Metabolic behaviors of different grapevine cultivars, grown in the same greenhouse, were characterized through a 1H NMR-based metabolomic approach. Pattern recognition, including the principal component analysis, revealed clear dependence of the grape metabolome on the grape cultivar. Interestingly, high accumulations of proline in the purebred grape cultivars of Cabernet Sauvignon, Merlot, and Chardonnay were found, whereas the proline levels were depleted in the crossbred grape cultivars of Steuben, Campbell Early (C. E.), and Seibel. Intrinsic levels of alanine, glutamine, and trans-feruloyl derivative were highest in C. E. cultivar, which grows easily in a wild vineyard, particularly in Korea, suggesting that their levels play important roles in the improvement of resistance or adaptation of the plant to environmental stress, such as freezing stress during the winter season in Korea. The present study highlights that metabolomics is a powerful approach for better understanding the differences of intrinsic metabolic variables of grape berries among various grape cultivars and their associations with the plant physiological mechanisms.