Micro-methods to determine the composition of selected secondary metabolites in Colombian forages

Plants used as forage in livestock production are a source of secondary metabolites (SM) that are involved in important interaction in life, health, nutrition and animal performance. This study aimed to adapt and validate six spectrophotometric micro-methods for quantifying SM such as total phenols (TP), total tannins (TT), condensed tannins (CT), total saponins (TS), total alkaloids (TA), and total sterols (TE), in more than 700 Colombian tropical forage samples. The sample and reagent quantities used in the assays were reduced, preparation conditions were modified in some steps, and the colorimetric reactions were performed in 96-well microplates. The micro methods were validated for linearity, limits of detection and quantification, accuracy, precision, and percent recovery in materials such as grasses, legumes, and tree forages from six Colombian ecoregions. The coefficients of variation (CV), repeatability and reproducibility for all methods were less than 0.4 and 5.6%, respectively. The recoveries ranged from 84.8 to 88.4% for TP, 65.7 to 92.3 for TT, 59.8 to 80.2 for CT, 49.9 to 69.5 for TA, and 61.9 to 78.8% for TS. Micro assays promote minimal waste generation in the environment and allow the processing of a larger number of samples, thereby reducing reagent consumption. This is particularly useful in the analysis of forage materials when making decisions about the effects and functionality of these components in animal diets and their performance.