Bitter gourd is renowned for its various bioactivities, including antioxidant, antiglycation, and antiplatelet aggregation. However, the bioactivity and bioavailability of plant extracts could be low. Biotransformation through Lactobacillus plantarum fermentation is capable of enhancing these beneficial properties. Therefore, the present work aimed to investigate the physical and pH profiles, total phytochemical contents, bioactive contents, and bioactivities of the juices of unfermented bitter gourd (UBG) and fermented bitter gourd at 48 (FBG-48) and 96 (FBG-96) h. Fermentation gradually altered the physical and pH profile of bitter gourd, while non-significant decrease was seen in the total flavonoid content (p > 0.05). The highest total phenolic content was observed in FBG-48, followed by UBG and FBG-96. UBG exhibited the highest total triterpenoid content, followed by FBG-48 and FBG-96. Interestingly, fermentation increased the antioxidant, antiglycation, and anti-aggregation activities of bitter gourd. FBG-48 demonstrated the highest antioxidant and antiglycation activities, with 10.77 and 8.68% higher activity, respectively, compared to UBG and FBG-96. Meanwhile, FBG-96 exhibited the highest antiglycation activity, with 60% increase. These could have been attributed to the biotransformation of tannic acid into syringic acid, and momordicoside Q into kuguacin P, along with the potential release of p-coumaric acid and caffeic acid from the cell walls. These findings demonstrated bitter gourd’s promising role in type-II diabetes complication treatment, particularly through its protein aggregation inhibition activity, and that fermentation could increase its bioactivities.