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The primary mechanism of drug resistance is the reduction of intracellular accumulation of azole in C. albicans worldwide ( Berkow and Lockhart, 2017 Campitelli et al., 2017 Pristov and Ghannoum, 2019). However, high administration frequency and long duration treatment of FLC contribute to the rising number of drug resistant C. Currently, fluconazole (FLC), a highly selective inhibitor of fungal cytochrome P-450 sterol C-14 alpha-demethylation, is the most widely administered antifungal for treating invasive, life-threatening fungal infections ( Robbins et al., 2017 Revie et al., 2018). Systemic candidiasis is a serious healthcare-associated infection in Europe and US, and associated with high mortality rates (40%) among hospitalized patients, particularly in individuals with hematological malignancies, undergoing major surgery, cytotoxic chemotherapy, and organ transplantation ( Magill et al., 2014 Bongomin et al., 2017 Pappas et al., 2018 Hou et al., 2022). albicans) is one of the most common commensal fungal species located in the gastrointestinal and reproductive tracts of healthy individuals, causing both mucosal and systemic infections in immunocompromised individuals ( Brown et al., 2012 de Oliveira Santos et al., 2018 Quindós et al., 2018). Taken together, we established a highly accurate, efficient, and reproducible method for quantifying the intracellular content of FLC in C. Furthermore, this method was successfully applied and validated in 36 clinical isolated strains. The overall recovery of FLC from samples was higher than 77.61%. The precision and accuracy of the samples were relative standard deviations (RSD) < 1.0% for intra-day and RSD < 0.51% for inter-day. The linear for this method were in the range from 5.0 to 1000.0 ng/mL. The multiple reaction monitoring transitions (precursor to product) were monitored for FLC m/z 307.1 → 238.2 and for the internal standard KCZ m/z 531.2 → 489.1. FLC and ketoconazole (KCZ, internal standard) were monitored in positive mode using electrospray ionization source.

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The extracts were separated on a Zorbax SB-C18 column using a mobile phase of acetonitrile (solvent A) and deionized water plus 0.1% formic acid. The entire extraction approach was simple, precise and reliable. albicans cells with Precellys homogenizers and FLC was extracted with methylene chloride. The cell lysates were prepared by lysing C. In this study, a simple and sensitive liquid chromatography with tandem mass spectrometry (LC-MS/MS) method was developed to quantify the intracellular fluconazole (FLC) content in the opportunistic human fungal pathogen Candida albicans. The combination therapy based on existing antifungal agents is well-established to overcome drug resistance and restore antifungal efficacy against drug-resistant strains. Systemic candidiasis is the fourth leading cause of healthcare-associated infections worldwide. 4Department of Pharmacy, Minhang Hospital, Fudan University, Shanghai, China.3Department of Pharmacy, Mudanjiang First People’s Hospital, Mudanjiang, China.

2School of Pharmacy, Naval Medical University, Shanghai, China.1Institute of Vascular Disease, Shanghai TCM-Integrated Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, China.Xiaofei Wang 1,2,3†, Xiaojuan Wang 2,4†, Tongkai Cai 1,2, Yulin Qin 2,4, Ling Li 1, Yuanying Jiang 2*, Bing Han 4* and Yongbing Cao 1*

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