Research on medicinal plant resources in coastal districts of Thai Binh province and recommendation of measures of conservation and sustainable development

used for the drug to save human lives. These compounds are screened on only about 6% of total plant species. Therefore, it is extremely great demand to exploit and process these plant resources for the treatment of serious diseases such as AIDS, cancer, diabetes, .... The world today has more than 35,000 plant species used as medicine. About 2,500 species of medicinal plants are traded in the world, 90% of which are wild. Medicinal plant resources are potentially huge treasures that can help the world human beings take care of health in a timely and effective manner. 1.1.4. Research on conservation of medicinal plant resources in the world Currently there are only a few hundred species in the world, 20 to 50 species in India, 100 to 250 species in China, 40 species in Hungary, 130 to 140 species in Europe. 10 In 1993, there were 8,619 protected areas worldwide. By 1997, there were 12,754 protected areas recognized by the United Nations. In addition, more than 17,500 other points were not included in the UN list due to substandards. In addition, there are about 2,000 botanical gardens all over the world, each garden is kept and planted up to several thousand species, including many species of medicinal plants. The conservation of medicinal plant resources in the world is carried out in two main forms: In situ and ex situ conservation. 1.2. Research on medicinal plant resources in Vietnam 1.2.1. Situation of investigation and statistics In the history of Vietnam there have been many famous physicians who made researches about medicinal plants, such as Chu Tien with "Full draft of basic medicinal plants"; Tue Tinh with "The South’s magical medicine"; Hai Thuong Lan Ong with "Lan Ong’s mind"... The Party and State always pay attention to the investigation and research of medicinal plant sources in Vietnam. On February 27, 1955, President Ho Chi Minh set out the way to build Vietnam's scientific, national and popular medicine based on the combination of traditional medicine and modern medicine. The Ministry of Health is also interested in creating favorable conditions for oriental medicine to develop. The Institute of Medicinal Materials (Ministry of Health) together with the system of pharmaceutical research stations have investigated in 2,795 communes and wards of 351 districts and towns of 47 provinces and cities throughout the country to make significant contributions in investigating, collecting medicinal plant resources and sharing experience of using medicinal plants in traditional folk medicine. 11 1.2.2. Usable value and economic value According to the Institute of Medicinal Materials (2002), Vietnam has 3,948 species of medicinal plants, belonging to 1,572 branches and 307 plant families, surpassing 3,200 species recorded in the Vietnamese Medicinal Dictionary. Of which, over 90% were wild and 144 species were included in the "Conservation of rare and threatened medicinal plants in Vietnam" (2003) and the "Vietnam Red List of Medicinal Plants" (2006). According to IMS Health, Vietnam is in the list of 17 countries with a growing pharmaceutical industry. Pharmaceutical market in Vietnam has the highest growth in Southeast Asia, about 16% annually. In 2013, the total drug consumption is estimated to be $3.3 billion, with the potential for export of pharmaceuticals reaching $40 million to $50 million, which is estimated to increase to $10 billion by 2020. 1.2.3. Development potentials The number of medicinal herbs used in medicine is increasingly being recognized, of which over 90% of the total number of medicinal plants grows naturally. But through the survey, this number can be actually higher because knowledge about medicinal plants of some of the ethnic minorities is limited. Vietnam has 54 ethnic groups with different customs and traditions, they have been accumulating knowledge and experience of using plants to prevent and treat diseases to survive and develop. With abundant plant resources together with the knowledge and experience of medicinal plants, this is a great potential to invest in the research and creation of new medicines with high therapeutic effect. 