Research article Application of morphoanatomy and microscopy in authentication of three species of traditional Chinese herbs of Moghania Yan-fei Huang a, c , Chao-qin Ren b, c , Wei Yuan a, c , Yuan Liu a, * , Zhi-feng Zhang a , Lu-yang Lv a a Ethnic Medicine Institute, Southwest University for Nationalities, No. 16, South 4th Section, 1st Ring Road, Chengdu, Sichuan 610041, PR China b Department of Biology and Chemical, Aba Teachers College, ShuimoTown, Wenchuan County, Sichuan 623001, PR China article info Article history: Received 29 July 2012 Accepted 2 September 2012 Available online 11 January 2013 Keywords: Moghania Microscopy authentication Morphoanatomy Traditional Chinese herbs abstract Background: Moghania philippinensis (Merr. et Rolfe) Li, M. macrophylla (Willd.) O. Kuntze and M. ferruginea (Wall. ex Benth.) Li. (Leguminosae) has not the national standards now, and they are just only recorded in appendix of “Pharmacopoeia of the People’s Republic of China” (2010 and 2005 edition). But their roots are the crude herbs recourses of the primary ingredient in the famous prescription of “Female Moghania Tablet” for curing colpitis in China. Moreover M. philippinensis was also recorded by the appendix “Pharmacopoeia of the People’s Republic of China” (1977 edition). M. strobilifera (Linn.) Ait, M. glutiaosa (Prain) Y.T.Wei, and M. Latifolia Benth are still medical used in some local region of Guangxi and Yunnan Province of China. So their crude herbs recourses are confused in Chinese Crude Herbs Market. Objective: To distinguish three species of Moghania and ensure their safety and efficacy. Materials and methods: Three species of M. macrophylla (Willd.) O.Kuntze, M. Latifolia Benth and M. glutiaosa (Prain) Y.T.Wei from Guangxi, Yunnan and Sichuan Province of China (SWUN) (Table 1). Fresh materials were fixed in FAA. Samples were passed through the traditional ethanol and dime- thylbenzene series, embedded in paraffin, and then sliced with a microtome and stained with safranine- fast green and finally mounted. The leaf lower epidermal cells are obtained by the practical peeling technique and clearing method with nail polish blot. The materials were dissociated by nitric acid chromate method. The samples of herbs were powdered. The average number of palisade cell under the epidermal cells is called Palisade Ratio (PR). Since palisade ratio is consistent and different from plant to plant, this parameter can be used in authentication. Sto- matal Index (SI) is a basilic parameter in authentication, which can be obtained from the following formula: Stomatal Index ¼ (number of stomata/every millimeter-square) 100/(number of stomata/ every millimeter-square þ number of epidermal cell/every millimeter-square). Mesophyll tissue is divided up by slendest vein, which is called vein islet. The numbers of vein islet in every epidermal cell is called Vein Islet Numbers (VIN), which also can be used as a reliable parameter in authentication since it is consistent in a species of plant. The results were studied using light microscope according to the usual microscopic techniques. Results: The three species of Moghania were studied to compare the distinguishing morphoanatomic details of root, stem, leaf and petiole, dissociation and powders. The microscopic features were sys- tematically described and illustrated. And detailed key authentication parameters based on these ana- tomic characteristics were presented. A key was constructed to the three species for convenient classification. Microscopy can be unambiguously used to authenticate and distinguish three species of Moghania. Copyright Ó 2013, Phcog.Net, Published by Reed Elsevier India Pvt. Ltd. All rights reserved. 1. Introduction Premenstrual symptoms, colpitis and hyperplasia of mammary glands are three of the common female gynecologic diseases. Among them, the colpitis is the multiple and difficult gynecologic diseases, bringing chronic pains of women. * Corresponding author. Tel.: þ86 28 8552 2322, þ86 1380 8091 609. E-mail addresses: [email protected](Y.-f. Huang), [email protected](C.-q. Ren), [email protected](W. Yuan), [email protected](Y. Liu), [email protected](Z.-f. Zhang), [email protected](L.-y. Lv). c Contribution equally/sharing first authorship. Contents lists available at SciVerse ScienceDirect Pharmacognosy Journal journal homepage: www.elsevier.com/locate/phcgj 0975-3575/$ e see front matter Copyright Ó 2013, Phcog.Net, Published by Reed Elsevier India Pvt. Ltd. All rights reserved. http://dx.doi.org/10.1016/j.phcgj.2012.12.003 Pharmacognosy Journal 5 (2013) 30e40
