On-line coupling of supercritical CO2 extraction with reversed-phase liquid chromatography for the quantitative analysis of analytes in aqueous matrices

The first report of on-line coupled supercritical fluid extraction (SFE) with reversed-phase liquid chromatography for the quantitative analysis of analytes in aqueous matrices is described. Two commercial systems (e.g. SFE and HPLC) were connected via a single six-port injection valve. By using water to eliminate residual decompressed CO2 gas in the solid-phase extraction trap, quantitative extraction and transfer were achieved for the target analytes (progesterone, phenanthrene, and pyrene) spiked in water, as well as in real samples (urine and environmental water). During each extraction, no restrictor plugging was realized. Extraction temperature and pressure were optimized. Different amounts of salt were added to the aqueous matrix to enhance ionic strength and thus extraction efficiency. Methanol and 2-propanol were used as CO2 modifiers. Compared with dynamically mixing modifier with the CO2 extraction fluid, pre-spiking the same amount of modifier in the extraction vessel enhanced the recovery approximately 30% for progesterone, phenanthrene, and pyrene due to a "co-extraction effect".     

黄鳝肌肉中11种性激素的气质联用检测方法

报道了一种同时筛选、定量和确证黄鳝中的11种性激素的灵敏、特异可靠的检测方法。对5.0 g肌肉样品,采用β-葡糖苷酸酶(来自Helix pomatia)对提取物水解后,经叔丁基甲醚提取,正己烷脱脂,再经C18小柱和NH2小柱净化,七氟丁酸酐对其进行衍生,最后利用GC-MS进行检测。采用外标法定量,通过对空白样品添加标准品,使之理论含量为1.00,2.50和5.00μg.kg-1,检出限在0.50~1.00μg.kg-1,平均回收率范围为56%~103%,平均相对标准偏差范围为3.1%~29.5%。对不同批次添加水平为1.00μg.kg-1的回收率进行比较,相对标准偏差范围为4.8%~22.2%,重复性好。因此,本方法对目前动物源食品中的性激素监督检验非常有用。     

Sub-nanogram per litre detection of the emerging contaminant progesterone with a fully automated immunosensor based on evanescent field techniques

Progesterone is reasonably anticipated to be a human carcinogen based on sufficient evidence of carcinogenicity in experimental animals and it can be found in various surface waters which are partly used as drinking water resources. Therefore, immunoanalytical methods at a very low limit of detection (LOD) and a low limit of quantification (LOQ) are becoming more and more important for environmental analysis and especially for monitoring drinking water quality. Biosensors have suitable characteristics such as efficiency in allowing very fast, sensitive, and cost-effective detection. Here we describe a fully automated immunoassay for progesterone with a LOD in the sub-nanogram per litre range and a LOQ in the lower nanogram per litre range. In contrast to common analytical methods such as GC-MS or HPLC-MS, the biosensor used requires no sample pre-treatment and no sample pre-concentration. The basis of our sensitive assay is the antibody with a high affinity constant towards progesterone and the robust biosensor setup used.     

Development of a competitive lateral flow immunoassay for progesterone: influence of coating conjugates and buffer components

Several aspects of the development of competitive lateral flow immunoassays (LFIAs) are described. The quantitation of progesterone is taken as an example. The LFIA format consisted of a nitrocellulose membrane spotted with various progesterone conjugates as the test line. A mixture of primary antibody and secondary antibody adsorbed to colloidal carbon was used for signal generation. A digital scanner and dedicated software were used to quantitate the response. A reappraisal of the checkerboard titration, often used in the optimisation of immunoassays, is discussed. Surprisingly, the highest sensitivity of the LFIA format (IC₅₀ of 0.6 μg L⁻¹ progesterone in buffer) was achieved by using a high coating concentration of the analyte–protein conjugate and a high dilution of the antibody solution. Immediate addition of all reagents in LFIA was superior to premixing the components and allowing prereaction. Of several blocking agents tested bovine serum albumin was superior in performance, whereas the combination of ovalbumin and progesterone substantially influenced test results.     

