Black Cohosh Cultivation in Missouri, and Quantification of its Medicinal Compounds in Response to Various Cultivation Practices

A horticultural research program on black cohosh (Actaea racemosa or Cimicifuga racemosa) was initiated in 2001 under the auspices of the Center for Phytonutrient and Phytochemical Studies, a research consortium between the University of Missouri - Columbia and the Missouri Botanical Garden. The consortium is funded by a substantial grant from the U.S. National Institutes of Health.

Background

Black cohosh is an attractive perennial herb in the buttercup family (Ranunculaceae) native to the midwestern and eastern United States. The species has been documented as occurring in 30 Missouri counties, all south of the Missouri River (Steyermark, 1977). It is generally found growing in rich, fertile, well-drained soils as an understory plant in deciduous forests, suggesting that the species may be suitable for cultivation under tree crops in an agroforestry-type setting.

Black cohosh has become a very popular and costly medicinal herb in recent years, with consumer demand increasing in North America and Europe. The root has been used historically to treat a variety of human ailments including sore throat, rheumatism, menstruation and uterine difficulties, menopausal and premenstrual syndrome symptoms, and as an astringent, diuretic, anti-diarrheal, cough suppressant, and diaphoretic (Tyler, 1993). Presently the herb is approved in Germany for treating premenstrual discomfort and menopausal ailments, and is commonly used by North American women to replace or supplement estrogen treatments in hormone replacement therapy. The root putatively contained estrogens or produced some sort of estrogenic activity in humans, but this has recently been discounted (Li, et al. 2002). Nevertheless, the root has been clinically successful at treating menopausal hot flashes and is therefore in great and growing demand. A number of phytochemical compounds from black cohosh are of medicinal interest, including actein, deoxyactein, acetyl-acteol, cimicifugoside, cimicifugin, cimicigenol, and caffeic acid.

Much of the black cohosh presently being marketed is wild-harvested, with very few plants under managed cultivation. While the plant is not considered rare, it is by no means abundant. Over-harvesting of wild plants and poaching in parks and nature reserves is becoming an important issue. 95% of the U.S. wild harvest is exported, and the species is being considered for inclusion under the Convention on International Trade of Endangered Species (CITES). This entire scenario underscores the need to study the cultivation and propagation of black cohosh. The plant appears to be fairly easy to cultivate, but very little information is available for those who may wish to grow it commercially. The species may be especially well-suited as a horticultural crop for Missouri farmers wishing to diversify their operations.

Objectives

Eight experiments are presently underway. We are studying and documenting a variety of horticultural issues related to the cultivation and utilization of this valuable crop. Our goal is to understand more about the nature of the species and its medicinal compounds while developing detailed recommendations for farmers wishing to cultivate the plant profitably.

Methodology

Our cultivation experiments are being conducted at two geographically different sites in Missouri: the University of Missouri-Columbia’s Southwest Research Center, Mt. Vernon (37° 4' lat, 93° 53' long, and 378 m alt), and the Shaw Nature Reserve of the Missouri Botanical Garden, Gray Summit (38° 28' lat, 90° 48' long, and 186 m alt). The soil at Mt. Vernon is a prairie-derived Hoberg silt loam (fine-loamy, siliceous, mesic Mollic Fragiudalf) that is moderately well drained. The soil at Gray Summit is a Hartville silt loam (fine, mixed, mesic Aquic Hapludalf) with a 5% slope. Soils were amended with compost and basic nutrients as deemed appropriate after laboratory analysis. A shade house measuring 240 by 40 feet and built of cedar poles, steel cabling, and woven shade cloth was erected at each site. Each shade house is divided into 12 plots of 40 by 20 feet each. Shade cloth with various degrees of shading can be attached over each plot, or left uncovered if desired. Both locations also contain natural woodlands where additional black cohosh plantings have been established. Much of the original plant material for these experiments was acquired from Elixir Farm Botanicals, LLC, Brixey, MO. The material under study has been vouchered and confirmed as Actea racemosa [17 Sept. 2002, A. Thomas 35 (MO); 16 June 2003, A. Thomas 36 (MO); 18 June 2003, A. Thomas 37 (MO)].