1.2.4. Research and preservation of medicinal plant resources in Vietnam 12 The Government and the health sector have made efforts to conserve the biological resources in general and medicinal plant resources in particular. The State’s projects for preserving medicinal plants (Preserving medicinal plant gene sources - Institute of Medicinal Materials, Ministry of Health) or preserving models of medicinal plant gene sources in the State projects or projects of Non- governmental organizations (Conservation of medicinal plants of Dao people in Ba Vi, Ha Tay - CREDEP, Conservation of medicinal plant gene sources - Ministry of Health, Model of preservation and development of medicinal plants in Sa Pa; Medicinal plants in Na Ot, Son La, etc. of the Institute of Ecology and Biological Resources) have been established to maintain the conservation of rare medicinal plants. Over the past time, Vietnam has been maintaining the primary medicinal plants in the Nature Reserves. So far, the conservation of medicinal plants has been expanded to cover many national parks and nature reserves in Vietnam. 1.3. Research on biological active ingredients from medicinal plant resources in coastal districts of Thai Binh province Thai Binh is a province with many medicinal plants of high economic value, however, the research on medicinal plant resources is very limited. Some outstanding researches: - In 2013, Khai Ha Pharmaceutical and Medical Equipment Trade Joint Stock Company developed a theme of "Applying scientific and technological advances in producing and processing Angelica sinensis (Oliv.) Diels and Achyranthes bidentata Blume, for socio- economic development in Thai Binh Province”. - Mr. Nguyen Xuan Quynh, University of Natural Sciences, Vietnam National University, Hanoi wrote a theme "Researching the 13 current state of biodiversity and building a database on biological resources for management purposes, using the flora ecosystem in Tien Hai district, Thai Binh province". - In 2014, Le Minh Ha, Institute of Natural Products Chemistry, Vietnam Academy of Science and Technology has the topic: "Research and develop the process of extracting rotundin substance in limestone Stephania glabra (Roxb.) Miers planted at a pilot scale for trial production, applied in Thai Binh province. " 1.3.1. Situation of research of the Physalis angulata locally and internationally According to author Vo Van Chi, in Vietnam, there are four species of the Solanaceae family including P. angulata, P.alkekengi, P.peruviana and P.minima. Contrary to Vietnam, species belonging to the Physalis family are favourite species of research and investment in the world. Many positive results have been published by scientists around the world. In terms of chemistry, the main class of Physalis is the withanolide, followed by labdane diterpene, sucrose esters, flavonoids, ceramides and others. Compounds of isanolide are divided into two forms, the unaltered withanolide and the altered withanolide. 1.3.2. Situation of domestic and foreign research on Cudrania tricuspidata: The scientific name of this plant is Cudrania tricuspidata (Carr.) Bur, belonging to the family of Moraceae, under many other names. In China, roots and bark the trees are used to make herbal teas and functional drinks from thousands of years ago. Cudrania tricuspidata has also been used as a traditional medicine for the 14 treatment of inflammatory diseases, cancer, hepatitis, influenza and neuropathy. In recent decades, Cudrania tricuspidata is considered one of the traditional remedies used to treat cancer in Korea. Studies on chemical compositions show that Cudrania tricuspidata contains high levels of xanthone and flavonoid compounds with anti-cancer, anti-inflammatory, anti-obesity, and neuroprotective effects. The xanthone compounds called miraclexanthone D, L, M show strong activity against gastric cancer cells. Flavonoids containing prenyl such as senegalensin and isoerysenealine E have anti-inflammatory activity by inhibiting the production of NO in mouse macrophages; whereas xanthones contain the prenyl branch (for example, cudratrixanthones C, G-I, O, 3-O-methylcudratrixanthone G) and isoflavonoids (eg, cudraisatrix fl avones H-J) have a protective effect on the nervous system. In addition, enzyme inhibitors for tyrosine phosphatase 1B, neuraminidase, α-glucosidase, and tyrosinase enzymes are also found in Cudrania tricuspidata. 15 CHAPTER 2 SUBJECT, LOCATION, CONTENTS AND RESEARCH METHODS 2.1 Object, location and time of study The medicinal plant resources in vascular plants in the districts of Thai Thuy and Tien Hai in Thai Binh province. Time of study: 2014 – 2016. 2.2. Research contents 2.2.1. Research on plants - Investigating the composition of medicinal plants. - Current situation of exploiting, using and role of medicinal plants in socio-economic life of Thai Binh province. - Study on assessment and identification of threatening factors affecting Thai Binh medicinal plant resources. 2.2.2. Preliminary research on the chemical composition and bioactivity of some valuable medicinal plants - Sampling, methanol extraction for active screening. - Select some good activities to study the chemical composition. - Re-assess the biological activities of the isolated clean substances. 