Microscopic Identification of Zhuang Medicine Laportea bulbifera (Sieb.et Zucc.) Wedd.

Haicheng WEN Deqiang DOU Songji WEI Caiyue LIN Xuesong WANG Zhonghua DAI Wei WEI

Abstract [Objectives]This study was conducted to investigate the microscopic identification characteristics of Laportea bulbifera （Sieb. et Zucc.） Wedd.[Methods]The characteristics of L. bulbifera leaf， stem and powder were identified by microscopy.[Results]The main microscopic identification characteristics of the tissue structures of L. bulbifera leaf and stem at different positions and L. bulbifera powder were determined.[Conclusions]The microscopic identification results are characteristic， and could serve as the identification basis of L. bulbifera. These results could provide references for the quality control of L. bulbifera.

Key words Zhuang medicine; Laportea bulbifera （Sieb.et Zucc.） Wedd.; Microscopy; Identification

Zhuang medicine has a long history， with distinct national character， territoriality and traditional feature. Zhuang ancestors accumulated a wealth of experience on the use of Zhuang medicine， and a great deal of folk prescriptions， compound prescriptions， empirical prescriptions and secret prescriptions are widely used in the Zhuangsinhabited areas. Laportea bulbifera （Sieb. et Zucc.） Wedd. is generally used as a kind of medicinal herb， and widely used by Zhuang medicine in clinic as well. L. bulbifera is also known as Mafengcao in Fuchuan County of Guangxi Province[1]. The whole plant and root both could be used as a medicine， which has the effects of dispelling wind and eliminating dampness， invigorating the circulation of blood， and inducing diuresis and dissolving calculi， and is mainly used for treating malnutritional stagnation[2]， urolithiasis， rheumatism， limb numbness， injuries from falls， fractures， contusions and strains traumatic injury， unknown pyogenic infections， and infantile fever and cough[3]. L. bulbifera is widely distributed in Guangxi Province， more abundant in Napo， Debao and Fuchuan. Currently， few studies have been conducted on the microscopic identification of L. bulbifera. In this study， L. bulbifera stem， leaf and powder were identified by microscopy， providing reference and foundation for further development and utilization of the medicinal value of L. bulbifera and the study on its quality standards.

Materials and Methods

Experimental material

L. bulbifera was collected from Luomeng Village， Jinxiu Town， Jinxiu County. The material was identified by professor Wei from Guangxi Medical University as the stem and leaf of perennial herb L. bulbifera （Sieb. et Zucc.） Wedd. in Urticaceae family.

Instruments and reagents

Following instruments were used： DF20 table type continuous feeding pulverizer （Wenling Linda Machinery Co.， Ltd.）， HM355S type paraffin slicing machine （Leica， Germany）， Nikon Eclipse 80i type optical microscope （Nikon， Japan）， and DM2500M biological microscope （Leica Instruments Co.， Ltd.）. All the reagents were analytically pure.

Experimental methods

Fresh L. bulbifera stems and leaves were collected and prepared by paraffin section method， and prepared into a permanent slide through double staining. The transection was observed with an optical microscope[4-5]. In order to prepare a powder slide， the stem and leaf sample was dried， pulverized and sieved with a 60mesh sieve， and the slide was prepared with dilute glycerol. The slides were observed under a DM2500M biological microscope， and photographed with an imaging system.

Results and Analysis

L. bulbifera stem transection

The stem is quasicircular， with five vertical ridges. There is a column of epidermic cells， and those at the ridges have obviouslythickened cell wall. Nonglandular hair could be observed. There are several columns of collenchyma cells at the inner side of epidermis， which are arranged closely. The epidermis is narrow， and includes several columns of thinwall cells. The vascular bundles are nonuniform in size. The rays are wide， and include 2-10 columns of cells. Secretory cells are often interspersed at the connection part of rays and the cortex. Broad medulla is located in the center. The structure is shown in Fig. 1.

L. bulbifera leaf transection

Each of the upper and lower epidermis includes one column of cells. The upper epidermal cells are square or oval， large in size; and unicellular nonglandular hair could be observed， with obviously expanded base part. There is one column of palisade tissue cells; and the spongy tissue cells are in a loose arrangement. There are several columns of collenchyma cells at the inner side of upper and lower epidermis of midrib; and the upper epidermis of midrib has much nonglandular hair. The vascular bundles of midrib are collateral， hemispherical， and the phloem is narrow. The structure is shown in Fig. 2.

L. bulbifera powder

The powder is brown. There are many starch grains in the shape of an umbilical point， with a diameter of 5-40 m. They show obvious laminated striation， and generally aggregate or exist in thinwall cells. Halfcompound starch grains could be observed. The nonglandular hair is unicellular， warty with expanded base part， has a length of 150-180 m and contains yellow substances. The fibers form bundles， and have a large linear cavity， and pits could be observed obviously. The fibers may also be interspersed in a single form. The ducts are spiral vessels， bordered pit vessels and scalariform vessels， with a diameter of 15-100 m. The secretory cells are quasicircular， with a diameter of 110-180 m. The epidermic cells are in the shape of an elongated spindle， have slightlythickened wall and show a mosaic arrangement.

Discussion

It could be seen from above results that the leaf and stem of L. bulbifera have obvious transection characteristics. It could be known from the microscopic identification of L. bulbifera powder that the epidermic cells are in the shape of an elongated spindle; there are many starch grains， and halfcompound starch grains could also be observed; the nonglandular hair is unicellular， warty with expanded base part; the ducts have different shapes. The leaf transection also shows unicellular nonglandular hair with obviouslyexpanded base part. The stem transection shows narrow cortex，of the medicinal plants in Lindernia had obvious "genus" characteristics， and all had a snakedshaped peak nearby.

