Background In women of reproductive age, polycystic ovarian syndrome (PCOS) is a frequent endocrinopathy and a heterogeneous, enigmatic condition that is still not fully understood. It begins to manifest between the ages of 18 and 45, and it may take years for its clinical manifestation to manifest. Vitamin D affects progesterone production in human granulosa cells, anti-müllerian hormone (AMH) signalling, follicle-stimulating hormone sensitivity, and ovarian follicular growth and luteinization as part of its physiological role in reproduction. Increased LH/FSH ratios can occur in ovulatory women with the polycystic morphology, but they may not be seen in a single blood sample. With this context in mind, the current study was conducted to determine and evaluate the biochemical tests in individuals with clinical symptoms of polycystic ovarian syndrome, such as Vitamin D, LH/FSH ratio, and serum insulin levels. Aim and Objectives 1. The objective of this study is to estimate the levels of Vitamin D, LH/FSH ratio and serum insulin in women with PCOS. 2. To find the correlation of LH/FSH, with Vitamin D in PCOS Patients and compare with controls. Materials and Methods The Biochemistry Department and the Department of Gynaecology at SGT Hospital, Budhera, Gurugram's Faculty of Medicine and Health Sciences, conducted this hospital-based cross sectional study. We will use 100 healthy volunteers who are age-matched from the general population as the control group. A signed and informed permission will be obtained from each individual in both groups once they have been fully informed of the study's objectives. Before beginning the sample collection, institutional ethical committee approval will be sought. On the fully automated Biochemistry analyzer MAGLUMI 1000, serum insulin, LH/FSH, and vitamin D levels will be determined using a chemiluminescence-based immunoassay (CLIA). Results Patients with PCOS had a significantly higher level of a parameter, such as serum insulin, with a p value of < 0.0001. The mean and standard deviation of the LH/FSH ratio were also substantially greater in the cases (1.60±0.59) than in the control groups (2.73±1.06). Statistically speaking, the p value was very significant at< 0.001. It indicates that both groups' LH/FSH levels significantly changed. p value <0.0001 indicates that the level of vitamin D is lower in cases when compared to controls. LH/FSH ratio and Vitamin D have negative correlation.