## 3 Results and Discussion
### Extracellular Enzyme Profiles per API ZYM(r) Kit for Biosolids  
The mineralisation of organic matter is mainly mediated by microorganisms, which possess the adequate enzymatic machinery to obtain the energy necessary to carry out their metabolic processes [39; 134]. The enzymes involved in the degradation of organic matter can be divided into hydrolytic enzymes that are responsible for the acquisition of C, N and P to support primary and oxidative metabolism, oxidative enzymes that degrade recalcitrant compounds such as lignin during in the acquisition of nutrients [135; 136]. These groups of enzymes may respond differently to the addition of organic amendments such as biosolids. [PERSON] et al. [62] stated that the addition of biosolids to soils can improve and maintain biological activity in soils over the long term. However, little information is available on the enzymatic activity of biosolids due to the lack of methodologies or analytical methods that provide information quickly and inexpensively. In recent decades, the API ZYM(r) system has been used and validated to evaluate enzymatic profiles of soils, compost, activated sludge and sediments [34; 36; 37; 38; 39; 44; 134; 137; 138]. However, the API ZYM(r) system has not been used directly and over time in biosolids samples as a system for the rapid determination of 19 extracellular hydrolytic enzymatic reactions related to the C, N and P cycles. The objective of using the API ZYM(r) system to evaluate biosolids at different season of the year was to quickly obtain information on the extracellular enzyme profile of biosolids. With the information obtained, it could be used as a background on the possible positive or negative effects on soil functional diversity when biosolids are used as an organic amendment. The extracellular enzyme profile of the biosolids analysed for each season of year is shown in Figure 4. The enzyme activity was classified into 5 families: (1) phosphatases (catalyses the removal of phosphate groups from substrates as nucleic acids), (2) esterases-lipases (hydrolyse ester bonds mainly in fats and oils), (3) aminopeptides (hydrolyse peptide bonds in peptides), (4) proteases (hydrolyse peptide bonds in proteins) and (5) glycosyl hydrolases (catalyses the hydrolysis of glycosidic bonds in carbohydrates). The enzymatic activities presented differences and similarities for each season of the year. In spring, a mainly moderate enzymatic activity was obtained (Figure 4). Within the 19 enzymes determined in biosolids in March and April, only 1 enzyme showed no activity, and it belonged to the family of glycosyl hydrolases. In biosolids produced in May, there were no activities of 3 enzymes, 2 glycosyl hydrolases and 1 protease. Low enzymatic activity in 4, 7 and 5 enzymes were detected in biosolids for March, April, and May, respectively. These enzymes belonged to the families esterases-lipases, aminopeptides, proteases and glycosyl hydrolases (Figure 4). The enzyme activity of the biosolids was moderate for March, April, and May in 10, 6 and 8 enzymes, respectively, and were distributed within the 5 families. In March, April and May biosolids, high enzyme activity was presented for 4, 5 and 3 enzymes, respectively, mainly from the phosphatase family. In summer, the enzyme profile of biosolids showed a predominantly high enzyme activity for all enzyme families. Undetected enzyme activity was obtained for 1 glycosyl hydrolase enzyme in June and July biosolids, while in August there were 2 enzymes, 1 glycosyl hydrolase and the other aminopeptidase. In June, low biosolids biological activity was obtained for 3 enzymes of the glycosyl hydrolases and aminopeptides families. Moderate summer enzyme activity was presented in 9, 6 and 3 enzymes for June, July and August biosolids, respectively (Figure 4). The esterases-lipases and aminopeptides families were the main ones that presented moderate enzymatic activity. The high enzyme activity for June, July and August biosolids was exhibited in 6, 12 and 4 enzymes, respectively.