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Quinolizidine alkaloids (QAs) are toxic secondary metabolites of the <i>Lupinus</i> species, the presence of which limits the expansion of lupin beans consumption, despite their high protein content. Evaluation of the level of alkaloids in edible <i>Lupinus</i> species is crucial from a food safety point of view. However, quantitation of QAs is complicated by the fact that not all important alkaloids used for quantitation are commercially available. In this context, we developed a method for the simultaneous quantitation of eight major lupin alkaloids using quantitative NMR spectroscopy (qNMR). Quantitation and analysis were performed in 15 different seed extracts of 11 <i>Lupinus</i> spp. some of which belonged to the same species, with different geographical origins and time of harvest, as well as in all aerial parts of <i>L. pilosus</i>. The mature seeds of <i>L. pilosus</i> were found to be a uniquely rich source of multiflorine. Additionally, we developed a protocol using adsorption or ionic resins for easy, fast, and efficient debittering of the lupine seeds. The protocol was applied to <i>L</i>. <i>albus</i>, leading to a decrease of the time required for alkaloids removal as well as water consumption and to a method for QA isolation from the debittering wastewater.