Integrated MALDI-TOF MS, Microbiological, Physicochemical and Sensory Assessment of Spoilage in Vacuum-Packaged Chicken Breast During Refrigerated Storage

Spoilage in vacuum-packaged chicken breast is driven by coupled microbial succession and physicochemical changes that cannot be adequately described by a single indicator. In this study, MALDI-TOF MS-based species-level identification of culturable isolates was integrated with microbiological counts (total viable count, lactic acid bacteria, yeasts and molds, and Enterobacteriaceae), physicochemical parameters (pH, water activity, CIE $L^{*}{a}^{*}{b}^{*}$, and total volatile basic nitrogen (TVB-N)), and sensory evaluation of odor, appearance/color, surface texture/slime and overall acceptability (trained panel, $n=8$) during 15 days of storage at 4 °C. Associations among variables were assessed using Spearman correlation analysis. MALDI-TOF MS identified 625 isolates belonging to 67 species across 19 families. The microbial community shifted from an initially diverse flora toward late-stage dominance by Latilactobacillus sakei, L. curvatus, Hafnia alvei, Serratia spp., Carnobacterium maltaromaticum and Brochothrix thermosphacta, while Candida zeylanoides persisted throughout storage. TVC exceeded 7 log CFU/g, and TVB-N increased from 10.65 to 23.20 mg N/100 g ($p<0.05$). TVB-N showed strong positive correlations with all microbial groups ($r_s\ge 0.90$, $p<0.01$) and with seven microbial families at the family level. Hafniaceae dominance coincided with a transient mid-storage decrease in pH, consistent with the deaminative activity of H. alvei. $b_{in}^{*}$ showed significant associations with four microbial families and with both microbial counts and TVB-N, supporting its value as a practical spoilage indicator. Sensory evaluation identified Day 13 as the rejection point, corresponding to TVC of 6.79 log CFU/g and TVB-N of 20.80 mg N/100 g, with simultaneous deterioration of odor and appearance, in contrast to the sequential pattern typically reported under aerobic conditions. To our knowledge, this is the first study to integrate time-resolved MALDI-TOF MS-based family-level profiling with physicochemical and sensory monitoring in vacuum-packaged chicken breast stored at 4 °C, offering a condition-specific framework for shelf-life assessment.