We made slight modifications and finalized the gradient as presented in Table . The formation of water-rich layer on the surface of stationary phase is quite important for interaction with the analytes to ensure consistent retention of compound. Hence, we specifically extended the equilibration time as it is a critical step in HILIC for a stable chromatography. illustrates the specific site prone to fragmentation of acyl-CoA. The neutral loss of 507 was chosen as the product ion (Q3) for the sMRM mode as it was the most intense and common fragment among acyl-CoAs, as observed in our study and supported by other literatures .