Monitoring of biological activity for target enzyme is important for its production in recombinant expression systems. Previously, we demonstrated that green fluorescent protein (GFP) as a fusion partner is successfully tooled for facile, in vivo, and non-invasive quantification of target enzyme levels based on a linear relationship between GFP fluorescence and enzyme (chloramphenicol acetyltransferase; CAT) activity. Here, we investigated the effects of culture environmental variations (initial glucose amount, surface aeration, and inducer concentration) on correlation between GFP fluorescence and CAT activity, and established a general linear correlation as a unique criterion for quantitative monitoring of CAT biological activity. This general correlation for GFP fusion strategy can be applied for non-invasive and on-line monitoring of recombinant enzyme production under various culture conditions without further experimental calibrations.