Experiment 1: Quantification of the production and accumulation of medicinal compounds in black cohosh plant tissues of known age in response to three shading regimes.

Little is known about the appearance, abundance, and quality of various phytochemicals in black cohosh tissues as the plants age and mature. Furthermore, the response of the plant in terms of production and accumulation of these phytochemicals to stress and various shading regimes is unknown. Standards have recently become available for the phytochemicals 27-deoxyactein and cimiracemoside A, allowing us to quantify amounts of these phytochemicals in plant tissues. This experiment will answer questions about when and under what conditions (including stress) these phytochemicals are produced and in what quantity. Plant roots of known age have been harvested annually for three years. We are documenting production of the phytochemicals of interest from plants grown under the various shading regimes (0, 40, and 80%), and at what age specific phytochemicals appear and become most abundant. Black cohosh seeds of southwest Missouri provenance were stratified and germinated in late winter 2000/01 by Elixir Farm Botanicals, LLC, Brixey, MO. Approximately 205 seedlings were transplanted at each site, with 16 to 17 plants per shade treatment planted 0.5 m apart. The experiment was planted 13 July 2001 at Mt. Vernon and 16 July at Gray Summit. Harvests have now been conducted in 2002, 2003, and 2004.

Experiment 2: Root tissue and phytochemical yield from black cohosh in response to plant spacing and three shading regimes.

This study is evaluating black cohosh plants grown at various spacings and under varying degrees of shading to determine the optimum horticultural conditions for healthy plant growth and maximum yield of large, marketable roots and potent medicinal compounds. Other data useful to growers, such as insect and disease prevalence, and seed production under the various shading and spacing regimes will also be documented. Large, robust, uniform black cohosh root crowns, grown by Elixir Farm Botanicals, were planted 11 and 15 Oct. 2001 at Mt. Vernon and 11, 12, 18, and 22 Oct. at Gray Summit. Before planting, crowns were diligently sized, sorted, and randomized to ensure the most uniform and random sizes possible per plot. Three shade treatments (0, 40, and 80% shade) were randomly assigned to the 12 shade house plots. Within each plot, 36 root crowns were planted 0.5 m apart and 36 plants 1.0 m apart, each in a square, block-type arrangement of 6 by 6 plants, for a total of 864 plants per site. The 20 exterior plants per block serve as border plants, while the interior 16 plants are considered the experimental plants. The spacing treatments are considered split-plots within shading treatments. Additional same-aged crowns were planted next to this study for occasional monitoring and sacrifice throughout the duration of the study. Harvest is anticipated in fall, 2004.

Unfortunately, most of the plants at the Gray Summit site succumbed over the first winter to a Phytophthora fungal root rot. This diagnosis was confirmed by the University of Missouri Plant Diagnostic Clinic, although the species of Phytophthora was not determined. While unfortunate, this situation actually presents a unique opportunity to conduct additional important research on dealing with such potential root rots, which may be important to farmers wishing to establish black cohosh as a crop (see Experiments 5, 6, and 8). The experiment continues at the Mt. Vernon site.

Experiment 3: Evaluating the suitability of black cohosh as an understory agroforestry intercrop with black walnut.

