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Since inflammatory bowel disease is a progressive disease, optimal treatment early in the disease course is paramount. Still, the use of conventional drugs is accompanied by side effects. The herbal therapies for gastrointestinal disorders are traditionally widespread in Asia, while in Western countries have been adopted in recent years. Andrographis paniculata is a herbaceous plant used in anti-inflammatory treatment. To improve its clinical application, an analytical method with the potential to define a relationship between dose and plasma concentration of active ingredients is prerequisite. Considering the increasing demand for eco-friendly bioanalytical methods, our research aimed to implement new sample preparation technique microextraction by packed sorbent for extraction of marker compounds of A. paniculata from plasma. This green procedure using polymeric sorbent based on the poly (styrene-divinylbenzene) allowed miniaturization of sample volume (100 µL), decrease in consumption of solvents, and high recovery (92–98%). A highly selective and sensitive reversed-phase ultrahigh-performance liquid chromatography method has been developed and validated. The newest greenness assessment software AGREE calculator was used for evaluation of the method's environmental performance. It suggests that our method conforms to the eco-friendly conditions (score 0.8). The sample preparation was identified as weak point of the bioanalytical methods which highlights the importance of microextraction. [ABSTRACT FROM AUTHOR]