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People living in non-Mediterranean West Balkan countries have diets with a low <i>n</i>-3 polyunsaturated fatty acid (PUFA) content. Walnuts, a traditional Serbian food, could be an excellent source of <i>n</i>-3 PUFA. The first sub-study evaluated the fatty acid and mineral content of Serbian walnuts, demonstrating that walnuts had the high content of linolenic acid (C18:3, <i>n</i>-3 ALA). The second sub-study assessed the consumption of walnuts (<i>Juglans regia</i> L.) and total <i>n</i>-3-fatty acid intake in apparently healthy Serbian residents, using 24-h dietary recalls (<i>n</i> = 352). An inadequate intake of <i>n</i>-3 fatty acids and a low consumption of walnuts was seen. Additionally, we evaluated the fatty acid profile of healthy Serbian adults (<i>n</i> = 110) and finally, via a randomized intervention 4-weeks study, we assessed the effects of walnut consumption on <i>n</i>-3 fatty acid profile of participants (<i>n</i> = 18). The plasma content of <i>n</i>-3 PUFA was low and the <i>n</i>-6/<i>n</i>-3 ratio was high in our study participants. The <i>n</i>-3 plasma fatty acid profile was improved after 4 weeks of walnut consumption, meaning that ALA, eicosapentaenoic acid, and total <i>n</i>-3 were significantly increased. The results of our study pointed out the potential health benefits of walnuts consumption on amelioration of the <i>n</i>-3 fatty acid profile that should be taken into account in preventive management programs. The higher conversion of ALA to EPA (&gt;10%) in examined study participants, suggests the importance of a moderate walnut consumption.