Introduction Apocyni Veneti Folium (AVF) is a commonly used traditional Chinese medicinal herb for the treatment of hypertension. Chemical markers are crucial for the quality control of herbal medicines; however, the therapeutic components of AVF remain to be well elucidated. Objectives This study was intended to integrate serum pharmacochemistry and network pharmacology to identify chemical markers of AVF and establish an efficacy‐related quality control method of AVF. Material and methods Ultra‐performance liquid chromatography quadrupole time‐of‐flight tandem mass spectrometry (UPLC‐QTOF‐MS/MS) was applied to identify the absorbed AVF constituents in rat serum. Network pharmacology was further used to identify anti‐hypertension‐related chemical markers. Subsequently, a quantitative method was established using UPLC with diode array detection (DAD) and applied for quality evaluation of commercial AVF samples. Results Thirteen prototype constituents were unequivocally or tentatively characterized in serum samples, among which quercetin, kaempferol, hyperoside, isoquercitrin, chlorogenic acid, cryptochlorogenic acid, and neochlorogenic acid were identified as dominant chemicals related to anti‐hypertensive efficacy. The quantitative data showed that the total contents of seven marker components even showed 2‐fold variation among 14 batches of commercial AVF samples with RSD values ranging from 12.15% to 75.61%. Hierarchical cluster analysis and heatmap analysis showed that 14 batches of commercial AVF samples could be divided into three main groups. Conclusion The chemical markers obtained from this study could be applicable for efficacy‐related quality control of AVF. The efficacy‐related chemical markers of Apocyni Veneti Folium (AVF) were discerned by integrating ultra‐performance liquid chromatography quadrupole time‐of‐flight mass spectrometry (UPLC‐QTOF‐MS)‐based serum pharmacochemistry and network pharmacology. The results showed that four flavonoids (quercetin, kaempferol, hyperoside, and isoquercitrin) and three polyphenols (chlorogenic acid, cryptochlorogenic acid, and neochlorogenic acid) were dominant chemicals related to anti‐hypertension efficacy. Moreover, a quantitative method of these seven constituents was established using UPLC‐diode array detection (DAD) to evaluate the quality of commercial AVF samples.