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Last Update: 2014-07-24
Labisia pumila (Myrinaceae), is a subherbaceous plant with creeping stems and found mainly in the lowland and hill forests of Peninsular Malaysia at an altitude between 300 and 700 m (Burkill 1966). Decoction of L. pumila has been traditionally used by generations of Malay women to induce and facilitate childbirth, as well as being a post-partum medication to help contract the birth channel, to tone the abdominal muscles, maintain a healthy female reproductive system, to treat menstrual irregularities and to regain body strength (Burkill 1966; Wan Ezumi et al. 2007). The ability to enhance the contraction of the uterus has been proven (Cheong 2007). Increasing request for high yielding material of this plant from the herbal industry serve as catalyst for researchers to establish high quality planting materials to fulfill the industrial demand.
Therefore, in order to produce high quality raw materials to support Malaysian herbal industries, it is important to select genotype which either produced the maximum bioactivity and/or contained high quality active ingredients (Ibrahim et al. 1990) before it can be domesticated. FRIM has established a germplasm of 300 genotypes for both varieties L. pumila var. pumila and L. pumila var. alata (Farah Fazwa et al., 2010, 2012) with high total phenolic content which is more related to antioxidant activities. In order to produce high quality planting materials with traditional claim of L. pumila, a study will be conducted on the ability of the species to enhance contraction of the uterus. Furthermore, literature and patent search indicated that production of high yielding accessions based on uterus contraction activity has not been investigated and registered cultivar of L. pumila is unavailable.
In this study, aqueous extraction is one of the methods will be use for the extraction of the fresh leaves of the herbs. This method offers several advantages over conventional solvent extraction-less initial capital investments, safer operation, capability of discontinuous operation, and production of a variety of products. Another advantage of aqueous processing is the capability for utilization of certain chemicals to remove or inactivate undesirable substances (Carl M. Cater, 1974). A study showed that water extracts of L.pumila were able to displace estradiol binding to antibodies raised against estradiol, making it similar to other estrogens such as estrone and estradiol. The extract has also been found to produce a dose-response effect on the reproductive hormones of female rats, notably on the estradiol and free testosterone level.
For the planting part, the planting stocks are usually raised from seeds. The dependence on seeds as source of planting materials is inadequate as seeds are difficult to obtain due to depletion of mother plants. Propagation by seeds alone will not be able to meet the large number of planting stock required. Propagation technique by cuttings can be one of the alternative methods used (Hartman & Kester 1983). This method of propagation also offers useful tool for propagating the superior genotypes which will help to maximise quality and productivity. Rozihawati (2008) has reported the propagation techniques by cuttings, factors affecting rooting and nursery techniques of this species. Aminah et al. (2009) also reported the suitability of using cutting to propagate L. pumila using different leaf parts and areas. Research on propagating superior accessions of L. pumila based on total phenolic content has also been carried out (Farah Fazwa et al. 2013).
As a conclusion, this study will be carried out to to select high yielding accessions of L. pumila from established germplasm based on uterus contraction activity for breeding purposes; to propagate the selected high yielding accessions using vegetative techniques and to establish clonal bank from the selected high yielding accessions of L. pumila. It is expected that at the end of the study high yielding planting materials of L. pumila will be made available and transfer of technology conducted. These outputs are important to ensure sustainable supply of high yielding planting materials for the production of high quality materials to meet future industry demand.
1.1 Problem Statement
Phytochemical extraction is important for the selection of high yielding accessions of Labisia pumila. Literature and patent search indicate the selection of high yielding accessions based on uterus contraction activity have never been tested before.
1.2 Objective (s);
I. To select high yielding accessions of L. pumila from established germplasm based on uterus contraction activity for breeding purposes.
II. To propagate the selected high yielding accessions by vegetative techniques.
III. To establish clonal bank from the selected high yielding accession of L. pumila
2.0 LITERATURE REVIEW
Labisia pumila, commonly known as Kacip Fatimah in Malaysia, is a small herbaceous and slightly woody plant (Karimi et al., 2011). The habitat of this plant is in lowland primary forests at shady places or in secondary forests on humus-rich soils (Sunarno, 2005). There are three varieties of Kacip Fatimah had been recognized namely var. alata, var. pumila and var. lanceolata and each variety have its own use and function (Stone, 1988). Stone (1988) also identified that these varieties can be distinguished from each other by their petiole and leaf characteristic.
