Automated online dried blood spot sample preparation and detection of anabolic steroid esters for sports drug testing

Dried blood spots (DBSs) provide a valuable complementary sample matrix for routine doping analysis, and the full automation of analysis for DBS samples is achievable to avoid extensive and repetitive manual laboratory work. In the current study, a fully automated online DBS preparation and detection method for the screening and quantification analysis of 13 anabolic steroid esters by means of the DBSA-TLX-HRMS system was developed and validated, based on the purpose for the determination of the abuse of anabolic steroid esters in athletes. Validation of the method yielded linear (R² > 0.99), precise, and accurate (RSD% and Re% < 20% at low, medium, and high concentration levels) results. The LOD of testosterone laurate was established at 0.5 ng/ml and at 0.2 ng/ml for other steroid esters. The extraction recovery of the target compounds from DBS ranged from 10.5% to 88.9%, and matrix effects were moderate. Furthermore, the developed and validated method was applied in the analysis of DBS samples collected after the oral administration of a single dose of 80 mg testosterone undecanoate demonstrating its applicability. Evaluation of analyte stability showed that testosterone undecanoate are more stable (8 weeks) in DBS samples of administration study when stored in frozen (−20°C) condition compared with cold storage (4°C). Collectively, these findings demonstrate the applicability of automated DBS analysis in doping control for detection of anabolic steroid esters.     

Abnormal cardiac repolarization in anabolic androgenic steroid users carrying out submaximal exercise testing

1. The aim of the present study was to investigate the cardiovascular effects of anabolic androgenic steroid (AAS) abuse by comparing the electrocardiographic parameters before and after submaximal exercise between AAS users and non-AAS users. 2. A total of 22 men who regularly engaged in both resistance and aerobic exercise at fitness academies volunteered for the study (control group: n = 11, age 25 ± 4 years; AAS group: n = 11, age 27 ± 5 years). All subjects were submitted to submaximal exercise testing using an Astrand-Rhyming protocol. Heart rate and electrocardiography parameters were measured at rest and at the third minute of the post-exercise recovery time. 3. AAS users presented higher QTc and QTd at rest (10% and 55%, respectively) and at the post-exercise period (17% and 43%, respectively), compared with control subjects. The maximal and minimum QTc interval of the AAS group was significantly prolonged at the post-exercise period (12% and 15%, respectively). The haemodynamic parameters were similar in both groups (P > 0.05). The AAS group showed a lower heart rate recovery at the first minute after the test (P = 0.0001), and a higher exertion score (P < 0.0001) at a lower workload, compared with the control group. 4. Our results show that the QTc interval and dispersion are increased in individuals who abuse AAS, suggesting the presence of ventricular repolarization abnormalities that could potentially increase the risk of cardiac arrhythmias and sudden cardiac death.     

类固醇糖尿病;糖皮质激素;胰岛素

目的:分析儿童类固醇性糖尿病的临床特点,提高对该病的认识。方法:回顾性分析山西省儿童医院收治的2例类固醇糖尿病患儿临床表现、体征、实验室检查结果。结果:2例儿童类固醇性糖尿病均无多饮、多尿及体质量减轻等糖尿病典型临床表现,均以餐后血糖明显升高为主要临床表现。1例患儿于糖皮质激素治疗2个月后发生,激素总量1 700 mg,胰岛素治疗7周后血糖恢复正常;另1例患儿于激素治疗20 d后发生,激素总量1 000 mg,激素减量12周后,血糖恢复正常。结论:大剂量长疗程使用糖皮质激素易引起类固醇糖尿病,激素减量可能使血糖恢复正常,胰岛素为治疗类固醇糖尿病的首选药物。     

Cytotoxic steroid derivatives from the Vietnamese soft coral Sinularia brassica

Using various chromatographic separations, six ergostane-type steroids, including one new compound sinubrassione (1), and two pregnene-type steroid glycosides, including one new compound sinubrassioside (7), were isolated from methanol extract of the Vietnamese soft coral Sinularia brassica. The structure elucidation was confirmed by spectroscopic methods including 1D, 2D NMR and HR-ESI-MS. The cytotoxic activities of all the isolated compounds against three human cancer cell lines were also evaluated using MTT-based colorimetric assays.     

