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Kent E. Cushman, Rita M. Moraes, Patrick D. Gerard, Ebru Bedir, Bladimiro Silva, and Ikhlas A. Khan

Leaves of American mayapple (Podophyllum peltatum) are being investigated as an alternative and renewable source of podophyllotoxin, a pharmaceutical compound used in the manufacture of several drugs. This study examined long-term performance of mayapple populations subjected to different harvest strategies. A naturally occurring population in shade was subjected to leaf removal treatments of frequency (every year, every 2nd or 3rd year) or timing (early or late season). Plots were 1.0 m2, established during Spring 2001, and treatments were applied from 2001 to 2004. Control plots not previously harvested were also included each year. Plants did not tolerate the severest of leaf removal treatments: early harvest time in combination with annual harvest frequency. Early annual harvests reduced total leaf dry mass and total leaf area in a quadratic manner. Late harvest conducted annually, and early harvest conducted every other year, also reduced leaf dry mass and area but not as much as early annual harvest. Plants harvested every year, early, or early every year produced fewer sexual shoots than other treatment combinations. Contents of α-peltatin, β-peltatin, and total lignans were higher for leaves harvested early than those harvested late during each year of the study, demonstrating that lignan contents were affected by leaf age and not treatment. In conclusion, our results for plants grown in shade show that leaves can be removed late in the growing season every 2nd or 3rd year or early every 3rd year without reducing long-term performance of the population. This is more restrictive than that reported for populations in full sun where plants tolerated late harvests every year.

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Muhammad Maqbool, Kent E. Cushman, Rita M. Moraes, and Patrick D. Gerard

American mayapple (Podophyllum peltatum L.) is a rhizomatous, herbaceous perennial found in wooded areas of eastern North America and is a source of the pharmaceutical compound podophyllotoxin. This research was conducted to determine the optimum duration of low temperature exposure in overcoming dormancy of fall-harvested rhizome segments for subsequent use as propagules in greenhouse plantings. Two types of rhizome segments were harvested from the wild and used in this study: two-node rhizome segments consisting of a terminal node and its adjacent one-year-old node and one-node rhizome segments consisting of a single node, other than a terminal node, of unknown age or rhizome position. For growth cycle I, rhizome segments were exposed to low temperature (≈4 °C) for 30, 45, 60, 75, or 90 days, planted in pots, and grown in a greenhouse set at 21 °C. Shoot emergence, shoots per pot, and plant height were recorded. Leaves were removed from plants when senescence first became evident, and leaf area was recorded. For growth cycle II, rhizome segments remained undisturbed in the original pots and were exposed to low temperatures (≈4 °C) for 90 days. Pots were again placed in the greenhouse and shoot emergence, shoots per pot, plant height, and leaf area were recorded. Increasing duration of low temperature exposure of rhizome segments up to 75 days appeared to increase percent emergence and plant height and decrease days to emergence, though changes in greenhouse conditions over the study period may have also influenced shoot emergence and plant growth. Two-node rhizome segments exhibited higher percent shoot emergence, shoot longevity, leaf area, and plant height than single-node segments during each growth cycle. Two-node rhizome segments also exhibited fewer days to emergence during growth cycle I. Rhizome segments produced no more than a single shoot in growth cycle I, whereas more than one shoot was produced in growth cycle II. Most of the effects of low temperature exposure during growth cycle I persisted throughout growth cycle II. These results indicate that dormancy of mayapple rhizomes can be overcome with low temperature exposure and shoots can be induced to grow at any time of year.

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Kent E. Cushman, Muhammad Maqbool, Hemant Lata, Ebru Bedir, Ikhlas A. Khan, and Rita M. Moraes

Four levels of shade (0%, 30%, 55%, and 80%) were used to determine their effect on growth and lignan content of american mayapple (Podophyllum peltatum L.). Mayapple rhizomes were harvested from the wild and transplanted into plant beds on 20 Dec. 2001 using a randomized complete block design with four blocks. Growth and lignan content were recorded during spring of 2002 and 2003. Leaf samples were analyzed for the following lignans: podophyllotoxin, alpha-peltatin, and beta-peltatin. Increasing levels of shade increased shoot longevity, leaf area per plant (cm2/plant), and shoot height. Shade did not affect shoot emergence, total leaf area (cm2·m-2), or leaf dry mass (g·m-2 or g/plant). Regardless of year, podophyllotoxin and total lignan contents at 0% shade were significantly greater than those at 80% shade, and the overall trend was for decreasing contents with increasing shade. Shade did not affect alpha-peltatin content. Content of beta-peltatin was greatest at 0% shade compared to the other three shade treatments. Year affected alpha-peltatin and beta-peltatin contents, with less content of each in 2003 than in 2002. There were large numerical decreases in podophyllotoxin yield (podophyllotoxin content per unit area, mg·m-2) as shade increased from 0% to 80%, but these differences were only marginally significant (P = 0.0897). In contrast, podophyllotoxin yield was significantly greater in 2003 than in 2002 as total leaf area and dry mass significantly increased. Increasing levels of shade slightly decreased air and soil temperatures. Our results indicate that american mayapple is not a shade-requiring species. Under full sun (0% shade) shoots did not persist as long as under shade and leaves were smaller and thicker, but total lignan content was significantly greater than under shade. It appears that growers of specialty crops serving the pharmaceutical industry can establish and cultivate american mayapple under full sun, thus providing leaf biomass with high podophyllotoxin content while avoiding the cost of expensive shade structures.

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Rita M. Moraes-Cerdeira, Jeffrey V. Krans, James D. McChesney, Ana M.S. Pereira, and Suzelei C. Franca

Cotton fibers were tested as a substitute for agar in tissue culture. The cost of agar has prompted us to search for an alternative more economical medium support. Effectiveness as a medium support was evaluated in terms of callus maintenance and shoot organogenesis using Artemisia, Agrostis, and Taxus. Taxus and Agrostis calli cultivated on liquid media with cotton fiber as medium support (25 ml of medium per gram of cotton) grew better than calli on agar (0.8% w/v). There were no significant differences in shoot organogenesis of Artemisia and Agrostis grown in 25 ml of medium per gram of cotton from those grown in agar medium.