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Research article
Application of morphoanatomy and microscopy in authentication ofthree species of traditional Chinese herbs of Moghania
Yan-fei Huang a,c, Chao-qin Ren b,c, Wei Yuan a,c, Yuan Liu a,*, Zhi-feng Zhang a, Lu-yang Lv a
a Ethnic Medicine Institute, Southwest University for Nationalities, No. 16, South 4th Section, 1st Ring Road, Chengdu, Sichuan 610041, PR ChinabDepartment of Biology and Chemical, Aba Teachers College, Shuimo Town, Wenchuan County, Sichuan 623001, PR China
a r t i c l e i n f o
Article history:Received 29 July 2012Accepted 2 September 2012Available online 11 January 2013
Keywords:MoghaniaMicroscopy authenticationMorphoanatomyTraditional Chinese herbs
0975-3575/$ e see front matter Copyright � 2013, Phhttp://dx.doi.org/10.1016/j.phcgj.2012.12.003
a b s t r a c t
Background: Moghania philippinensis (Merr. et Rolfe) Li, M. macrophylla (Willd.) O. Kuntze andM. ferruginea (Wall. ex Benth.) Li. (Leguminosae) has not the national standards now, and they are justonly recorded in appendix of “Pharmacopoeia of the People’s Republic of China” (2010 and 2005 edition).But their roots are the crude herbs recourses of the primary ingredient in the famous prescription of“Female Moghania Tablet” for curing colpitis in China. Moreover M. philippinensis was also recorded bythe appendix “Pharmacopoeia of the People’s Republic of China” (1977 edition). M. strobilifera (Linn.) Ait,M. glutiaosa (Prain) Y.T.Wei, and M. Latifolia Benth are still medical used in some local region of Guangxiand Yunnan Province of China. So their crude herbs recourses are confused in Chinese Crude HerbsMarket.Objective: To distinguish three species of Moghania and ensure their safety and efficacy.Materials and methods: Three species of M. macrophylla (Willd.) O.Kuntze, M. Latifolia Benth andM. glutiaosa (Prain) Y.T.Wei from Guangxi, Yunnan and Sichuan Province of China (SWUN) (Table 1).Fresh materials were fixed in FAA. Samples were passed through the traditional ethanol and dime-thylbenzene series, embedded in paraffin, and then sliced with a microtome and stained with safranine-fast green and finally mounted. The leaf lower epidermal cells are obtained by the practical peelingtechnique and clearing method with nail polish blot. The materials were dissociated by nitric acidchromate method. The samples of herbs were powdered.The average number of palisade cell under the epidermal cells is called Palisade Ratio (PR). Since palisaderatio is consistent and different from plant to plant, this parameter can be used in authentication. Sto-matal Index (SI) is a basilic parameter in authentication, which can be obtained from the followingformula: Stomatal Index ¼ (number of stomata/every millimeter-square) � 100/(number of stomata/every millimeter-square þ number of epidermal cell/every millimeter-square).Mesophyll tissue is divided up by slendest vein, which is called vein islet. The numbers of vein islet inevery epidermal cell is called Vein Islet Numbers (VIN), which also can be used as a reliable parameter inauthentication since it is consistent in a species of plant. The results were studied using light microscopeaccording to the usual microscopic techniques.Results: The three species of Moghania were studied to compare the distinguishing morphoanatomicdetails of root, stem, leaf and petiole, dissociation and powders. The microscopic features were sys-tematically described and illustrated. And detailed key authentication parameters based on these ana-tomic characteristics were presented. A key was constructed to the three species for convenientclassification. Microscopy can be unambiguously used to authenticate and distinguish three species ofMoghania.
Copyright � 2013, Phcog.Net, Published by Reed Elsevier India Pvt. Ltd. All rights reserved.
Premenstrual symptoms, colpitis and hyperplasia of mammaryglands are three of the common female gynecologic diseases.Among them, the colpitis is the multiple and difficult gynecologicdiseases, bringing chronic pains of women.
Table 1Source of materials of the three species of M.
