A complete analytical protocol for the determination of 25 doping-related peptidic drugs and 3 metabolites in urine was developed by means of accurate-mass quadrupole time-of-flight (Q-TOF) LC–MS analysis following solid-phase extraction (SPE) on microplates and conventional SPE pre-treatment for initial testing and confirmation, respectively. These substances included growth hormone releasing factors, gonadotropin releasing factors and anti-diuretic hormones, with molecular weights ranging from 540 to 1320 Da. Optimal experimental conditions were stablished after investigation of different parameters concerning sample preparation and instrumental analysis. Weak cation exchange SPE followed by C18 HPLC chromatography and accurate mass detection provided the required sensitivity and selectivity for all the target peptides under study. 2 mg SPE on 96-well microplates can be used in combination with full scan MS detection for the initial testing, thus providing a fast, cost-effective and high-throughput protocol for the processing of a large batch of samples simultaneously. On the other hand, extraction on 30 mg SPE cartridges and subsequent target MS/MS determination was the protocol of choice for confirmatory purposes. The methodology was validated in terms of selectivity, recovery, matrix effect, precision, sensitivity (limit of detection, LOD), cross contamination, carryover, robustness and stability. Recoveries ranged from 6 to 70% (microplates) and 17–95% (cartridges), with LODs from 0.1 to 1 ng/mL. The suitability of the method was assessed by analyzing different spiked or excreted urines containing some of the target substances.

通过固相萃取（SPE）后的精确质量四极杆飞行时间（Q-TOF）LC-MS分析，开发了测定尿液中25种与掺杂有关的肽类药物和3种代谢物的完整分析方案在微孔板和常规SPE预处理上分别进行初始测试和确认。这些物质包括生长激素释放因子，促性腺激素释放因子和抗利尿激素，分子量范围为540至1320 Da。在研究了有关样品制备和仪器分析的不同参数后，确定了最佳实验条件。弱阳离子交换SPE，然后进行C18 HPLC色谱和准确的质量检测，为所有正在研究的目标肽提供了所需的灵敏度和选择性。可以将2 mg SPE在96孔微孔板上与全扫描MS检测结合使用以进行初始测试，从而提供了一种快速，经济高效且高通量的方案，可同时处理大量样品。另一方面，出于确认目的，选择方案是在30 mg SPE柱上提取并随后进行目标MS / MS测定。该方法论在选择性，回收率，基质效应，精密度，灵敏度（检测限，LOD），交叉污染，残留，稳健性和稳定性方面得到了验证。回收率范围为6％至70％（微孔板）和17-95％（墨盒），LOD为0.1至1 ng / mL。该方法的适用性通过分析含有某些目标物质的不同加标或排泄的尿液来评估。