Lutein is a xanthophyll family of carotenoids, found in flowers, vegetables, and fruits either in esterified or non-esterified fatty acid form. It is mainly administered in pharmacological products, dietary additives, the food industry, and animal feeding industries. This study was conducted on the ?Fandoghi‘ variety from the Markazi province for pistachio hull lutein extraction and quantification. This study aimed to assess the lutein in pistachio hull and optimize its extraction protocol by new extraction methods with emphasis on microwave-assisted method (MAE). The powder from dried pistachio hulls obtained from fresh raw un-hulled pistachios was applied for further analysis. An experimental design based on the central composite design was applied for the extraction using the MAE method and extraction optimization. The lutein contents were quantitatively analyzed using a validated LC-MS/MS method. According to the free form of lutein, Ethyl acetate was applied as an extraction solvent with the MAE method followed by the setting up of the extraction time, temperature, and solvent/sample ratio as variables. Under optimal experimental conditions corresponding to 5 min extraction time at 40°C, and 30 mg ml-1 of the solvent/sample ratio, the amount of lutein obtained from dried pistachio hull was 3.86 mg 100 g-1. The MAE method is a green, time-saving, and cost-effective method for lutein extraction from pistachio hull that can be suggested for lutein extraction from other plant materials and it can be applied in industrial scale.