This review describes the analytical methods that have been developed over the years to tackle the high polarity and non-chromophoric nature of amino acids (AAs). First, the historical methods are briefly presented, with a strong focus on the use of derivatization reagents to make AAs detectable with spectroscopic techniques (ultraviolet and fluorescence) and/or sufficiently retained in reversed phase liquid chromatography. Then, an overview of the current analytical strategies for achiral separation of AAs is provided, in which mass spectrometry (MS) becomes the most widely used detection mode in combination with innovative liquid chromatography or capillary electrophoresis conditions to detect AAs at very low concentration in complex matrixes. Finally, some future trends of AA analysis are provided in the last section of the review, including the use of supercritical fluid chromatography (SFC), multidimensional liquid chromatography and electrophoretic separations, hyphenation of ion exchange chromatography to mass spectrometry, and use of ion mobility spectrometry mass spectrometry (IM-MS). Various application examples will also be presented throughout the review to highlight the benefits and limitations of these different analytical approaches for AAs determination.