# Hake Fish Preservation Using Plant-Based Impregnated Polylactic Acid Food Films as Active Packaging
## 3 Results and Discussion
### Preservation of Hake Fillets
#### 3.2.2 Effects of Storage on the Physical and Chemical Properties
However, this result suggests that a PLA coating impregnated with natural extracts may effectively control moisture loss in fish fillets during cold storage. Regarding the pH, [PERSON] et al. [56] identified pH values as a biomarker for fish and meat freshness, noting that values over 7 suggest the onset of food deterioration. Figure 4c shows a separation of the pH increase tendency among the impregnated and non-impregnated films from day 2. On days 5 and 7, the hake fillets packaged in the impregnated films exhibited steady pH values, remaining slightly above 7, indicating a delay in spoilage. During the same period, the pH values of hake fillets packaged in the control films were statistically higher, suggesting a faster rate of spoilage. When fish are caught under stressful conditions, biochemical processes occur that affect the pH. Initially (post-catch), a decrease in pH can be observed due to glycogenesis reactions that release lactic acid, which accumulates in muscle tissue [57]. However, once the fish begins to deteriorate, basic compounds are generated that can increase the pH such as volatile bases, trimethylamine, and ammonia [58] as well as the formation of alkaline compounds due to microbiological activity or endogenous enzymes present in the fish [59]. Therefore, these changes in pH can be indicative of the degree of freshness of the fish and its quality. On the other hand, Figure 5 shows the evolution in the chemical parameters of the packaged hake fillets over the storage period. Figure 4: Evolution of **(a)** drip loss (%), **(b)** water activity (a\({}_{w}\)), and **(c)** pH during preservation at 4 \({}^{\circ}\)C at each sampling point. Data are expressed as the mean \(\pm\) standard deviation (\(n=3\)). Lowercase letters indicate differences between the impregnated and control samples at the same storage time. Capital letters indicate differences along storage days in the same treatment (\(p<0.05\)). The total volatile basic nitrogen (TVB-N) is one of the most widely used measurements of seafood quality and is a common chemical indicator of marine fish spoilage [60,61]. Both the bacterial activity and biochemical modifications resulting from the autolytic activity of fish produce a series of basic nitrogenous compounds, such as ammonia, trimethylamine (TMA-N), dimethylamine, and monoethylamine, known as the total volatile basic nitrogen (TVB-N), which are considered representative of this alteration [62]. The concentration of TVB-N in freshly caught fish usually varies between 5 and 20 mg 100 g\({}^{-1}\)[63]. Based on previous studies [64, 65, 66, 67], [PERSON] proposed that the quality classification of fish and fish products regarding TVB-N values would be \"high quality\" up to 25 mg 100 g\({}^{-1}\), \"good quality\" up to 30 mg 100 g\({}^{-1}\), \"limit of acceptability\" up to 35 mg 100 g\({}^{-1}\), and \"spoilt\" above 35 mg 100 g\({}^{-1}\)[59]. [PERSON] et al. [34] determined that a gelatin/chitosan coating could improve the quality of fresh fish fillets and prevent spoilage when refrigerated at 4 \({}^{\circ}\)C. They showed that the TVB-N for the control group increased dramatically from 3.59 mg 100 g\({}^{-1}\) at day 0 to 93.52 mg 100 g\({}^{-1}\) at day 17. This result suggests that the freshness of the control filled decreased quickly and became spoilt during cold storage. In this study, the TVB-N (Figure 5a) showed the same trend. The samples exposed to the impregnated and control films increased from 10.20 mg 100 g\({}^{-1}\) at day 0 to values near 80 mg 100 g\({}^{-1}\) at day 12. There comes a time of advanced deterioration in which not only volatile bases are produced, but also acids, depending on the types of microorganisms developing in the environment, which is why volatile bases remain or fall and the pH drops (Figure 4c). TMA-N (Figure 5b) is the main component of the TVB-N fraction. This compound is produced by the bacterial reduction of trimethylamine oxide (TMAO) and is considered responsible for the characteristic \"unpleasant smell\" of fish [68].