S.no
Taxon Batches Locality Elevation(m)
Date ofcollection
Voucher
1 M. macrophylla(Willd.) O. Kuntze
No.1 Sichuan,China
500 April,2007
Y. Liu 07041001(SWUN)
No.2 Sichuan,China
500 April,2007
Y. Liu 07041002(SWUN)
No.3 Yunnan,China
551 May,2007
CZ. Peng07051003(SWUN)
2 M. latifolia Benth No.1 Yunnan,China
551 April,2008
CZ. Peng08041001(SWUN)
No.2 Yunnan,China
551 April,2008
CZ. Peng08041002(SWUN)
No.3 Yunnan,China
551 April,2008
CZ. Peng08041003(SWUN)
3 M. glutiaosa(Prain) Y.T.Wei
No.1 Guangxi,China
570 April,2008
B. Dai08041001(SWUN)
No.2 Guangxi,China
570 April,2008
B. Dai08041002(SWUN)
No.3 Guangxi,China
570 April,2008
B. Dai08041003(SWUN)
Table 2Comparison of morphological characters of the three species of the whole plant of M.
Taxon M. macrophylla (Willd.) O. Kuntze
The whole plant About 0.5e3 m, erect, shrub
Young stem Columniform bvious ridge, anddensely covered soft hair.
Dried root Grey or brown-red colored, andinfarctate and heavy
Ternated compound Texture and hair Papery or thin leathery, covered nhair except main vein of lowersurface, and lower surface withbrown and black glandular dots.
Form and size of blade 8e15 � 4e7 cm, the lower side isnot white-grey color.
Nowadays "Female Moghania Tablet" has the good clinic effectfor curing colpitis, and it is the good promised sales volume in thetraditional Chinese medicinal market. Moreover, it was the firstchoice for women to curing colpitis. The dried roots of Moghania isthe primary ingredients in the prescription of "Female MoghaniaTablet", which belongs to Leguminosae of Moghania philippinensis(Merr. et Rolfe) Li, M. macrophylla (Willd.) O. Kuntze andM. ferruginea (Wall. ex Benth.) Li. They were recorded in appendixof “Pharmacopoeia of the People’s Republic of China” (20051 and20102 edition). And M. philippinensis was also recorded by the ap-pendix “Pharmacopoeia of the People’s Republic of China” (1977edition). However, the "Pharmacopoeia of the People’s Republic ofChina" (2010 edition and earlier) has not accepted the qualitystandard of them.
China is the native producing region of 6 species ofMoghania and1 varieties, and 6 species of them are popularly used in traditionalChinese herbs in southwest of China.3 M. philippinensis (Merr. etRolfe) Li and M. macrophylla (Willd.) O. Kuntze are widely dis-tributed in southwest of China, such as Yunnan, Guangxi, Guang-dong, Jiangxi, Fujian, Taiwan, Hubei, Hunan, Sichuan Province ofChina and so on, especially in Guangxi and Yunnan Province ofChina. Their dried roots have the good medicinal functions of alle-viating the pain ofwind-dampness, lumbago, arthralgia, and lumbarmuscle strain, gynecopathy, breast diseases, impotence and seminalemission and soon.4 And theyare also used byHani, Dai, Zhuang andYao Nationality. But their wild resources and cultivated resources
M. latifolia Benth M. glutiaosa (Prain) Y.T.Wei
About 1e2 m, erect, shrub About 0.5e1 m, erect, suffrutescent,ramose
Triangular prism, and denselycovered ferrugineous soft hair.
Columniform, and densely coveredgolden base-expanded glandularhair and grey hair.
Brown red colored, not infarctateand heavy easy to smashed
Brown-red colored
o Papery or thick papery, coveredshort soft hair, and lower surfacewith brown and black glandulardots.
Papery or thick papery, coveredshort soft hair, and lower surfacewith brown and red glandular dots.
8e14 or more longer � 4e6 cm orbroader, the lower side is white-grey color.
4e9 � 1.5e3 cm, young leaf purplecolored; the lower side is white-grey color.
3e10 cm 1.5e4 cm2e6 mm 2e6 mm
y
Single to three, axillary oracrogenous, 3e11 cm;inflorescence axis, hypsophyll andpedicel covered ferrugineous softhair.
