Several studies described variations in lactose percentage (LP) in dairy cows during udder inflammation or negative energy balance (NEB). However, their joint effects on LP were never described together. This was the aim of a longitudinal observational study performed on 5 Quebec dairy farms using automatic milking systems. Quarter milk samples were collected every other week from 5 to 300 DIM. Quarter health status was described by combining inflammation status (SCC- or SCC+: < or ≥100 000 cells/mL) and microbial status (Path- or Path+: presence or not of pathogens on a milk culture). Cows with NEB in early lactation (DIM <70) were identified using milk BHB content: 0.15 mM = BHB-; 0.15–0.19 mM = BHB+; > 0.19 mM = BHB++. A total of 14,583 quarter foremilk samples were collected from 380 lactating cows. The quarter level LP was analyzed using a mixed linear regression model with the following fixed effects: quarter health status, parity, delay between last milking and sampling, quarter milk yield (in kg/d), DIM, and herd. A random quarter intercept with a repeated measures correlation structure and a cow random intercept were also specified. The LP of SCC+ quarter milk was lower (−0.17 ± 0.014%; P < 0.01) for both primiparous and multiparous. LP was also lower in the presence of streptococcus dysgalactiae and Staphyloccocus aureus regardless of the quarter inflammation status (SCC- or SCC+): −0.13 ± 0.029% for primiparous and −0.11 ± 0.027% for multiparous cows (P < 0.01). BHB+ and BHB++ cows in early lactation had lower LP (−0.04 ± 0.012% and −0.12 ± 0.021% respectively) compared with BHB- cows (P < 0.01). A larger decrease of LP was observed in SCC+ quarters from BHB+ and BHB++ cows, respectively −0.19 ± 0.021% and −0.27 ± 0.020% and that, in both parity groups (P < 0.01). Moreover, the cumulative effect of the quarter health status and NEB on milk LP was greater with higher NEB. Our findings suggest the necessity to jointly take in consideration both quarter health status and NEB when using LP as a health indicator.