Assessment, identification and characterization of germplasm is a significant link among the conservation, utilization and improvement of plant genetic resources. In current investigation 133 genotypes of (Capsicum annuum) were evaluated at morphological level for 9 quantitative and 21 qualitative traits, the comprehensive approach of these studies delivers useful information on variability of traits for consecutive two years 2016-17. The genotypes were also evaluated for macro, micro elements, antioxidants and proximate analysis. But very little variation was found at proximate level. Among minerals potassium, iron and calcium showed wide range of variability. Potassium ranged from 209.0 to 352.3 mg/100g. Iron displayed variation ranged from 56.2 to 139.4 mg/100g and calcium content varied from 20.2-135.2 mg/100g. Additionally a precise, simple and accurate HPLC technique was used for determination of antioxidants (ascorbic acid and beta carotene) showed wide variation. Ascorbic acid ranged from 85.99 to 317.65 mg/100g whereas beta-carotene varied from 129.37 to 1318.07 ug/g. Furthermore, capsinoids, unique feature of Capsicum annuum were also determined by HPLC technique, exhibited immense variations for capsiacin ranged from 5.96 to 3924.27 ug/g, whereas range of dihydrocapsiacin varied from 1.96 to 1428.05 ug/g among diverse genotypes. Cluster analysis exposed grouping of genotypes based on similarities indices regardless of geographical origin. Correlation of yield with agronomic trait presented complex pattern of association. Principal component analysis was performed to observe the percentage contribution of each parameter to total biochemical variability. The first principal component (PC1) elucidated 64.53 % of variation. (PC2) exhibited 32.5 % of variability among genotypes for all biochemical traits. Germplasm displayed substantial variation, on the basis of agronomic performance, 20 genotypes were identified for future breeding either through direct exploitation or involved in the improvement of Capsicum through breeding programs. For better understanding molecular diversity was tested through 29 SSR markers to detect polymorphism in 133 genotypes. An average of 2.0 allele per marker was perceived. In the conclusion 28 characters studied in a set of 133 genotypes displayed a wide range of variability for yield, single fruit weight, plant height, potassium, iron, calcium, ascorbic acid, beta-carotene and capsinoids contents.