The determination of trace residues and contaminants in food has been of growing concern over the past few years. Residual antibacterials in food constitute a risk to human health, especially because they can contribute to the transmission of antibiotic-resistant pathogenic bacteria through the food chain. Therefore, to ensure food safety EU and USA regulatory agencies have established lists of forbidden or banned substances and tolerance levels for authorized veterinary drugs (e.g. antibacterials). In addition, the EU Commission Decision 2002/657/EC has set requirements about the performance of analytical methods for the determination of veterinary drug residues in food and feedstuffs. During the past years, the use of powerful mass spectrometric detectors in combination with innovative chromatographic technologies has solved many problems related to sensitivity and selectivity of this type of analysis. However sample preparation still remains as the bottleneck step, mainly in terms of analysis time and sources of error. This review covering research published between 2004 and 2008 intends to provide an update overview of the past five years, on recent trends in sample preparation for the determination of antibacterial residues in foods, making special emphasis in on-line, high-throughput, multi-class methods and including several applications in detail.