Structure determination of compounds from alpine plants now possible with just 2 grams of flowers: National Museum of Nature and Science

A research group including the National Museum of Nature and Science has established a method capable of determining the structures of phenolic compounds contained in alpine plants using a minute flower sample of just 2 grams. Due to global warming and the resulting expansion of deer habitats that lead to grazing damage, population sizes of alpine plants are declining, making it increasingly difficult to collect samples. On the other hand, these plants are known to contain various polyphenols that enable them to endure harsh environments, and some of these components have proven useful for drug discovery. Being able to determine chemical structures from such trace amounts will help protect rare plants, offering the distinct advantage of allowing research to be conducted sustainably.

Senior Curator Yoshinori Murai at the Department of Botany, National Museum of Nature and Science (also a Guest Associate Professor at Tokyo University of Agriculture and Technology), who specializes in plant science, environmental adaptation, and chemical ecology, has been conducting research to extract and determine the structures of chemical substances contained in alpine plants. Every year from May to October, he goes mountaineering to collect plant samples for his research, while also engaging in conservation activities such as covering rare plants with nets to protect them from deer grazing. He has cultivated over 100 types of alpine plants collected so far in greenhouses and artificial climate chambers at the Tsukuba Botanical Garden (Tsukuba City, Ibaraki Prefecture), meticulously investigating their components while ensuring their preservation.

Alpine plants can unfold their leaves and grow in harsh environments exposed to intense sunlight, including ultraviolet rays, as well as severe cold. To adapt to these environments, they are known to contain phenolic compounds, which are organic compounds. Phenolic compounds are widely recognized as polyphenols. Certain components have already been utilized by humans as medicines and foods or applied to chemical products like agricultural medications. However, many alpine plants include rare, endangered, or endemic species. Large quantities cannot be brought back when collecting samples due to permissions/regulations and ethical considerations regarding environmental conservation.

Previously, the mainstream method for analyzing components contained in plants was structure determination using analytical instruments such as Nuclear Magnetic Resonance (NMR). However, this method required preparing at least several milligrams of purified powder samples, which translated to needing approximately dozens of grams of raw plant samples like flowers or leaves.

Murai and his colleagues established a method to crystallize trace amounts of purified samples by refining the solvent composition. By analyzing the crystals obtained through this method using either electron diffraction or X-ray diffraction, they successfully determined the structures of more than 10 components from the white flowers of D. lapponica subsp. obovata, which blooms on mountains from central Honshu northward. These components were found to include quercetin glycosides, which possess strong antioxidant properties and ultraviolet absorption capabilities.

X-ray diffraction is utilized for single crystals with a size of approximately several dozen micrometers (a micrometer is one-millionth of a meter), while electron diffraction is used for even smaller crystal samples of about a few micrometers. Although the flowers of Diapensia lapponica subsp. obovata are small, measuring about 1 cm in diameter, and the leaves are only 5 to 10 mm, this experiment successfully clarified the structures of over 10 components from an exceptionally small flower sample of just 2 grams.

Murai stated, "Reducing the required sample size by even a few grams will make it easier to advance future research. I want to apply this to plants with narrow distributions, such as species endemic to Japan or endangered species." Using this method not only reduces the environmental burden on natural habitats but also offers the advantage of allowing analysis from small samples of rare, miniature plants being cultivated inside artificial climate chambers.

Due to global warming, impacts are beginning to emerge, such as plants that originally grew wild at lower altitudes now being observed at higher elevations. According to Murai, grazing damage by deer is worsening in areas like the Southern Alps, where the mountain scenery is beautiful and precious creatures like the rock ptarmigan can be found. He emphasizes that alongside protecting rare animals, the conservation of various plants growing on the mountains must be addressed with urgency.

This research was conducted with support from the Grants-in-Aid for Scientific Research (KAKENHI) by the Japan Society for the Promotion of Science and the National Museum of Nature and Science's integrated research project funding for "Science of Extreme Environments." The results were published on February 22 in the online edition of Journal of Molecular Structure, a Dutch scientific journal. A joint announcement was made on March 31 by the National Museum of Nature and Science, Rigaku Corporation (Akishima City, Tokyo Prefecture), Asterism LLC (Taito City, Tokyo Prefecture), and Tokyo University of Agriculture and Technology.