Single or several, acrogenous oraxillary, 1.5e5 cm; inflorescenceaxis, hypsophyll and pedicelcovered golden base-expandedglandular hair and grey hair.
Almost no 1e10 cmAlmost no Almost noPetal is red-purple or pink witha longer calyx. Androecium isdiadelphous. Ovary is ellipticcovered soft hair with thin style.
Petal is yellow as long as calyx.Androecium is diadelphous. Ovaryis elliptic.
dPod is elliptic with 1.2e1.6 cm longand 7e8 mm wide, coveredferrugineous short soft hair. Theapex has a small acuate beak.
Pod is elliptic with 1e1.4 cm longand 0.5e0.7 cm wide, covered lightgolden base-expanded glandularhair.
1e2, globose and black, about 2mm 1e2, almost globose and brown-black, about 2 mm
The whole year From Feb to MayThe whole year From Feb to May560e2100 m About 1400 mMainly distributed in Yunan andGuangxi Province of China.
Mainly distributed in Dayao,Jinghong and Mengna county ofYunan Province and Longzhoucounty of Guangxi Province ofChina.
Y.-f. Huang et al. / Pharmacognosy Journal 5 (2013) 30e4032
are not enough to satisfy the sustainable needs of traditional Chi-nese medicinal market for making of "Female Moghania Tablet".
In addition to that of them, M. strobilifera (Linn.) Ait, M. latifoliaBenth andM. glutiaosa (Prain) Y.T.Wei are still medical used in somelocal region in southwest of China. And they are also regarded ashaving the same medicinal function as that of them. They arepopularly medical used by Hani, Dai, Zhuang Yao and Dai Nation-ality of China.3 To ensure their effective usage, the most urgentrequirement is to guarantee authenticity. A few studies have beencarried out for this purpose. However, systematic information ofmicroscopic authentication and morphoanatomy has not beenreported for the three species in this area. In order to distinguishthe three species and find out whether the local medical usedspecies of M. latifolia Benth and M. glutiaosa (Prain) Y.T.Wei have
Fig. 1. Photos of plants of the three species of M. A: The whole plant. B: Ph
any other similarities withM. macrophylla (Willd.) O. Kuntze or not,which is recorded in appendix of “Pharmacopoeia of the People’sRepublic of China” (20051 and 20102). Microscopic authenticationwas adopted, since it is a facile, inexpensive, and objective methodwhich has been successfully applied in “Pharmacopoeia of thePeople’s Republic of China”,2 the American Herbal Pharmacopoeia(Upton, 2003), and the Japanese Pharmacopoeia (Society of Japa-nese Pharmacopoeia, 2001). Microscopy technique was also fre-quently reported in the literature for authentication.5e7
A series of related studies of M. has been conducted by ourresearch group.8e11 In order to get fresh material of M., we madeon-site investigations in Guangxi, Yunnan and Sichuan Province ofChina and collected both fresh wild and cultivated plants ofM.macrophylla (Willd.) O. Kuntze,M. latifolia Benth andM. glutiaosa
otos of stem. C: The upper surface of leaf. D: The lower surface of leaf.
(Prain) Y.T.Wei. The native people told us that the local medicalused species ofM.was regarded as lower quality than the species ofrecorded in appendix of “Pharmacopoeia of the People’s Republic ofChina” (2010 and 2005 edition), and cannot be purchased in thetraditional Chinese crude native herbs market.
For the sake of offering a scientific data for further research toensure their safety and efficacy, in the present work, a systematicand detailed pharmacognostical study of three species of M. nativeto Guangxi, Yunnan and Sichuan province of China was undertakento establish a method which would facilitate authentication ofthese species. Subsequently, representative and unique micro-scopic features were characterized and recorded by digital colorphotography.
2. Materials and methods
2.1. Materials
Three species ofM.were collected during flowering and fruitingtime from Guangxi, Sichuan and Yunnan province of China, andauthenticated by Prof. Yuan Liu (Ethnic Medicine Institute, South-west University for Nationalities, Chengdu, PR. China), Bin Dai(Ethnic Medicine Institute of Guangxi, Nanning, PR. China) andChaozhong Peng (Yunna Branch, Institute of Medicinal PlantDevelopment). Voucher specimens were deposited in the Herbar-ium of Ethno-herbs, Ethnic Medicine Institute, Southwest Univer-sity for Nationalities (SWUN) (Table 1).
