Effects of Plant Growth Regulators on Micropropagation and Silymarin Accumulation in Milk Thistle (Silybum marianum L.) In Vitro

Abstract

Milk thistle (Silybum marianum L.) is a valuable medicinal plant known for its hepatoprotective compound, silymarin. However, the silymarin content varies with genotype, cultivation methods, climatic conditions, and harvest time. Micropropagation provides a solution by producing genetically uniform plantlets and offering an in vitro system for studying secondary metabolite biosynthesis. Therefore, we established a micropropagation protocol and assessed the effects of plant growth regulators (PGRs) on seed germination, shoot organogenesis, and silymarin accumulation. Surface-sterilized seeds achieved the highest germination rate (53.33%) on MS medium supplemented with 2 mg/L gibberellin (GA3). For shoot induction, 0.5 mg/L 6-benzylaminopurine (BA) yielded the highest number of shoots, averaging 5.69 shoots per explant after 4 weeks. Meanwhile, 0.5 mg/L meta-Topolin (mT) was most effective for shoot multiplication, resulting in a 4.97–fold increase in shoot number after three weeks. Furthermore, culture on medium supplemented with 0.5 mg/L mT significantly enhanced the accumulation of silymarin, photosynthetic pigments, total flavonoids, and total phenolics, as compared with media containing BA or thidiazuron (TDZ). These findings provide an efficient micropropagation protocol and demonstrate the significant influence of PGRs on both plant growth and silymarin production in S. marianum.