Production of deuterated biomass by cultivation of Lemna minor (duckweed) in D₂O

Main conclusion: Common duckweed Lemna minor was cultivated in 50% D₂O to produce biomass with 50–60% deuterium incorporation containing cellulose with degree of polymerization close (85%) to that of H₂O-grown controls. The small aquatic plant duckweed, particularly the genus Lemna, widely used for toxicity testing, has been proposed as a potential source of biomass for conversion into biofuels as well as a platform for production of pharmaceuticals and specialty chemicals. Ability to produce deuterium-substituted duckweed can potentially extend the range of useful products as well as assist process improvement. Cultivation of these plants under deuterating conditions was previously been reported to require addition of kinetin to induce growth and was hampered by anomalies in cellular morphology and protein metabolism. Here, we report the production of biomass with 50–60% deuterium incorporation by long-term photoheterotrophic growth of common duckweed Lemna minor in 50% D₂O with 0.5% glucose. L. minor grown in 50% D₂O without addition of kinetin exhibited a lag phase twice that of H₂O-grown controls, before start of log phase growth at 40% of control rates. Compared to continuous white fluorescent light, growth rates increased fivefold for H₂O and twofold for 50% D₂O when plants were illuminated at higher intensity with a metal halide lamp and a diurnal cycle of 12-h light/12-h dark. Deuterium incorporation was determined by a combination of ¹H and ²H nuclear magnetic resonance (NMR) to be 40–60%. The cellulose from the deuterated plants had an average-number degree of polymerization (DP_n) and polydispersity index (PDI) close to that of H₂O-grown controls, while Klason lignin content was reduced. The only major gross morphological change noted was root inhibition.