DNA modifications are epigenetics markers that modulate gene expression. The enzymes that apply and erase these markers are emerging targets of biopharmaceutical intervention, as changes in DNA modification are metabolic switches that respond to stimuli and can reflect or cause disease. Accurate detection of epigenetic changes is increasing important to research, therapeutic development and clinical care. We present a direct injection mass spectrometry (DI-MS) platform that accurately, precisely, and quickly quantitates global levels of DNA cytidine methylation (5mC) and hydroxymethylation (5hmC) in less than one minute per sample. Our platform combines an Advion TriVersa NanoMate coupled online to a Thermo Scientific Orbitrap Fusion Lumos. Following digestion to nucleosides, the DNA samples are analyzed at the rate of < 1 min per injection with comparable detection limits of 0.63 ng/µL and 0.31 ng/µL, respectively. In contrast, the detection limits for 5mC and 5hmC in state-of-art nano liquid chromatography (LC) coupled to online mass spectrometry (nLC-MS) are notably different (0.04 ng/µL and 2.5 ng/µL, respectively). The high sensitivity of DI-MS is achieved by optimizing sample buffer composition, the source fragmentation energy, and the radio frequency of the instrument ion funnel. Accuracy of the analysis was optimized using in-source fragmentation to obtain a population of up to 93% of just the nucleoside base. By bypassing chromatographic separation, we avoid potential issues of carryover and batch effects due to potential column contamination across samples. Moreover, we minimize the bias of differential chromatographic retention, which impacts the limit of detection of molecules that do not efficiently bind to our columns during loading, e.g. hmC. The speed of DI-MS opens for the analysis of >1,000 samples per day, achieving a scale comparable to high-throughput sequencing. We are also applying DI-MS to the identification and quantification of other biomolecules including amino acids, peptides and proteins and polynucleotides.