Fruit vinegar is typically produced through a two-stage deep liquid fermentation involving alcohol fermentation (Saccharomyces cerevisiae) and acetic acid fermentation (Acetobacter pasteurianus). In order to enhance the flavor and texture of sea buckthorn fruit vinegar, Lactobacillus fermentum was introduced into the alcoholic fermentation stage. At the end of fermentation, the total acid and acetic acid of sea buckthorn (Hippophae rhamnoides L.) fruit vinegar were both enhanced compared with sea buckthorn vinegar brewed through the traditional liquid fermentation method, and in terms of the main active ingredients, the total flavonoid content was slightly enhanced. Non-targeted metabolomics (LC-MS) was used to characterize the characteristic metabolite profiles during the fermentation process. A total of 55 differential metabolites, including organic acids, flavonoids, and amino acids, were identified, and the contents of citric acid, malic acid, and manganic acid, which are the sources of the irritating taste of sea buckthorn berry vinegar, were significantly reduced. In addition, the co-fermentation of Lactobacillus fermentum promoted both glycolysis and the TCA cycle and also led to a significant up-regulation of aromatic metabolites, such as ethyl acetate, ethyl lactate, and ethyl caproate. These results will provide new information on the dynamics of the characterized metabolites during the fermentation of sea buckthorn fruit vinegar.