The invention discloses highland barley flavored beef jerky with barley fragrance and a preparation method of the highland barley flavored beef jerky. The highland barley flavored beef jerky is prepared from the following raw materials in parts by weight: 16-18 parts of highland barley, 240-250 parts of beef, 18-20 parts of barley seedling leaves, 6-8 parts of common turnips, 8-10 parts of semen cassiae, 2-3 parts of villous amomum fruits, 1-2 parts of radix angelicae, 1-2 parts of fennel fruits, 1-2 parts of nardostachyos root and rhizome, 6-8 parts of table salt, 4-6 parts of mango cores, 2-3 parts of Chinese peashrub roots, 2-4 parts of pomegranate seeds, 2-4 parts of pineapple peel, 4-5 parts of yellow ginger powder, 6-8 parts of tender shoot of Koelreuteria paniculata, an appropriate amount of pomegranate seed oil and 20-30 parts of haw wine. The highland barley flavored beef jerky with barley fragrance, prepared by the preparation method disclosed by the invention, is fragrant, crisp and chewy, integrates the fragrance and the flavor of the highland barley and the fragrance and the flavor of barley, can clear the intestines, ease constipation, lose weight, strengthen bodies, and enhance the immunity of organisms, and is unique in flavor; according to the formula of the highland barley flavored beef jerky with barley fragrance, varied traditional Chinese medicine ingredients are also added, so that health-care functions are improved, and the highland barley flavored beef jerky with barley fragrance has the efficacies of invigorating the stomach, promoting digestion, clearing the lung, promoting blood circulation, resisting oxidation, clearing away heart fire, reducing pathogenic fire and the like; the beef wrapped with barley straws is roasted, so that the barley fragrance is generated, the highland barley flavored beef jerky is rich in flavor, and the highland barley flavored beef jerky with barley fragrance is a novel flavored jerky product with the advantages of being environmentally-friendly, health-care, comprehensive in nutrition, leisure, convenient and the like.