Baicalin is a flavonoid that has an influence on molecular processes. It possesses anticancer, anti-inflammatory, antiviral, antioxidative, and antithrombotic properties. It was found that baica-lein treatment attenuated the damage of the mammary gland induced by LPS, suppressed the activity of myeloperoxidase, TNFα, and IL-1β in mice with mastitis. The aim of the study was a pilot analysis of baicalin tolerability after intramammary (IMM) administration and its impact on somatic cell count (SCC) after multiple IMM treatment on dairy cows with clinical mastitis. Moreover, the determination of baicalin in milk was performed by the sensitive ultra-high per-formance liquid chromatography with tandem mass spectrometry. The pharmacokinetic analyses were performed using Phoenix® WinNonlin® 6.4 and ThothPro v 4.1 software. Twelve dairy cows with clinical mastitis were selected for this study. The pharmacodynamic endpoint was SCC level and the clinical investigation was also carried out. Baseline SCC analysis was performed every 24 h among all cows three days before the first dose (B1–B3). After the baseline monitoring, 8 days of treatment (T1–T8) was performed and 8 days within recovery period SCC level was observed (R1–R8). Starting from T1 to T8, a decrease of SCC in relation to baseline was characterized by a declining trend. The presented results confirm the effect of baicalin on the reduction of SCC in mastitis in dairy cows after this therapy. The current study has shown that baicalin accumulation was not confirmed.