Preparation of fish samples

Fresh water rainbow trout (Oncorhynchus mykiss) with an average weight of 1500 g were purchased from a public market (Shiraz, Iran) and transferred to the laboratory packed in an insulated ice box. TMAO content of the rainbow trout was reported to be 0.31 g kg⁻¹ (wet weight). However, this value can be changed depending on the nutrition and the season (RØrvik et al. 2000). The fishes were aseptically headed and filleted by knife to obtain uniform cubical samples of equal weight (approximately 30 g). All the fish samples were immediately placed in sterilized petri dishes. A number of alginate beads as the pH indicator were fixed on the inner side of the lid of the petri dishes using a liquid stick such that the beads would be facing the headspace standing 3 cm above the fish samples. Closed petri dishes were stored at room temperature for 64 h and at 4 °C for 12 days for spoilage monitoring. At specified intervals during the storage period, the samples were analyzed to detect any chemical or microbial changes. Reference petri dishes contain no fish samples were also maintained over the same experimental period as the ones containing fish samples.