Sabo, Ian, Jonathan L. Horton*, H. David Clarke, and Jennifer Rhode Ward. Biology Department, University of North Carolina Asheville. *Corresponding author
(Presented at The Future of Ginseng and Forest Botanicals Symposium, July 12-14, 2017, Morgantown, WV)
Abstract
Panax quinquefolius L. (American ginseng) is an economically important, but increasingly threatened, herbaceous perennial plant native to eastern North America. The roots have long been prized in traditional Asian medicine, and are increasingly being used in producing herbal supplements for other markets. Much research is focused on ginsenosides, medicinally-active compounds found in ginseng roots. Given the conservation concerns regarding wild P. quinquefolius, and the use of roots for both commercial and research activities, we began experimenting with a partial-root harvest method in 2014. This approach was designed to extract tissue for ginsenoside analysis without reducing the plant’s fitness or resulting in mortality. Partial-root harvest samples were taken from 57 plants in four wild populations from western North Carolina. Of the 57 plants subjected to partial-root harvest in 2014, 51 (89%) reemerged in 2015 and 45 (79%) reemerged in 2016. These resprout rates were similar to paired unharvested plants (86% and 81%, respectively). Fitness (berry production) was not affected by partial-root harvest, however, there were short-term effects on growth. Harvested plants had significantly shorter stems and smaller leaf area than unharvested plants in the first year after harvest. However, there were no significant differences in any plant metric between these two groups of plants by the second year after harvest. Our results suggest that this method could be an effective way for researchers to reduce their impact on wild ginseng populations.
Keywords: American ginseng, non-destructive harvest, Panax quinquefolius, phytochemical analysis, sustainability
Introduction
Panax quinquefolius L. (American ginseng) is an economically important, perennial herb endemic to the deciduous forests of eastern North America (Anderson et al. 1993). The roots have long been prized in traditional Asian medicine, and are beginning to be used more in North America and Europe as herbal supplements (Schlag and McIntosh 2006). This use has led to an increased demand for the roots of this species, resulting in overharvesting, increasing rarity, and loss of genetic diversity in much of its native range (Cruse-Sanders and Hamrick 2004). Researchers are interested in determining the composition of root phytochemicals, particularly the triterpenoid saponins known as ginsenosides (Qi et al. 2011), that are major medicinally-active compounds. Previous studies have demonstrated that plants’ chemotypes vary between organs (leaves and root) in cultivated (Li et al. 1996) and wild (Searels et al. 2013) plants, and among roots of individual plants (Searels et al. 2013; Schlag and McIntosh 2013). While some variation can be attributed to genetic factors (Schlag and McIntosh 2013), the production of certain ginsenosides (Rb1, Rd, Rc) seems to be environmentally determined (Lim et al. 2005). However, most previous research isolating root ginsenosides for analysis has relied on total root harvest killing the plant.
Given the conservation concerns regarding American ginseng, we began experimenting with a non-destructive, partial-root harvest method in 2014 in an attempt to extract tissue for ginsenoside analysis without increasing plant mortality or reducing plants’ vegetative fitness. We predicted that carefully harvesting small amounts (~ 300 mg) of fresh tissue, then immediately replanting the remaining root, would have no negative effects on growth or survival compared to similarly sized unharvested plants. If successful, this partial-root harvest method would allow researchers to assess root phytochemicals with less overall mortality.
Materials and Methods
For this study, we monitored 114 mature (three or four leaf) P. quinquefolius plants (Fig. 1) from four protected wild populations in western North Carolina. Of these plants, 57 were partially root harvested in 2014, while the other 57 were unharvested. When roots were exposed, side roots branching from the main root were harvested (Fig. 2). If no side root was present, then part of the side of the main root was harvested with care to not damage the vascular cylinder in the center of the root. Roots were then replanted into the same spot from which they were dug.