小鼠子宫腺肌病模型病灶中雌孕激素受体的表达及意义

目的探讨雌激素受体(ER)和孕激素受体(PR)在子宫腺肌病模型病灶中的表达及临床意义.方法用异体垂体移植法建立ICR小鼠子宫腺肌病模型,应用免疫组化SP法检测正常子宫内膜、模型子宫内膜及病灶异位子宫内膜中ER、PR的表达.结果垂体移植4个月和6个月后,均成功诱发子宫腺肌病模型,不同时间诱发模型的病灶大体结节数和HE染色评分之间比较,均无统计学差异(均P>0.05);建模6个月后模型病灶的结节显著大于建模4个月后(P<0.05).建模后4个月,模型子宫内膜和病灶异位子宫内膜以及正常子宫内膜中ER和PR表达的阳性率比较,均无统计学差异(均P>0.05);建模后6个月,3种子宫内膜中ER阳性率分别为55.07%、31.63%和24.74%;PR阳性率为48.40%、27.48%和22.70%;3者比较均有统计学差异(均P<0.05).结论子宫腺肌病的发生、发展可能与子宫内膜局部ER、PR的高表达相关.     

A convenient enantioselective synthesis of 3-asymmetrically substituted oxindoles as progesterone receptor antagonists

A convenient enantioselective synthesis of 3-asymmetrically substituted oxindoles is reported. Compound (2) prepared by radical cyclisation of (1) was used for the synthesis of racemic and enantiomerically pure 3-asymmetrically substituted oxindoles. Desulfurisation of (2) using Raney Ni yielded the racemate (5). Addition of (S)-1-phenylethanol to compound (2) yielded the diastereoisomer (21) the structure of which was determined using X-ray crystallography. Using a sequence of steps (21) was converted to the enantiomer (8). The enantiomer (9) was similarly prepared from (2) using (R)-1-phenylethanol.     

Synthesis of 6-Hydroxy Derivatives of Steroidal Hormones by SeO<Subscript>2</Subscript> Mediated Oxidation

Summary. Selenium dioxide oxidation of molecules with cyclopentanoperhydrophenanthrene skeleton and allylic moieties, such as the well known human steroidal hormones progesterone and testosterone enables the syntheses of potential active 6-hydroxysteriods.Current address: Institute of Pharmacy, Department of Pharmacognosy, University of Innsbruck, A-6020 Innsbruck, Austria     

Towards the Expression of Sex Hormone Receptors in the Human Vocal Fold

BACKGROUND: The human larynx is assumed to be a steroid receptor target organ. There are only very limited data on the evidence of steroid receptors in the vocal folds, although voice alterations due to hormonal influence and treatment have been found. GOAL OF THE STUDY: To investigate the expression of estrogen alpha, progesterone, and androgen receptors in human vocal folds (vocalis muscle, glands, lamina propria, epithelium). METHODS: Immunohistochemically, vocal fold cadaver specimens of 15 autopsied patients (6 women, 9 men), which were taken approximately 4 to 8 hours postmortem were investigated. Furthermore, one (male) vocal fold biopsy obtained intraoperatively during a laryngectomy was tested. RESULTS: No specific immunohistochemical staining for the different types of steroid hormones investigated could be observed in either the postmortem taken biopsies nor the intraoperatively one. However, several unspecific staining patterns could be observed. CONCLUSION: The results of this study contradict recently published data and question the expression of sex hormone receptors in the vocal folds. Main causes of false interpretations of unspecific staining are discussed.     

Preliminary report on hormone receptors in the human vocal fold

There has been an ongoing effort to describe the physiologic factors associated with perceived and/or measured human voice changes that occur with age. In our study we focused on possible endocrine involvement on voice by using immunohistochemical staining to observe hormone receptor presence in vocal folds from 42 deceased subjects (fresh cadavers), male and female, ranging in age from 2 months to 82 years (average 37.7 years). On statistical analysis, age and gender were found to be associated with progesterone receptor staining of the glandular nuclei (young>old P = 0.013; male>female, P = 0.060). Gender was associated with androgen receptor staining in glandular cytoplasm (male>female, P = 0.014) and progesterone receptor staining in the epithelial cytoplasm (male>female, P = 0.039). No statistical significance was noted in other categories. CONCLUSION: Hormone receptors are found in the nucleus and cytoplasm of cells in the vocal fold with statistically significant differences in age and gender distribution.