Glyphosate and intermediates/impurities of production are separated on an Obelisc N HILIC/ion-exchange column by a combination of HILIC and ion-exchange mechanism. Method can be used in analysis of glyphosate, iminodiacetic acid, (N-phosphonomethyl)-imminodiacetic acid, diethanolamine and related impurities in reaction mixtures, waste and ground waters. Detection techniques are LC/MS, ELSD, CAD and UV.