Gastric content is a complex biofluid within the human stomach which has an important role in digestive processes. It is believed that gastric content may be a contributory factor in the development of upper gastro-intestinal diseases. In this work, selected ion flow tube mass spectrometry (SIFT-MS) has been applied to the quantification of volatile organic compounds (VOCs) in the headspace vapor of gastric content samples, which were retrieved from three groups of patients, including those with gastro-esophageal cancer, noncancer diseases of the upper gastro-intestinal tract, and a healthy cohort. Twelve VOCs have been investigated in this study; the following 7 VOCs, acetone, formaldehyde, acetaldehyde, hexanoic acid, hydrogen sulphide, hydrogen cyanide, and methyl phenol, were found to be significantly different between cancer and healthy groups by the Mann-Whitney U test. Receiver operating characteristics (ROC) analysis was applied for the combined VOCs of acetaldehyde, formaldehyde, hydrogen sulphide, and methyl phenol to discriminate cancer patients from healthy controls. The area under the curve (AUC) was 0.9. This result raises the prospect that a VOC profile rather than a single biomarker may be preferable in the molecular-orientated diagnosis of gastro-oseophageal cancer, and this warrants further investigation to assess its potential application as a new diagnostic test.