One of the most important aspects of cell metabolism in eukaryotic cells is the compartmentalization of reactions. Nicotinamide (pyridine) dinucleotides (NAD+, NADP+, NADH, and NADPH) were discovered over a century ago as obligatory cofactors for fermentation in yeast extracts. Since then, these molecules have been documented as key participants in a wide range of cellular processes, including biosynthesis, signal transduction, redox homeostasis, and ATP production. NADPH can be generated by various pathways by different enzymes. The level and cellular localization of NADPH might regulate its biological roles. The last few years have perceived the engineering of genetic tools for monitoring NAD(H) and NADP(H) compartmentalization and metabolism. Here, we provide an integrated view on the pathways underlying the basic principles of NADPH production and its subcellular compartmentalization..