Traditionally, Labisia pumila have been widely used as medicinal plant especially related to women health care. It is locally known to be used for induce and facilitate childbirth as well as a post partum medication to help contract the birth channel, to tone the abdominal muscles and to regain body strength by traditional uses (Wan Ezumi et al., 2007). Other traditional uses include treatment for dysentery, flatulence, rheumatism, dysmenorrheal and gonorrhea (Mohamad Norhaiza et al., 2009; Burkil, 1996). Recently, it was reported that the bioactive compounds of L. pumila consisted mainly resorcinols, flavonoids and phenolic acids (Ibrahim, 2010). Polyphenolic contain in L. pumila have the ability to prevent cancer and heart disease (Tsao & Deng, 2004).
The main objective of this study is to select the high yielding of Labisia pumila based on contraction activity. The uterine contractility activity was carried out using procedure by Ayinde et.al. Estrus in the uterus of female Sprague Dawley rats will be induced using diethylstilbestrol (i.p.). The effects of L. pumila on the uterus contraction activity will be measured using Powerlab system (ADIntruments Pty. Ltd.) and compared to oxytocin as the positive control drug. The high yielding accessions of L. pumila will be selected based on of the highest contraction activity.
The selected L.pumila will undergo propagation using vegetative techniques to ensure sustainable supply of high quality planting material which benefit the producer as well as the consumer.
Objective 1: To select high yielding accessions of L. pumila from established germplasm based on uterus contraction activity for breeding purposes.
a) Phytochemical Extraction and Chemical fingerprinting of L. pumila aqueous extract
Plant leaf will be cleaned and leave to dry in at oven at controlled temperature until reached desired moisture content. 300 g dried and ground leaf will be extracted by refluxing using hot distilled water. The filtrate is then concentrated and dried using freeze-dryer. The crude obtained will be divided into two analysis, bioactivity and chemical fingerprint analysis (UV, HPLC and HPTLC). While, the FTIR fingerprint will be carried out on the dried and ground leaves. PCA analysis also will be carried out.
b) Uterus contraction activity of L. pumila
The uterine contractility activity will be carried out using procedure by Ayinde et al. 2006. Estrus in the uterus of female Sprague Dawley rats will be induced using diethylstilbestrol (i.p.). The effects of L. pumila on the uterus contraction activity will be measured using Powerlab system (ADIntruments Pty. Ltd.) and compared to oxytocin as the positive control drug. The high yielding accessions of L. pumila will be selected based on of the highest contraction activity. This bioactivity facility will be established in FRIM.
Objective 2: To propagate the selected high yielding accessions by vegetative techniques.
a) Multiplication of high yielding accession by vegetative technique
Multiplication of high yielding accessions of L. pumila will be done by cuttings. This method will be used to produce the high yielding materials of L. pumila. From each of selected accessions, 60 ramets will be further produced. These clones will be raised and maintained at FRIM’s nursery for six to eight months before they can be transplanted to field. The survival rate and growth performance (e.g height, leaf number, biomass) of these rooted plants will also be monitored and assessed during this stage
Objective 3: To establish clonal bank from the selected high yielding accession of L. pumila
a) Establishment of clonal bank plot
The clonal bank plot will be laid out in a Completely Randomized Block Design (RCBD). These selected clones will be arranged per replicate in three experimental blocks. For each replicate, the size of planting bed will be 1 meter x 2 meter. This experimental design will be applied to both varieties.
Upon completion of this project:
1. Phytochemical Extraction and Chemical fingerprinting of L. pumila aqueous extract and also test for uterus contraction activity of L. pumila on rats
2. Multiplication of high yielding accession by vegetative technique
3. Establishment of clonal bank plot from the selected high yielding accession of L.pumila
Last Update: 2014-07-22
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