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This invention provides a prickly pear and Job's tears yogurt and its preparation method, belonging to the field of yogurt preparation technology. The preparation method of the prickly pear and Job's tears yogurt includes the following steps: soaking Job's tears and mixing them with water, then successively pulping, gelatinizing, and then hydrolyzing with α-amylase at high temperature to obtain a Job's tears enzymatic hydrolysate; inoculating the Job's tears enzymatic hydrolysate with probiotic BLH1 seed liquid for fermentation; after fermentation, adding prickly pear juice to the fermentation liquid for further fermentation to obtain a prickly pear and Job's tears fermentation liquid; mixing milk powder, water, and sucrose evenly, homogenizing, sterilizing, cooling, and then mixing evenly with the prickly pear and Job's tears fermentation liquid, inoculating with lactic acid bacteria seed liquid for fermentation, and after fermentation, ripening to obtain the prickly pear and Job's tears yogurt. The prickly pear and Job's tears yogurt provided by this invention has a good taste, high sensory evaluation, significantly retains functional components such as phenols, flavonoids, and vitamin C, and greatly increases GABA content and antioxidant activity.