Isolation and characterization of the hydrophilic BNI compound, 6-methoxy-2(3H)-benzoxazolone (MBOA), from maize roots

Abstract

Background and aimsBiological nitrification inhibition (BNI) is a chemical ecological phenomenon whereby plants specifically suppress nitrification by releasing inhibiting compounds from roots, an effective strategy for improving nitrogen uptake by limiting nitrogen losses from agricultural fields. During this study, we have aimed at characterizing hydrophilic BNI activity released from maize roots to understand the chemical basis for BNI function in maize.MethodsMaize plants were grown hydroponically and root exudates were collected using water-based solutions and hydrophilic BNI activity was extracted. We isolated the target BNI compounds by a combination of chromatographic techniques and bioassays using a recombinant luminescent ammonia-oxidizing bacterium Nitrosomonas europaea (pHLUX20).ResultsWe identified 6-methoxy-2(3H)-benzoxazolone (MBOA) as the responsible BNI compound with a median effective dose (ED50) = 0.76 μM. MBOA inhibited the conversion of NH3 to NH2OH as well as NH2OH to NO2− in N. europaea, suggesting that MBOA blocks both ammonia monooxygenase and hydroxylamine oxidoreductase enzymatic pathways. Treatment with MBOA significantly suppressed NO2− and NO3− production during soil incubation, but this activity was reduced subsequently due to biodegradation of MBOA by soil microbes. A quantification experiment revealed that MBOA accounted for nearly 50% of the total BNI activity in hydrophilic and hydrophobic exudates from maize roots. A soil incubation test showed that two previously identified benzoxazinoids, HDMBOA and HDMBOA-β-glucoside, can be eventually transformed into MBOA.ConclusionWe elucidated MBOA as the key component of BNI in maize. Collectively, the present findings will serve as the groundwork for construction of an advanced environment-friendly agricultural system.