2.2. Apparatus
All the transverse sections of the materials were prepared usinga KD-1508microtome (Jinhua Kedi Instruments, China). An imagingsystem consisting of an optical microscope equipped witha micrometer and a digital camera for acquisition of photographs(BX41 standard laboratorymicroscope of Olympus, Japan) was usedfor photography.
2.3. Reagents
FAA (50% ethanol: formaldehyde: Glacial acetic acid ¼ 89: 5: 6)and ethanol series (in gradient of 30%e100%) were prepared forspecimen fixation and dehydration, respectively. Safranine T in50% alcohol, and safranine-fast green solution were prepared forspecimen staining. Chloral hydrate and diluted glycerin were pre-pared according to procedures described in Pharmacopoeia of thePeople’s Republic of China (The State Pharmacopoeia Committee ofChina, 2010). Dimethylbenzene, paraffinwax, and Canadian balsamused in this study were all of chemical grade.
Table 3Main comparison of transverse sections, dissociation and powders of root, stem, leaf and
Taxon M. macrophylla (Willd.)O. Kuntze
Brown-red inclusion (mm),quantity, local site
9e35; a lot of; root, stemleaf and petiole
Calcium oxalate crystal (mm),quantity, local site
5e35, a lot of; root, stem,leaf and petiole
Stone cell (mm), quantity, local site 5e50, a lot of; root, stem,leaf and petiole
Epidermal cells ofstem and leaf
Non-glandular hair HasGlandular hair in No
Stoma Type ParacyticWidth 2.40e3.03Length 16.67e19.69Stomatal index 10.8e12.1
2.4. Methods
Fresh materials of three species of M. were divided into appro-priate sizes and fixed in FAA. Samples were passed through thetraditional ethanol and dimethylbenzene series, embedded inparaffin according to the technique described by Ruzin (1999), andthen sliced with amicrotome to suitable thickness and stainedwithsafranine-fast green (Berlyn andMiksche,1976; Johansen,1940)andfinally mounted in Canada balsam for observation. Leaves of thesamples were prepared according to the practical peeling techni-que and clearing method (Berlyn and Miksche, 1976; Leng Ping-sheng, Wang Wen-he, et al 2007). The samples of three species ofM.were powdered and cleared in chloral hydrate, then mounted inglycerin. Three samples were studied for the key authenticationfeatures.
3. Results
3.1. Plant morphology
The gross morphology between three species is quite differentfrom each other. We listed the distinguishable characters in Table 2and showed them in Fig. 1.
3.2. Microscopic characteristics
Characteristic microscopic differences of three species intransverse section of root, stem, leaf and petiole; and surface viewof leaf under epidermis; and herbs dissociation and powders aresummarized in Table 3 and pictures of the three species are shownin Figs. 2e6.
3.2.1. Microscopic characteristics of transverse sections3.2.1.1. Root. M. macrophylla (Willd.) O. Kuntze: The outline isalmost circular. Phellem layer is 5e6 layers tangentially prolongedbrown cork cells, arranged closely. The outer-layer is sometimesruptured and exfoliated. The cortex consisted of 2e3 layers pa-renchyma cells, oval or subrounded, arranged loosely. Phloem andxylem fiber are bundled; the cortex and phloem are narrow, butxylem is very wide with the ratio of 5e7; cambium is not obvious;phloem and xylem rays are composed of 3e6 layers of cells; two orthree vessel bundles are closely packed together occasionally, ma-jority of them radially arranged in xylem; parenchyma cells ofcortex and phloem contained brownered inclusion about 14e16 mm in diameter, stone cell about 8e20 mm in diameter,calcium oxalate crystal about 7e13 mm in diameter, starch grainabout 50e60 mm in diameter and except a few of them existed inxylem rays.
petiole of M.
