The need to improve the physicochemical properties of sea buckthorn juice and the bioavailability of carotenoids is a major challenge for the field. The effects of different natural emulsifiers, such as medium-chain triglycerides (MCTs), tea saponins (TSs) and rhamnolipids (Rha), on the physical and chemical indexes of sea buckthorn juice were studied. The particle size of sea buckthorn juice and the carotenoids content were used as indicators for evaluation. The effects of different addition levels of MCT, Rha and TS on the bioavailability of carotenoids in sea buckthorn juice were investigated by simulating human in vitro digestion tests. The results showed that those emulsifiers, MCT, Rha and TS, can significantly reduce the particle size and particle size distribution of sea buckthorn juice, improve the color, increase the soluble solids content, turbidity and physical stability and protect the carotenoids from degradation. When the addition amount of Rha was 1.5%, the total carotenoids content (TCC) of sea buckthorn juice increased by 45.20%; when the addition amount of TS was 1.5%, the total carotenoids content (TCC) of sea buckthorn juice increased by 37.95%. Furthermore, the bioaccessibility of carotenoids was increased from 36.90 ± 2.57% to 54.23 ± 4.17% and 61.51 ± 4.65% through in vitro digestion by Rha and TS addition, respectively. However, the total carotenoids content (TCC) of sea buckthorn juice and bioaccessibility were not significantly different with the addition of MCT. In conclusion, the findings of this study demonstrate the potential of natural emulsifiers, such as MCT, Rha and TS, to significantly enhance the physicochemical properties and bioavailability of carotenoids in sea buckthorn juice, offering promising opportunities for the development of functional beverages with improved nutritional benefits.