We measured morphological data – reproductive status, number of berries (if present), number of leaves, total number of leaflets, stem height (cm), peduncle length (cm, if present), largest leaf rachis (cm), largest leaflet length (cm), and largest leaflet width (cm) in 2014 before partial-root harvest and again in 2015 and 2016 mid growing season. Leaflet length and width were used in an allometric equation developed by Mooney and McGraw (2009) to calculate leaf area of the largest leaf.
We used Analysis of Variance to compare the number of berries, number of leaves, number of leaflets, stem length, and leaf area between partial-root harvested plants and unharvested plants before harvest and one year and two years after harvest. Survivorship was compared between the partial-root harvested plants and the unharvested plants one and two years after harvest using Chi-squared tests.
Results and Discussion
Before harvest in 2014, there were no differences in morphological metrics between partial-root harvested and unharvested plants (Table 1; Fig. 3), confirming that control and partial-root harvest plants were appropriately paired. There was no significant difference in reemergence between partial-root harvested and unharvested plants either the first year (χ2 = 0.988, p = 0.568, harvested 51/57 – 89% and 49/57 – 86%) or second year (χ2 = 0.999, p = 0.815, harvested 45/57 – 79% and unharvested 46/57 – 81%) after harvest.
Table 1. Statistical results from Analysis of Variance comparing morphological parameters between partial-root harvested and unharvested plants.
Fitness (berry production) was not affected by partial-root harvest, however, there were short-term (one year) effects on growth. Mean number of berries decreased in the first year after harvest (2015) for both non- harvested and partial-root harvested plants, and this decrease was not significantly different between the two groups (Table 1; Fig. 4). Possible reasons for this decrease in berry production across both groups and all populations could include inter-annual differences in temperature and/or precipitation, although these factors were not addressed in this study. Neither the number of leaves nor the total number of leaflets per plant differed between partial-root harvested and unharvested plants in either year after harvest (Table 1, Fig. 4 & 5). The differences in number of leaves and leaflets between years is also likely due to environmental conditions and future research should explore the effects of environmental variation of growth. Because ginseng growth is determinant and buds are formed late the previous summer, it is likely environmental conditions in late summer the year before that determine leaf and leaflet number in the current year. Both stem length and leaf area decreased between 2014 and 2015 for partial-root harvested but not for unharvested plants, and these differences were significant (Table 1; Fig. 4). This decrease in stem length and leaf area in partial-root harvested plants could be a response to simulated root herbivory from the partial-root harvest. Root and rhizome herbivory has caused size class reversion and dormancy in subsequent years (Farrington 2006).
By the second year after harvest (2016), number of berries, leaves and leaflets and stem length were all slightly lower than they were before harvest (Fig. 5), but were not significantly different between partial-root harvest and unharvested plants. Leaf area of unharvested plants was higher, while area of partial-root harvest plants was lower than pre-harvest values (Fig. 5). These values were highly variable in both groups, however, and were not significantly different between them (Table 1).
In summary, partial-root harvest had no significant effect on survivorship one or two years after harvest. There were significant decreases in stem length and leaf area in partial-root harvested plants in the first year after harvest, but these were not present in the second year, suggesting that partial-root harvest has little long-term effect on growth or survivorship over time. Berry production was lower in 2015 relative to 2014, but the decrease occurred in both partial-root harvested and unharvested plants suggesting other factors such as inter-annual variation in weather.
Because the partial-root harvest method had no significant effect on survivorship in either year and had marginal and short-lived effects on morphology, it could prove to be an effective non-destructive method for ginsenoside extraction for research applications. The partial-root harvest method could be used in research projects designed to test the environmental and genetic factors affecting the production of secondary metabolites while preserving plant material in the field. One hindrance to studying the phytochemical makeup of wild American ginseng populations has been the need to destructively harvest root tissue. This method will allow researchers to sample plants from wild populations without causing long-term population declines.
Acknowledgements
Funding was provided by the North Carolina Biotechnology Center (BRG-1206). Members of the UNCA Botany Research Group (L. Barton, J. Burroughs, S. Comito, L. Howe, M. Jasper, C. Lahue, M. Shattelroe, and A. Warren) assisted with field surveys.
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