This experiment, initiated in summer, 2002, will help determine the suitability of black cohosh as an intercrop with black walnut (Juglans nigra) in an agroforestry setting. Many species of plants are stunted or killed by "juglone", a substance produced by the roots of black walnut trees. It is not known if black cohosh is susceptible to juglone, and whether or not the species may be compatible with black walnut. If black cohosh, a shade-loving plant, could be grown under black walnut as an agroforestry intercrop, such a system may provide excellent opportunities for Missouri farmers to grow and harvest two compatible crops together. The study is being conducted at both the Mt. Vernon and Gray Summit sites. At each site, 20 black walnut, 10 white oak (Quercus alba), and 10 mixed hickory (Carya spp.) seedlings were transplanted into 5-gallon pots in spring, 2002. We anticipate the trees filling the pots with roots during the growing season. In spring, 2003, we planted one black cohosh and one tomato plant in most of the pots, with single species in other pots. Tomato is extremely sensitive to juglone and will serve as a control: it should be affected by the walnuts but not by oak or hickory. We can then observe the black cohosh for similar results, assuming that if tomato is harmed and black cohosh is not, that black cohosh is not susceptible to juglone. If for some reason the tomato plants in the black walnut pots are not damaged, we might assume that not enough juglone has been produced to validate the experiment. The experiment is being continued through 2004.

Experiment 4: Quantification of the production, accumulation, and breakdown of medicinal compounds in black cohosh root, leaf, and stem throughout the growing season.

The root of black cohosh has traditionally been used for medicinal purposes. Little is known about the presence, production, accumulation, and breakdown of various phytochemicals in black cohosh roots and other tissues throughout the growing season. In other medicinal crops, such as ginseng, researchers have determined that desirable phytochemicals commonly occur in high quantities in plant tissues other than those that are traditionally consumed. Standards have recently become available for 27-deoxyactein and cimiracemoside A, allowing us to quantify these phytochemicals in plant tissues. Approximately 175 vigorous, mature root crowns were planted in established woodlands at Shaw Nature Reserve on 9 Oct., and at Southwest Center on 7-8 Nov. 2002 for this study. Harvest of six random plants at each location was conducted every two weeks from leaf emergence in spring to leaf senescence in fall, 2003 and 2004. Harvested plant tissues are separated into leaf, stem, flowers, fine roots, and crown roots, then washed, chopped up, frozen, and freeze-dried. Tissues are then analyzed for the two phytochemicals.

Some preliminary results are presented in the two tables below. The phytochemical 27-deoxyactein is the compound by which dosage of black cohosh root is now standardized. Interestingly, compared with crown root (the tissue most often consumed) we found approximately 10 times the amount of this compound in leaf tissue and 50 times the amount in flowers (Figure 1). Comparatively less was found in fine root and stem. We saw virtually no change (accumulation or degradation) in content of 27-deoxyactein in any of these plant tissues throughout the season. With cimiracemoside A (Figure 2), we found no detectable amount of this substance in leaves, and varying amounts in other plant tissues. In general, more cimiracemosode A was found in roots compared with stem and flower. Again, no clear seasonal pattern of accumulation or degradation of this substance was detected.

Experiment 5: Effect of planting depth and fungicide treatment on establishment of black cohosh root crowns in a heavy soil.

A shade house was constructed at Shaw Nature Reserve, Gray Summit, MO for the purpose of studying black cohosh cultivation. Experiments were initiated in fall, 2001 by planting large root crowns. Unfortunately, the majority of the root crowns were attacked by a Phytophthora fungal root rot disease over the first winter and perished. While unfortunate, this situation presented a unique opportunity to study the establishment of black cohosh under less than ideal conditions; a situation likely faced by many farmers wishing to cultivate the plant commercially. We took advantage of the known presence of this especially virulent Phytophthora population at the site, and evaluated the susceptibility of the root crowns at two planting depths in combination with a fungicidal treatment. During the growing season, the shade house was covered with a fabric blocking 40% of the available sunlight.

On 8 Oct. 2002, 288 large, robust black cohosh root crowns were planted to initiate this experiment at Shaw Nature Reserve. Four treatments were replicated 12 times across the shade house with 6 plants per treatment plot. Treatments were 1) shallow planting, 2) deep planting, 3) shallow planting with fungicide, and 4) deep planting with fungicide. The tops of the root crowns were set just under the soil surface for shallow planting and approximately 2.5 inches for deep planting. Two days after planting, a soil drench fungicidal treatment was prepared by diluting 21.3 % a.i. mefenoxam [N-(2,6-dimethylphenyl)-N-(methoxyacetyl)-D-alanine methyl ester] (trade name Subdue Maxx) at a rate of 1 fl.oz. per 3 gal. water, then applying 1 pint of this solution to 1 ft² of soil surface surrounding the appropriate plants. Survival and vigor of plants in response to these four treatments were documented during the 2003 and 2004 growing seasons. Results have not yet been evaluated.

