hydrogen

The continual growth in transportation fuels and more strict environmental legislations have led to immense interest in developing green biomass energy. In this work, a proposed catalytic transformation of oxygenated organic compounds (related to bio-oil) into pure hydrogen was desighed, involving the catalytic reforming of oxygenated organic compounds to hydrogen-rich mixture gas followed by the conversion of CO to CO₂ via the water gas reaction and the removal of CO₂. The optimization of the different reforming catalyst, the reaction conditions as well as various sources of oxygenated organic compounds were investigated in detail. The production of pure hydrogen, with the H₂ content up to 99.96% and the conversion of 97.1%, was achieved by the integrated catalytic transformation. The reaction pathways were addressed based on the investigation of decomposition, catalytic reforming, and the water gas reaction.     

含氧有机物催化重整制备纯氢

研究了一种新的利用含氧化合物制备纯氢的催化变换过程,该过程耦合了含氧化合物的催化重整、水煤气变换反应和CO₂去除步骤. 详细研究了重整催化剂的筛选、反应条件以及不同的含氧化合物催化重整行为. 利用所述集成方法获得的最高氢气浓度为99.96vol%和最大转化率为97.1mol%. 此外,通过含氧化合物的解离、催化重整和水煤气变换反应研究,探讨了含氧化合物制备纯氢的相关反应路径.     

Application of liquid chromatography coupled with electrospray ionization time-of-flight mass spectrometry for screening and quantitative analysis of C21 steroids in the roots and rhizomes of Cynanchum paniculatum

A liquid chromatography-mass spectrometry method is presented for the quantification of C21 steroids in the roots and rhizomes of Cynanchum paniculatum. Eight C21 steroids, including five steroidal aglycones and three steroidal glycosides, were simultaneously analyzed by liquid chromatography coupled with electrospray ionization time-of-flight mass spectrometry. The extracted ion current chromatograms were extracted from the total ion current chromatogram using characteristic ions produced by target compounds for peak determination. Chromatographic separation was achieved on a C18 reversed-phase column within 60 min, using an acetonitrile/water gradient. For comparision, six C. paniculatum samples from different locations were investigated by the established method, and the results indicated that the different geographical origin significantly influenced the C21 steroid composition. The method was observed to have the necessary sensitivity, selectivity, precision, and accuracy, and to be suitable for quality control of herbal medicines and their preparations.     

Development and validation of a qualitative screening method for the detection of exogenous anabolic steroids in urine by liquid chromatography-tandem mass spectrometry

A screening method for the urinary detection of 34 exogenous anabolic steroids has been developed. The method involves an enzymatic hydrolysis, liquid-liquid extraction and detection by liquid chromatography-tandem mass spectrometry. The use of some adducts such as [M+NH₄]⁺, [M+CH₃COO]⁻ and [M+H+MeOH]⁺ was necessary in order to detect some analytes at the required level (lower than 10 ng/ml). Two transitions were selected for each analyte. Different concentration factors have been studied in order to increase the sensitivity. A concentration factor of 50 was selected for the screening method although the high ion suppression observed under these conditions can hamper its application as a quantitative method. The method was validated and limits of detection were obtained by spiking ten different blank urine samples at five different concentration levels. Up to 29 analytes were detected in all spiked urines at the required level. Limits of detection between 1 and 10 ng/ml were obtained for most analytes which fulfil current requirements. The applicability of the method was shown by analysing positive samples.     

Ionization of anabolic steroids by adduct formation in liquid chromatography electrospray mass spectrometry

The ionization of 46 anabolic steroids has been studied. The absence of basic or acidic moieties in most of these analytes makes their direct ionization as [M + H]+ by atmospheric pressure interfaces difficult. The formation of adducts with different components of the mobile phase has been found to be an efficient way to ionize anabolic steroids by electrospray. Different mobile phases using methanol (MeOH) or acetonitrile as organic solvent and HCOOH, Na+ or NH4+ as additives have been tested to favor the adduct formation. A direct correlation between the chemical structure of the anabolic steroid and the possibility to ionize it in a particular chromatographic condition has been found. According to their ionization, anabolic steroids can be divided into seven different groups depending on both the nature and the relative position of their functional groups. The formation of different adducts such as [M + Na + MeOH]+ or [M + H + CH3 CN - H2O]+ is required in order to ionize some of these groups and the optimal mobile phase composition for each group of anabolic steroids is proposed. Despite the ionization limitations due to their chemical structure, most of tested anabolic steroids could be ionized using the adduct formation approach.