2.2.3. Proposed solutions for effective management, conservation and sustainable exploitation of medicinal plant resources in Thai Binh province 2.3. Research methods 2.3.1. Preparation - Collecting materials and books related to the field of study. - Collaborating with specialized agencies (plant chemicals). - Agreement on technical measures with instructors and prepare financial and logistical conditions for the investigation. 2.3.2. Methods of plant research - Inheritance method - Method of field survey by routes - Definition of scientific names 16 - Analysis of collected information and data - Quick interview method 2.3.3. Methods of chemical composition research a. Methods of isolation of compounds Primarily chromatographic methods, including conventional column chromatography, with adsorbents, are conventional phase silica gels, as well as ODS, YMC (RP-8 or RP-18) silica gel blends combined with chromatography (TLC), thin layer chromatography (PTLC) and high performance liquid chromatography (HPLC). b. Methods for specifying the structure of compounds Modern spectral methods including: IR infrared spectroscopy; Ultraviolet UV; Mass spectra of MS; One-way (1D) 1-H NMR, 13C- NMR, DEPT; Two-dimensional (2D) resonance spectra: COSY, HMQC, HMBC, NOESY... 2.3.4. Methods of expected biological activity research a. Cytotoxic activity assay on some human cytotoxic cell lines b. Antioxidant activity assay: - Check the antioxidant capacity of the active substance on liver cells. c. Evaluation of antimicrobial activity assay 17 CHAPTER 3 RESEARCH RESULTS AND DISCUSSION 3.1. Current situation of medicinal plant resources in coastal districts of Thai Binh province 3.1.1. Diversity of medicinal plants resources 3.1.1.1. List of medicinal plants The research has recorded 346 species belonging to 268 genera of 94 families belonging to 3 branches of vascular plant used as medicine in two coastal districts of Thai Binh province. A list of medicinal plants has been established with detailed data as shown in Appendix 1. Compared with the number of medicinal plants in Vietnam (4,472 species), the number of species is not high (346 species) but within the limited area of this locality, the resources of medicinal plants here are quite rich and diverse. 3.1.1.2. Diversity in families Data from the list of medicinal plants shows that: Of the 6 vascular plant branches in Viet Nam, there are 3 branches in Thai Thuy and Tien Hai districts of Thai Binh province: fern, pine and orchid branch. In these three branches of medicinal plants available here, the degree of diversity is quite high. However, the composition of the branches distributes unevenly, in which the dominance of medicinal plants in the jade branch accounts for 97.87% of the total number of medicinal plants collected. The remaining two branches account for a very small share. 3.1.1.3. Diversity in taxonomy The two branches of fern and pine are very small, the majority of which is still the orchid branch with a rate of 99.26%. 3.1.1.4. Diversity of species 18 Similar to the distribution in families and genera, 346 species of medicinal plants were identified in the study area, concentrated in orchid, accounting for 99.40% of total species recorded. Regarding the fern and the pine branches, there is only 1 species in each branch. The results also show that the number of medicinal plants is unevenly distributed among families. There are many familes that have only one species of medicinal plants, three families have over 15 species and five families have 10 to 15 species. Therefore, we can confirm that the number of species used as medicine in Thai Thuy, Tien Hai districts of Thai Binh province mainly in the orchid branch. 3.1.1.5. Diversity of life forms Regarding the life forms, it showed that the group of Phanerophytes (Ph) was the highest, superior to other groups. Followed by a group of 1-year Therophytes (Th), Hemicrytophytes (Hm) and the least is Crytophytes (Cr). Medicinal species in the coastal districts of Thai Binh province do not have any species belonging to the Cherophytes group (Ch). In the Phanerophytes (Ph), Lianas (Li) had the highest proportion (18.78% of total species), followed by the Nanophanerophytes (Na) (17.91% of total species), Microphanerophytes (Mi) (6.07% of total species), Megaphanerophytes (Mg) (5.49% total number of species, herbaceous plants (Hp) (3.77% of total species). Succulent plants (1.44% of total species) and the least is parasitic or parasitic plants (Pp) (0.58% of total species). 3.1.1.6. Diversification of parts used as drugs * Diversity in the frequency uses of different parts of medicinal plants 19 Research shows that leaves are the most used part with 253 species, accounting for 73.12%; the second is root with 123 species, accounting for 35.48%; the third is the fruit with 120 species, accounting for 34.68%; the fourth is the seed with 104 species, accounting for 30.05%; the fifth is all parts of the plants with 89 species, accounting for 25.72% ... the lowest used is resin with 40 species, accounting for 11.56%. 