M. latifolia Benth M. glutiaosa (Prain) Y.T.Wei
, 10e35; a little; root,stem, leaf and petiole
10e30; a lot of; root, stem,leaf and petiole
5e25; a little; root, stem,leaf and petiole
5e100, a lot of; root, stem,leaf and petiole
10e45; a little; root, stem,leaf and petiole
10e35, a lot of; root, stem,leaf and petiole
Has A lot of and longer than othersNo A lot of basal expandedParacytic and infinitive Paracytic2.11e3.63 1.97e3.4213.56e21.05 13.05e18.4229.3e31.7 7.2e8.4
Fig. 2. Transverse sections of root of three species of M. ; A and B: Outline. C: Epidermis and cortex. D: Xylem and phloem. E. Brownered inclusion. F. Stone cell. 1. Cork layer;2. Cortex; 3. Xylem rays; 4. Brownered inclusion; 5. Stone cell; 6. Phloem; 7. Vessel; 8. Calcium oxalate crystal.
Y.-f. Huang et al. / Pharmacognosy Journal 5 (2013) 30e4034
The microscopic characteristics of root of the other species aresimilar toM.macrophylla (Willd.) O. Kuntze, but there are also somedifferences among them. For example, the size and density ofbrown-red inclusion, stone cell and calcium oxalate crystal aredifferent from each other.
Micrographs of detailed comparison of three species are shownin Table 3 and Fig. 2.
3.2.1.2. Stem. M. macrophylla (Willd.) O. Kuntze: The outline is fiveirregularly waved curving shaped. The epidermal cells are squareand single-layered, coveredwith thin cuticle and single or 2e3 cellsnon-glandular hair and contained the green and brown-red
Fig. 3. Transverse sections of stem of three species of M. A and B: Outline. C: Libriform fiber.Gb: Calcium oxalate crystal in polarized. I: Non-glandular hair and glandular hair. 1. EpiderBrown inclusion; 9. Stone cell; 10. Calcium oxalate crystal; 11. Non-glandular hair; 12. Glan
inclusion. Cortex is narrow and consisted of 4e5 layers parenchymacells with brown inclusion about 9e22 mm in diameter, a lot ofstone cells about 5e20 mm in diameter. The phloem and vascularbundle is narrow and parenchyma cell contained stone cell about10e25 mm in diameter, brown inclusion about 14e22 mm indiameter, brown-red inclusion about 9e35 mm in diameter, andcalcium oxalate crystal about 10e30 mm in diameter. Two or fivevessel bundles are closely packed together occasionally, majority ofthem radially arranged in xylem. Pith is very wide.
The microscopic characteristics of stem of other species aresimilar toM.macrophylla (Willd.) O. Kuntze, but there are also somedifferences among them. For example, glandular hair and the
Y.-f. Huang et al. / Pharmacognosy Journal 5 (2013) 30e4036
quantity of inclusion, stone cell and calcium oxalate crystal aredifferent from each other.
Micrographs of detailed comparison of three species are shownin Table 3 and Fig. 3.
3.2.1.3. Leaf and petiole3.2.1.3.1. Leaf. M. macrophylla (Willd.) O. Kuntze: Bifacial leaf,
both upper and lower epidermises are composed of oblong orsubrounded and single-layered cells that are arranged closely andcovered with cuticle. Upper epidermis is covered consisted of 2e3cells non-glandular hair. Stomata can be observed in lower epi-dermis. The palisade tissues are 1e2 layer, do not pass throughmidrib; spongy cells are 1e2 layer, oval, arranged loosely withbroad intercellular space and contained brown inclusion and cal-cium oxalate crystal. Vascular bundle of lateral veins can beobserved. Vascular bundles of the midrib are of the ectophloic type,half circular-shaped, and enclosed by 8e9 layer parenchyma cellsunder upper epidermis and 4e5 layer parenchyma cells underlower epidermis. The cambium is not conspicuous.
The anticlinal walls of the leaf epidermal cells are wavy. Stomatatype is paracytic, which is 2.40e3.03� 16.67e19.69 mmdispersedlydistributed, and guard cell is kidney-shaped and 2 subsidiary cellswith cuticula trace. The palisade ratio is 10.8e12.1.
Micrographs of detailed comparison of three species are shownin Table 3 and Fig. 4.
3.2.1.3.2. Leaf petiole. M.macrophylla (Willd.) O. Kuntze: Almostcircular with five irregularly waved curving shaped. Epidermis iscomposed of oblong or subrounded and single-layered cells thatare arranged closely with brown inclusion and single cell non-glandular hair. Vascular bundle is 6e7 ringed and ectophloic type.Fiber bundle is 2e4 layer and waved ringing. Cambium is notobvious. Single or 2e3 vessel bundles are closely packed togetheroccasionally, majority of them radially arranged in xylem. Pith isvery wide with redepurple inclusion in parenchyma cell. Upperepidermis of leaf wings is covered with cuticle, and lower epi-dermis of leaf wings is covered nonglandular hair. Vascular bundleof lateral vein can be observed in the cortex of leaf wings.
