Introduction
Biuret is a common impurity in urea fertilizers. It is a byproduct of the urea granulation process and is formed by the thermal condensation of two urea molecules. When urea fertilizers are added to arable lands, biuret, a contaminant in these fertilizers, is also applied. Biuret in the soil is decomposed by microorganisms and eventually produces ammonia and carbon dioxide, relatively slow but the same as urea’s fate (Aukema et al., 2020; Cameron et al., 2011; Robinson et al., 2018). Biuret is not known to be toxic to animals; however, excess biuret can stunt plant growth and cause chlorosis of leaves (Mikkelsen, 1990). Consequently, the permissible biuret concentration in many countries is 1.2% for urea fertilizers.
Previous studies on biuret toxicity have shown that it inhibits protein synthesis in plants (Ogata T. & Yamamoto M., 1959; Webster et al., 1957). A recent experiment also found that biuret toxicity can alter the expression levels of many genes involved in environmental stress response (Ochiai et al., 2020). However, the mechanisms underlying biuret toxicity are still not well understood. Clarification of this mechanism may help prevent potential plant injury.
We previously found that rice plants that were overexpressingbiuret hydrolase from a soil bacterium had an enhanced biuret tolerance (Ochiai et al., 2020). The experiment used15N-labeled biuret to show that biuret hydrolase overexpressing plants take up more biuret than wild-type plants. However, the form of15N in plants after uptake is not known. In this study, we examined the biuret accumulation, in wild-type andbiuret hydrolase overexpressing rice plants using HPLC-UV, to understand how the biuret concentration in plants causes injury.
In addition, we hypothesized that biuret inhibits the metabolism of compounds with a similar structure, specifically ureido compounds. As an ureido, we focused on allantoin because of its multiple roles and importance to plants. Allantoin, a compound composed of a hydantoin ring and ureido group, is an intermediate in the purine degradation pathway. It contains four nitrogen (N) atoms per molecule and contributes to N recycling in plants (Soltabayeva et al., 2018). Allantoin and allantoic acid are the dominant forms of assimilated-N transported from roots to shoots through the xylem in tropical leguminous plants (Schubert et al., 1986). Many plant species accumulate allantoin under abiotic stress such as salinity, drought, and heavy metal toxicity (Casartelli et al., 2019; Kaur et al., 2021; Lescano et al., 2016; Nourimand and Todd, 2016; Watanabe et al., 2013). The accumulated allantoin can enhance the abiotic stress tolerance of plants (Watanabe et al., 2013). We investigated the effect of biuret on the accumulation and metabolism of allantoin in rice seedlings.
Furthermore, we performed a metabolome analysis using rice suspension cells to examine metabolite changes under biuret toxicity.