果叶马尾藻中一个新的甾体成分

从采自文本北海的褐藻果叶马尾藻（Sargassum carpophyllum）中分离得到一个结构新颖的甾体化合物，经IR，MS，2DNMR等波谱技术中，确定为：24-乙基-3-乙氧甲酰基-3-羟基-A-失碳-5-，24（28）-胆甾二烯-2-酮。这是首次报道从天然界获得A-nor甾体羧酸酯类化合物。     

Direct analysis in real time ‐ high resolution mass spectrometry (DART‐HRMS): a high throughput strategy for identification and quantification of anabolic steroid esters

High throughput screening is essential for doping, forensic, and food safety laboratories. While hyphenated chromatography‐mass spectrometry (MS) remains the approach of choice, recent ambient MS techniques, such as direct analysis in real time (DART), offer more rapid and more versatile strategies and thus gain in popularity. In this study, the potential of DART hyphenated with Orbitrap‐MS for fast identification and quantification of 21 anabolic steroid esters has been evaluated. Direct analysis in high resolution scan mode allowed steroid esters screening by accurate mass measurement (Resolution = 60 000 and mass error < 3 ppm). Steroid esters identification was further supported by collision‐induced dissociation (CID) experiments through the generation of two additional ions. Moreover, the use of labelled internal standards allowed quantitative data to be recovered based on isotopic dilution approach. Linearity (R² > 0.99), dynamic range (from 1 to 1000 ng mL^‐1), bias (<10%), sensitivity (1 ng mL^‐1), repeatability and reproducibility (RSD < 20%) were evaluated as similar to those obtained with hyphenated chromatography‐mass spectrometry techniques. This innovative high throughput approach was successfully applied for the characterization of oily commercial preparations, and thus fits the needs of the competent authorities in the fight against forbidden or counterfeited substances. Copyright © 2014 John Wiley & Sons, Ltd.     

若干抗真菌甾体生物碱类似物的合成

为寻找新型抗真菌药物,从易得原料3β-乙酰氧基-5,16-孕甾二烯-20-酮(6)经10或11步反应合成了7个抗真菌甾体生物碱类似物,其中19个为新化合物,化合物3a～c和5体外试验抗真菌活性与氟康唑等同或更强。     

Effect of glucocorticoid administration on the steroid profile

The steroid profile (SP) is a powerful tool to detect the misuse of endogenous anabolic androgenic steroids in sports, and it is included in the Athlete Biological Passport (ABP). Glucocorticoids (GCs), which are widely prescribed in sports and only prohibited in competition by systemic routes, inhibit the hypothalamic‐pituitary‐adrenal axis. Since the metabolites monitored in the SP have a partial adrenal origin, their excretion in urine might be altered by GCs consumption. The aim of the present work was to investigate if GCs administered by either systemic or local routes could influence the SP parameters. Three of the most frequently detected GCs in sports (prednisolone, betamethasone, and triamcinolone acetonide) were administered to healthy male and female volunteers (n=40) using different administration routes (topical, oral, and intramuscular administration at different doses). In total, 66 administrations of GCs were performed. Urine samples were collected before and after GCs administration. The SP was measured using gas chromatography‐mass spectrometry. The excretion rates of the SP metabolites decreased after systemic GCs administration. This excretion decrease showed to be associated with the dose and the administration route. However, the individual evaluation of the SP ratios (T/E, A/T, A/Etio, 5αAdiol/5βAdiol, and 5αAdiol/E) led to normal sequences for all the conditions tested. Therefore, GCs administration did not produce misinterpretations on the ABP evaluation. According to these results, GCs administration should not distort the establishment of normal ranges of the SP ratios, and does not need to be considered a confounding factor in the SP evaluation.     