Experiment 6: Effect of fall versus spring planting and fungicide treatment on establishment of black cohosh root crowns in a heavy soil.

A second experiment designed to take advantage of the Phytophthora root rot situation described above compared fall versus spring planting of black cohosh in combination with a fungicide treatment. While fall planting is generally preferred, the dormant root crowns lie susceptible to root rot for several months over winter until spring growth commences. Overwinter cold storage of roots followed by spring planting may provide an alternative where heavy soils are involved and root rot is more likely.

The experiment was initiated in fall, 2002 at the Shaw Nature Reserve shade house, which is covered with a fabric blocking 40% of the available sunlight during the growing season. Four treatments of 6 plants each were established with 12 replications across the shade house, for a total of 288 plants. Treatment plots were situated randomly within each replication area of the shade house. Treatments are 1) fall planting, 2) fall planting with fungicide, 3) spring planting, and 4) spring planting with fungicide. On 8 Oct. 2002, the fall planting of 144 plants was completed while the spring planting of 144 stored roots was installed on 2 April 2003. The tops of all root crowns were set approximately 1 inch below the soil surface. Two days after the fall planting, and eight days after the spring planting, a soil drench fungicidal treatment was prepared by diluting 21.3 % a.i. mefenoxam [N-(2,6-dimethylphenyl)-N-(methoxyacetyl)-D-alanine methyl ester] (trade name Subdue Maxx) at a rate of 1 fl.oz. per 3 gal. water, then applying 1 pint of this solution to 1 ft² of soil surface surrounding the appropriate plants. Survival and vigor of plants in response to these four treatments were documented during the 2003 and 2004 growing seasons. Results have not yet been evaluated.

Experiment 7: Effect of gibberellic acid and ethephon (2-chloroethylphosphonic acid) on germination of black cohosh seeds.

Achieving reliable and consistent germination of expensive black cohosh seed is one of the most important obstacles to the establishment of black cohosh as an alternative agricultural crop. Upon harvest, the seeds apparently go into various stages of dormancy that can make germination very uneven and unreliable. Some seeds can remain dormant for years. Many techniques are used on seeds of various plant species to overcome dormancy and produce consistent germination. Two commonly-used substances that often produce good results are gibberellic acid and ethephon. Gibberellic acid is a plant hormone often produced by germinating seeds. Ethephon is an ethylene-releasing substance that stimulates seed germination in many species. Numerous reports have been published indicating that these two substances, alone or in combination, produce good germination in a variety of species (e.g., Sari et al., 2001). Theses studies are being initiated in summer, 2003. Various concentrations and combinations of gibberellic acid and ethephon will be used to treat black cohosh seeds before sowing them on moistened filter paper and placing them in an incubation chamber. Results from this initial seed germination study may lead to further studies toward streamlining germination of black cohosh seeds.

Experiment 8: Fungal Diseases of Black Cohosh that may be Important in Cultivation.

A disease now called "Black Root and Crown Rot of Black Cohosh" has been characterized. This disease (singly or in combination with others) has devastated cultivated plants at our Gray Summit site. After morphological characterization, three Phytophthora species and a number of Pythium species have been associated to the disease. It is possible that the Phytophthora species play a major role in the problem and that perhaps some moderately pathogenic Pythium species are part of a disease complex. We are confirming the identity of these organisms via molecular characterization. The Koch’s Postulate procedure is being followed to elucidate the true pathogenicity of these fungi with black cohosh.

References

Li, W., S. Chen, D. Fabricant, C. Angerhofer, H. Fong, N. Farnsworth, and J. Fitzloff.