Molecular identification

Cao[18]established an NJ tree of Lindernia plants in Sichuan Province by DNA barcode technology， and the results were basically consistent with those of classical morphological classification.

Chemical Components

Iridoid glycosides： Lin[10]preliminarily judged that L. crustacea and L. antipoda both contained iridoid glycosides.

Sterols： Lan[19]isolated five crystals from the whole grass of L. ciliate and identified three of them assitosterol， stigmasterol and lupeol by IR， UV， Ms and NMR spectroscopy.

Organic acids： Cao[20]detected seven medicinal plants in Lindernia All. by HPLC to contain oleanolic acid and chlorogenic acid.

Phenylethanoid glycosides： Zhang[21]isolated such five caffeoyl phenylethanoid glycosides as plantainoside A， linderruelliosides A， leucosceptoside A， linderruellioside B and desrhamnosylverbascoside from L. ruellioides.

Phenylpropanoids： Jiang[22]from Liuzhou Institute for Food and Drug Control in Guangxi Province detected verbascoside in Yao medicine L. ruellioides by TLC and HPLC.

Pharmacological Effects

Zhang[21]evaluated and compared the invitro antioxidant capacities of five caffeoyl phenylethanoid glycosides isolated from L. ruellioides by ·OH， O-2· and DPPH· scavenging experiments， and the results showed that the caffeoyl phenylethanoid glycosides isolated from L. ruellioides all had higher antioxidant capacities against ·OH and DPPH· and certain antioxidant capacity against O-2·， and within certain ranges， with the concentrations of the compounds increasing， their antioxidant activity increased， showing certain doseeffect relationships. Pan[23]established an A549 lung cancer mice transplantable tumor model and studied the effects of extractions of L. procumbens （Krock.） Phileox on A549 lung cancer mice transplantable tumor. The results showed that the extractions of L. procumbens remarkably inhibited the growth of A549 lung cancer， showing a tumor inhibition rate of 40%. Compared with the negative group， there was no effect on the body weight of A549 lung cancer BALB/cnu mice. Liu[24]established a transplanted tumor model， and observed the effects of L. procumbens extractions and its complex extractions on H22 liver cancer ascites lump and Lewis lung cancer and on C57BL/6 mice organ index such as spleen and thymus. The results showed that L. procumbens extractions and its complex extractions had the tendency to prolong the lifecycle of mice bore H22， and also inhibited C57BL/6 mice Lewis lung cancer， and the inhibition rate was over 30%. Compared with the negative control， no statistical difference was found on C57BL/6 mice organ index such as spleen and thymus. Pan[25]established S180 tumor carneus and EL4 lymphoma transplanted tumor models to experimentally study L. procumbens and its complex extractions on transplantable tumor. The results showed that L. procumbens in all dose and its complex extractions in high and moderate dose level could inhibit the growth of S180 tumor， and the inhibition rate was over 40.0% （P<0.05）. The tumor inhibition rate was 21.5% （P>0.05） in low dose. The inhibition rates of L. procumbens and its complex extractions on EL4 lymphoma was over 30.0%. No statistical difference on organ index such as spleen and thymus was found between the extractions and the negative control. L. procumbens and its complex extractions could significantly inhibit the growth of S180 and EL4 with no dosedependence.

Clinical Application

Tan[26]soaked 1 kg of L. ciliatain in 4.5 kg of 38° Guilin flower wine， and drank 30-50 ml per night for 113 d， which cured his leg and foot joint pain. Lan[27]from the Health Center of Chengguan Commune in Yunxiao County， Fujian Province treated light infant diarrhea （simple dyspepsia） by decocting the fresh herb （50-100 g） in water， giving the decoction which was taken twice a day， one dose per day. In the case of severe infant diarrhea （toxic dyspepsia）， 100 g of fresh herb was decocted， the decoction was taken two doses daily， and the disease could be cured after three days of treatment. Liang[6]repeatedly interviewed the Yao people in Jinxiu County， and "Jinxiu County Folk Medicine Status Survey" summarized the clinical manifestations and treatment of the Sha syndrome. It was found that the L. ruellioides was used for the treatment of Miaoshe Sha （Mahuang Sha）， Yangmao Sha and Chen Sha as a Yao medicine. Li[28]used L. crustacea to treat 100 cases of snake bites， and received satisfactory results.

Summary and Outlook

In Guangxi Province， there are 16 Lindernia medicinal plants， which are widely used as Zhuang and Yao national medicines in the folk， indicating that they have certain medicinal value. However， the development and utilization of Lindernia plants is still in its infancy， and few studies have been conducted on their components， pharmacological effects and clinical application. Further research is needed in the future to provide an experimental basis for the development and utilization of these medicinal plants in Lindernia All.

Fengfeng XIE et al. Advances in Research on Medicinal Plants of Lindernia All. in Guangxi Province

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Editor： Yingzhi GUANGProofreader： Xinxiu ZHU

much narrow phloem and wide rays， while the medulla is broad and located in the center; secretory cells are often interspersed at the connection part of rays and the cortex; and the identification characteristics including nonglandular hair also could be seen. These microscopic identification characteristics provide reference for the microscope identification of L. bulbifera， as well as a foundation for further research and development of the Zhuang medicine L. bulbifera and the establishment of its quality standards.

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