糖皮质激素依赖性皮炎36例临床分析

目的分析导致激素依赖性皮炎的原因并观察其治疗效果。方法采用我科2008年10月至2009年10月共诊疗的36例激素依赖性皮炎临床资料分析,并对其进行综合治疗观察。结果 36例患者中以中青年女性为主,长期滥用或误用激素类制剂导致皮肤对激素产生依赖为其主要病因。综合治疗治愈28例,显效4例,无效4例,总有效率88.89%。结论激素依赖性皮炎目前已成为皮肤科常见病,避免滥用或误用外用激素药物是预防本病的关键,多药联合治疗激素依赖性皮炎有良好效果。     

Comparative safety evaluation of selective androgen receptor modulators and anabolic androgenic steroids

Introduction: Anabolic androgenic steroids (AASs) have been in use for decades for the treatment of short stature, severe burns, HIV wasting syndrome, osteoporosis, and anemia. However, their lack of selective effects on certain symptoms and unfavorable pharmacokinetic properties has limited their long-term usage in clinics.

Areas covered: Selective androgen receptor modulators (SARMs) have some advantages over AASs; they are highly specific for androgen receptors, are orally available, and, most importantly, act as strong receptor agonists in skeletal muscle and bone, and as weak agonists or antagonists in androgen-responsive tissues such as the prostate and sebaceous glands. The exact molecular mechanism, however, has not been fully elucidated. This article includes a toxicological review of major AASs, and a comparative safety analysis of major AASs and SARMs in clinical trials to evaluate the therapeutic potential of SARMs.

Expert opinion: Based on the robust tissue selectivity of SARMs over AASs, they are worth considering as a promising therapeutic option for the treatment of various muscle-wasting diseases.     

对类固醇糖尿病临床特点的护理观察

目的：观察糖皮质激素继发糖尿病（类固醇糖尿病）的血糖水平变化特点。方法：对116例被诊断为类固醇糖尿病的药疹患者和与之相匹配的普通2型糖尿病患者进行动态血糖水平监测,评价并比较两组患者间血糖水平变化的特点和差异。结果：类固醇糖尿病患者以午餐后至睡前血糖水平升高为主要表现,空腹血糖水平一般正常,而2型糖尿病患者的全天平均血糖水平及其波动和低血糖症发生率均低于类固醇糖尿病患者,全天血糖水平达标时间明显长于类固醇糖尿病患者。结论：类固醇糖尿病患者的血糖水平变化具有自身的临床特点,与普通2型糖尿病患者的血糖水平变化明显不同,其血糖水平变化与激素作用高峰有关,临床干预中应注意这一特点并采取相应措施。     

对类固醇糖尿病临床特点的护理观察

目的:观察糖皮质激素继发糖尿病(类固醇糖尿病)的血糖水平变化特点。方法:对116例被诊断为类固醇糖尿病的药疹患者和与之相匹配的普通2型糖尿病患者进行动态血糖水平监测,评价并比较两组患者间血糖水平变化的特点和差异。结果:类固醇糖尿病患者以午餐后至睡前血糖水平升高为主要表现,空腹血糖水平一般正常,而2型糖尿病患者的全天平均血糖水平及其波动和低血糖症发生率均低于类固醇糖尿病患者,全天血糖水平达标时间明显长于类固醇糖尿病患者。结论:类固醇糖尿病患者的血糖水平变化具有自身的临床特点,与普通2型糖尿病患者的血糖水平变化明显不同,其血糖水平变化与激素作用高峰有关,临床干预中应注意这一特点并采取相应措施。     

甲泼尼龙和维库溴铵联用相关急性类固醇肌病

1例65岁男性因肺炎合并呼吸衰竭,给予高流量吸氧,亚胺培南-西司他丁钠0.5 g、1次/8 h静脉滴注,万古霉素0.5 g、1次/8 h静脉滴注,莫西沙星0.4 g、1次/d静脉滴注,甲泼尼龙160 mg/d静脉滴注。治疗3 d病情控制不佳,行气管插管和呼吸机机械通气,因患者出现躁动应用维库溴铵4 mg静脉推注,4～6 h重复给药(16～20mg/d),同时甲泼尼龙减量至80 mg/d静脉滴注。治疗5 d患者出现睁眼困难,肌力下降,四肢腱反射减弱,肌酸激酶(CK)650 U/L,考虑为急性类固醇肌病。停用维库溴铵,改为丙泊酚,甲泼尼龙减量至40 mg/d静脉滴注。2 d后症状缓解,睁眼困难消失,四肢肌力逐渐恢复。5 d后患者CK 119 U/L,肌无力症状完全消失。     

