Background Traditional Chinese Medicine (TCM) Cuscutae semen (SC, Tusizi from Cuscuta chinensis Lam.) and Lycii fructus (FL, Gouqi from Lycium barbarum L.), are also used as a herb pair (SC-FL) to treat various ailments, including spermatogenic dysfunction (SD), a disease responsible for low fertility in males. Objective Herein, we will further determine the bioactive components, component targets and partial molecular mechanisms of the herb pair for the treatment of SD. Methods We employed the traditional Chinese medicine systems pharmacology database in combination with Ultra high performance liquid chromatography (UHPLC) analysis to analyze the active ingredients of the SC-FL herbal pair and explore its possible targets and underlying mechanism in treatment of spermatogenic dysfunction (SD). Moreover, we used a Sprague-Dawley rat model, generated by using glucosides of Tripterygium wilfordii (GTW), to evaluate the effect of SC-FL and the underlying mechanism on SD and reliability of certain key targets and pathways obtained from the pharmacology analysis. Results We identified 56 active ingredients in SC-LF affecting 41 overlapping gene signals that influenced SD treatment outcomes. 262 Gene Ontology (GO) terms and 170 pathways were yielded under analyses of Gene Ontology, Kyoto Encyclopedia of Gene and Genome pathway from which we predicted that cell proliferation and apoptosis were the primary biological processes (BP) involved in the treatment of SD by SC-LF. The results showed that SC-FL treatment significantly improved the testicular organ coefficient along with sperm count and motility, while reduced testicular damage and testicular tissue cell apoptosis in SD model. Mechanistically, SC-FL significantly upregulated the expression levels of anti-apoptotic proteins AR and BCL2 and downregulated those of pro-apoptotic proteins BAD, BAX, cleaved caspase 3, and CASP3. Conclusion Collectively, these results indicated that SC-FL elicited a protective effect by potentially regulating apoptosis, thus suggesting that it will represent an effective reagent for male infertility.