O Absorption, Translocation, and Metabolism of Prohexadione Calcium in Annual Bluegrass (Poa annua), and Three Turfgrass Species Goddard, M.J., J.B. Beam, S.D. Askew, Virginia Tech, Blacksburg O * O H * O O Partition R2 values Leaf 0.99 y = -0.02x2 + 1.18x + 45.04 Rinse 0.90 y = -9.76Ln(x) + 47.64 Foliage 0.98 y = 2.99Ln(x) + 5.08 Root 0.89 y = 1.48Ln(x) + 1.19 CO2 0.82 y = 1.06Ln(x) + 1.32 Introduction ______ Prohexadione Ca (PC) is an experimental turfgrass growth regulator that selectively controls or suppresses annual bluegrass in desirable turfgrass such as creeping bentgrass (Agrostis stolonifera), Kentucky bluegrass (Poa pratensis), and perennial ryegrass (Lolium perenne). PC inhibits gibberellin production in turfgrass similar to trinexapac-ethyl (Rademacher, 2000). Previous research indicates that annual and Kentucky bluegrass growth is suppressed more by prohexadione Ca than creeping bentgrass and perennial ryegrass. Methods and Materials cont._______ Plants partitioned into treated leaf, other foliage, roots, CO2, and rinse Radioactivity detected by LSS. Plant parts were combusted in a biological oxidizer, prior to LSS Radioactivity (% of recovered) Results _______ Annual and Kentucky bluegrass absorbed more prohexadione Ca than creeping bentgrass and perennial ryegrass when averaged over harvest timing and trial Neither translocation out of the treated leaf or metabolism of prohexadione Ca differed between species When averaged over species and trial, 22% of recovered prohexadione Ca was metabolized within 1 HAT, and plants metabolized an additional 0.7% each additional hour for a period of 48 hours Time after treatment (h) Figure 1: Effect of time after treatment on radioactivity distribution averaged over plant species and trial. Objectives ______ Determine absorption, translocation and metabolism rates of 14C PC when applied to foliage to elucidate reasons for differential plant response. Prohexadione Calcium (RF = 0.58) Metabolite 1 (RF = 0.12) Standard Kentucky bluegrass* treated leaf Radioactivity (CPS) Metabolite 2 (RF = 0.33) Materials and Methods ________ 2 laboratory trials, RCBD with 3 replications each Plants were field collected and thinned to one tiller Plants placed in 0.25% Hoagland's solution and maintained at 20/30 C night/day with an average irradiance of 80 µmol/m2/s PAR Spotted with three 1 µl drops of 14C PC with 91% purity and 2.1 kBq radioactivity, in 88% 1M HCL, 12% Acetonitrile, and 0.25% v/v nonionic surfactant, to newest fully expanded leaf Plants were harvested at 1, 12, 24, and 48 hours after treatment (HAT) Air derived from each plant was vacuumed through a 50 ml glass burette containing 20 g of glass beads and 20 ml of 3M NaOH and rinsed with 20 ml acetonitrile: water (3:7) 1 ml aliquot of rinse + 19 ml scintillation cocktail was analyzed with a liquid scintillation spectrometer (LSS) Radioactivity partitioning Species CO2 Rinse Treated leaf Foliage Roots --------------------------- % --------------------------- Annual bluegrass 4a 17c 53b 15 11a Creeping bentgrass 2b 31a 46b 11 9bc Kentucky bluegrass 3ab 15c 60a 12 10ab Perennial ryegrass 24b 54ab 13 7c Distance (mm) Figure 2: Radiochromatogram trace peaks of PC standard and Kentucky bluegrass 48 HAT, showing displacement of PC with polar metabolites. *Kentucky bluegrass values were adjusted to meet the levels of the standard. 100 Conclusion _______ Annual and Kentucky bluegrass sensitivity to PC is partially explained by increased absorption rates but may be due to factors other than translocation and metabolism 80 y = 0.69x + 74.16 R2 = 0.93 60 Prohexadione calcium Polar metabolites Radioactivity (% of recovered) 40 y = 0.65x + 25.62 R2 = 0.93 20 Reference _______ Rademacher, W. 2000. Growth retardants: effects on gibberellin biosynthesis and other metabolic pathways. Annu. Rev. Plant Physiol. Plant Mol. Biol. 51:501-531 10 20 30 40 50 Time after treatment (h) Figure 3: Effect of time after treatment on percent PC and polar metabolites in treated leaves, averaged over plant species and trials. Preliminary experiment determined that appreciable PC was lost as radioactive CO2 when exposed to plants Three drops (3 µl) of PC placed on each plant Chambers trapping air released from plant to be collected and analyzed for 14C CO2 Air vacuumed through burette containing glass beads with 3M NaOH to trap CO2 Plant parts were clipped and separated for analysis Treated leaves and other plant parts were cut, rinsed, dried, and combusted Absorbed PC was separated from metabolites via thin layer chromatography