A new pregnane steroid from the stems of Caralluma umbellata

A new pregnane steroid with an unusual formyl group (1) was isolated from the stems of Caralluma umbellata, together with three known steroid compounds. The structure of 1 was elucidated on the basis of spectral evidence including 2D NMR studies.     

Therapeutic potential of cationic steroid antibacterials

Antibiotics were one of the great health successes of the 20th century. Antibiotics, both naturally derived and synthetic, have resulted in huge decreases in both morbidity and mortality from bacterial infections. As a consequence, the ‘antibiotic age’ has changed public expectations about the results of infectious disease. However, this has led to high levels of inappropriate prescribing, where antibiotics may be administered to fulfil patient expectations rather than for clinical benefit. Along with unwise uses in agriculture and elsewhere, this has contributed to recent rises in numbers of antibiotic-resistant bacteria. As a result, many commentators have described this as the end of the antibiotic age and the term ‘superbug’ has entered the common vocabulary for multi-drug-resistant bacteria such as vancomycin-resistant Enterococcus, multi-drug-resistant Staphylococcus aureus and multi-drug-resistant Pseudomonas aeruginosa. In this context, an attractive approach for the development of antibacterial agents is the use of a new class of cationic steroidal compounds mimicking polymyxin activities. The permeabilization properties of these agents of the outer membranes of Gram-negative bacteria are reported in this review, as well as a discussion of literature results.     

Necrotizing fasciitis in a patient with bullous pemphigoid treating with systemic steroid

Abstract

Necrotizing fasciitis (NF) is a rarely seen, life-threatening soft tissue infection characterized by progressive necrosis of skin, subcutaneous tissues and fasciae. Herein, we present a case of NF in a patient with bullous pemphigoid. A 78-year-old female patient was admitted with the left leg pain, swelling, redness and blistering. Streptococcus pyogenes was isolated from the wound culture. In the skin biopsy and the left leg magnetic resonance imaging, signs of NF were observed. The diagnosis of NF was considered. Then, urgent surgical debridement was performed by the department of plastic surgery. While under treatment with intravenous antibiotics therapy, unfortunately the patient died of acute renal insufficiency.     

The effect of tea consumption on the steroid profile

Green tea (GT), along with its flavonol epigallocatechin?3‐gallate (EGCG), has shown to inhibit the UGT2B17 isoenzyme, which is highly involved in the glucuronidation of testosterone (T) and its metabolites. Since the steroid profile (SP) is composed of urinary concentrations of T and related metabolites excreted in both the free and the glucuronide fractions, GT consumption could alter the SP, leading to misunderstanding in doping controls. The aim of the present work was to study the effect of GT consumption on the SP. This study was performed with 29 male volunteers, which could be classified in 2 arms depending on their T/E values (0.12 ± 0.02, n = 12; 1.64 ± 0.90, n = 17). The clinical protocol was designed to evaluate the effect of GT administration on the SP biomarkers. Participants were asked to consume GT with a high content of EGCG for 7 days (5 GT beverages along the whole day for days 1–6 and 9 GT beverages on day 7, corresponding to 520 and 936 mg/day of EGCG, respectively). Urine samples were collected before and during GT consumption at different time periods. The SP was measured using gas chromatography–mass spectrometry. The excretion rates of the SP metabolites did not change after GT consumption. Moreover, the individual evaluation of the subject's steroidal biological passport resulted in normal sequences. The results obtained show that GT consumption does not distort the establishment of normal ranges of SP parameters. Therefore, GT consumption does not need to be considered a confounding factor in the SP evaluation.