3.1.1.7. Diversify in the number of plant parts used for medicine The highest proportion of plants using total parts for making drugs accounts for 146 species (42.20%); followed by plants using 01 part with 77 species, accounting for 22.20%; The third one is plants using 02 parts with 56 species, accounting for 16.20%; the fourth one is plants using 03 parts with 40 species, accounting for 11.60%, the lowest is plants using 4 parts with 27 species, accounting for 7.80%. 3.1.2. Distribution of medicinal plants The distribution of medicinal plants in two coastal districts of Thai Binh province is in the residential area with 170 species, accounting for 49.13% and growing wild in the fields, swamps, along the roads, and along the coasts with 176 species, accounting for 50.87%. 3.1.3. Potentials to cure diseases of medicinal plants (28 disease groups) The number of species that have the potential to cure gastrointestinal diseases accounts for the most species (181 species), followed by dermatological diseases (169 species), cough, blood cough (142 species), detoxification (101 species), gynecological diseases (95 species), rheumatism treatment (71 species), bite treatment (61 species), bone disorders, trauma (53 species), kidney disease treatment (46 species) (37 species), healing of eyes (species), healing 20 of hepatitis (35 species), sedative (34 species), cardiovascular treatment, blood pressure (33 species), cure diseases caused by worms, insects (31 species), cure burns (20 species), cure cirrhosis (18 species), cure asthma (17 species), cure TB (12 species), cure diabetes (12 species) malnutrition (9 species), hemorrhoids treatment (9 species), cancer treatment (8 species) (6 species), polio treatment (4 species), healing encephalitis (2 species) and treatment of obesity (2 species). 3.1.4. Some species have new uses There are 16 species with new uses that documents have not mentioned. There are two species of medicinal plants that have been identified as containing strong oxidizing and anti-cancer compounds (Maclura Cochinchinensis, Physalis angulata). 3.1.5. The rare and precious plant species listed in the Vietnam Red Book (2007) and the IUCN Red List (2014) in two coastal districts of Thai Binh province Only three species of the Least Concern category (including taxa are not considered to be conservationally endangered or nearly- threatened) according to IUCN (2014) and none of the species listed in the Vietnam Red Book (2007). 3.2. The research, exploitation and utilization of medicinal plant resources of people in two coastal districts of Thai Binh province 3.2.1. Exploitation situation of medicinal plants in the study area The survey results in Thai Thanh, Thai Do, Thai Thuong, Thuy Lien communes (Thai Thuy district), Tay Giang, Nam Phu, Nam Thanh, Dong Minh communes (Tien Hai district) showed that most people use medicinal plants for medical treatment. According to statistics of research on the use of medicinal plants in two coastal districts of 21 Thai Binh, nearly 50% of medicinal plants are collected from the wild. Of the total of 346 species of medicinal plants here, 13 species are exploited with high frequency. That's not a big number but looking at the table above we can see that the harvesting habit here is not sustainable (they take the whole plants). This situation may lead to the risk of decline of some medicinal plants. 3.2.2. Exploitation and processing of medicinal plants of the people in the study area The drug processing tools are very simple, people usually use only knife, then chop and dry or soak the plant parts. Such processing and preservation is so primitive so the proportion of moldy and rotten medicinal plants is quite high. According to physicians, most of the medicinal plant parts, whether they are leaves, stalks, roots or tubers, are chopped after harvesting, then are dried depending on the different types of diseases that have different ways of drying. 3.2.3. Traditional medicine and preparation There are 22 popular remedies to cure most common diseases with easy-to-find medicinal plants in the locality. 3.3. Screening the biological activity of some valuable species according to local knowledge We have selected 20 potential medicinal plants for research, to extract and screen biological activity to search for active substances that can protect and care human health. The samples of Annona squamosa, Annona glabra, Solnum nigrum L, Solanum Procumbens Lour, Suaeda maritima (L.) Dum and Datura metel show screening results of good activity. However, most of these samples have been studied in Vietnam. When 22 combining results of the active testing with the study of the world's research, we have discovered two samples of Maclura Cochinchinensis, Physalis angulata which contain many active substances that have not yet had any specific research in Vietnam. Based on this initial research, it is possible to orient and select two species of " Maclura cochinchinensis " and " Physalis angulata" should be put into the conservation plan, giving priority for future development. 