The spider mite, Tetranychus pueraricola (Ehara & Gotoh; Acari: Tetranychidae), is a serious pest in agriculture and horticulture. Application of chemical pesticides is the main mode of this pest control. Due to pesticide residues and resistance-induced resurgence of pests, there is a need to discover alternatives for spider mite management. GC16 comprises a mixture of calcium chloride (CaCl2, 45%) and lecithin (55%), which was recently found to have acaricidal properties. We evaluated the sublethal effects of GC16 on T. pueraricola using life table and enzyme [catalase (CAT), peroxidase (POD), superoxide dismutase (SOD), carboxylesterase (CarE), glutathione S-transferases (GST), and Ca2+-ATPase (Ca2+-ATP)] activity assays. The results showed that fecundity of T. pueraricola increased at LC30 but decreased at LC50 of GC16. The intrinsic rate of increase (r) of T. pueraricola decreased under the LC30 and LC50 of GC16. GC16 concentration and exposure time significantly influenced the activities of CAT, POD, CarE, GST, and Ca2+-ATP in adult mites. Twelve hours later after the treatment, GST and Ca2+-ATP activities were significantly inhibited by LC30 but enhanced by LC50. Moreover, the demographic parameter r and enzyme activities were negatively correlated. In sum, sublethal amounts of GC16 had an adverse effect on mites, and there was a trade-off between developmental performance and physiological enzyme activity of mites under GC16 stress, and GC16 showed an acaricidal potential for T. pueraricola. This work provides guidance for the application of GC16 to control T. pueraricola.