3.4. Summary of research results of Maclura Cochinchinensis and Physalis angulata 3.4..1. Research results of Maclura cochinchinensis sample * It has been isolated and determined the chemical structure of 15 compounds from the sample of Maclura cochinchinensis, ; Three new compounds are VPA4, VPA9 and VPA10. * Three compounds (VPA30, 31 and 32) were found to have potent cytotoxic activity against lung cancer cell lines A-549, Hela cervical cancer and PANC1 pancreatic cancer. * Detected two compounds (VPA6 and 13) with antioxidant activity, protecting liver cells * Liquorice extract of resin sample of Maclura cochinchinensis does not shown toxicity: LD50 is not detected 3.4.2. Research results of Physalis angulata sample * The chemical structure of 17 compounds from angulata sample has been isolated and identified; of which, a new compound found called MQ31 * MQ29 compound was found to have good cytotoxic activity for KB cell carcinoma cell lines, LU-1 lung cancer and MCF-7 breast cancer. 23 * Compounds MQ14-15, MQ18, MQ20, MQ24-26, MQ29 and MQ31 have good activity and high selectivity for HL-60 leukemia cells. Liquorice extract of resin sample of Physalis angulata does not shown toxicity: LD50 is not detected 3.5. Solutions for effective management, conservation and sustainable exploitation of medicinal plant resources in coastal districts of Thai Binh province. 3.5.1. Preserving medicinal plants Thai Binh has rich botanical resources such as: Solanum hainanense, Pandanus tectorius Sol, Premna integrifolia, Phyllanthus urinaria L, Cynara scolymus, Alisma plantago-aquatica, Adenosma glutinosum., Eleusine indica (L) Gaertn, Plantago asiatica L, Eclipta prostrata L, Styphnolobium japonicum (L.) Schott, Aloe vera L var... many of these species are wild in some localities of Thai Binh province; There are a number of species that have been noted and included in the small scale planning area of material planting production areas. Of these, a few are bought and process by traditional medicine companies and pharmaceutical companies in the province to produce into convenient products to promote health care for people such as tea-Phyllanthus Urinaria L, tea - Cynara Scolymus L, curmin... 3.5.2. Improving the effectiveness of the leadership and direction of the Party committees and local Defining clearly that the management and protection of forests is a key, urgent, regular and long-term task to contribute to mitigating natural disasters, protecting the ecological environment and responding to climate change. Creating jobs, raising incomes, 24 contributing to hunger eradication and poverty alleviation, improving the livelihoods of coastal inhabitants, ensuring national defense and security in coastal areas. Raising the role and responsibility of the heads of local agencies and organizations in forest protection and development. People's organizations are important contributors to the preservation and development of local medicinal plant resources. 3.5.3. Promoting the awareness raising activities Diversifying forms of propagation, advocacy and education of the Party's opinions and the State's law on management and development of the mangrove forest system (special-use forests, protection forests, etc.). Their role is particularly important for socio-economic development, ecological environment protection, limiting the impact of climate change, ecotourism potentials and potentials for making medicines (through mass media, timely rewarding, propaganda in agricultural extension programs, seminars, exchanges, fostering courses to improve knowledge, organizing contest competition, distributing leaflets, integrating them into cultural programs, coordinating with schools, organizing site training courses on Planting, caring and protecting forests ...). 3.5.4. Strengthening the effectiveness and efficiency of forest management, protection and development Pursuant to legal documents, policies and regulations of the Central Party and Thai Binh province for making rational proposals for planned planted herbal planting areas; in order to continue well implementing the existing projects from 1990 to now to contribute to the restoration and development of mangroves in Thai Thuy district and Tien Hai district, Thai Binh province should soon confirm the buffer zone of the Biosphere Reserve (in accordance with the 25 Biosphere Reserve management strategy up to 2020) to create a safety belt, to enhance the conservation ef