Micrographs of detailed comparison of three species are shownin Table 3 and Fig. 5.
3.2.2. Microscopic characteristics of dissociation and powders3.2.2.1. Root. M. macrophylla (Willd.) O. Kuntze: Light brown color.Cork cell is quadrate, about 31e52 mm in length. Vessel is mainlypitted vessel, 226e271 mm in length, 33e60 mm in width; Some-times reticulate vessel can be observed. Fiber is single or bundle,thin and long, linear cell cavity and obvious pit, 487e736 mm in
Fig. 4. Transverse sections of leaf of three species of M.; A: Blade. B: Midrib. C: Stomas. 1. Lower epidermis; 2. Upper epidermis; 3. Spongy parenchyma; 4. Palisade tissue; 5. Non-glandular hair; 6. Calcium oxalate crystal; 7. Brown inclusion; 8. Glandular hair; 9. Phloem; 10. Xylem; 11. Stomas.
length, 21e33 mm in width; parenchyma cell, which is rounded thefiber, contained calcium oxalate crystal with thick wall a little lig-nification or none lignification, called crystal fiber sheaths. Redepurple or brown inclusion can be observed. Stone cell is circleround or irregular shape with grey or brown color, a little pit about20e50 mm in diameter. Calcium oxalate crystal is about 20e35 mmin diameter. Starch grains are observed occasionally.
Micrographs of detailed comparison of three species are shownin Table 3 and Fig. 6A.
3.2.2.2. Stem. M. macrophylla (Willd.) O. Kuntze: Grass green color.Cork cell is quadrate, about 20e53 mm in length. Vessel is mainlypitted vessel, 210e230 mm in length, 50e65 mm in width; Some-times reticulate vessel can be observed. Fiber is single or bundle,thin and long, linear cell cavity and obvious pit, 150e360 mm inlength, 10e15 mm in width; parenchyma cell, which is rounded thefiber, contained calcium oxalate crystal with thick wall a little lig-nification or none lignification, called crystal fiber sheaths. Brown-red or brown inclusion can be observed. Stone cell is circle round orirregular shape with grey or brown color, a little pit about 20e40 mm in diameter. Calcium oxalate crystal is white-grey colorand about 10e35 mm in diameter. Starch grains are observedoccasionally.
Micrographs of detailed comparison of three species are shownin Table 3 and Fig. 6B.
3.2.2.3. Leaf. M.macrophylla (Willd.) O. Kuntze: Tender green color.Upper epidermic cell is irregularly small quadrate, about 9e18 mm
in length; the anticlinal wall of lower epidermic cell is wavedcurve, and paracytic type stomata is observed in lower epidermic;single non-glandular hair and the epipetalous trace is obvious.Parenchyma cell, which is rounded the fiber, contained calciumoxalate crystal. Vessel is scalariform, 90e250 mm in length, 12e26 mm in width. Fiber is slender and long cell, 180e360 mm inlength, 8e15 mm in width. Brown-red or brown-grey inclusion canbe observed. Calcium oxalate crystal is white-grey color and about5e20 mm in diameter. Stomata are obvious.
Micrographs of detailed comparison of three species are shownin Table 3 and Fig. 6C.
3.3. Key to the three species of M.
1. Twig and anthotaxy is densely covered with basal expandedgolden long glandular hair and grey slender hair; Epidermis ofstem and leaf is covered glandular hair. M. glutiaosa (Prain)Y.T.Wei
1. They have not above characters.2. Root is large and thick; the lower side of leaf is not whiteegrey
color. M. macrophylla (Willd.) O. Kuntze2. Root is not large and thick; the lower side of leaf is whiteegrey
color and leaflet is wider than others.$M. latifolia Benth.
4. Discussion
1. In this article, some reliable morphologic characteristics werespecified for identification of the three species of Moghania.
Fig. 5. Transverse sections of petiole of three species of M. A: Outline. B: Epidermis and cortex. C: Vascular bundle. 1. Epidermis; 2. Cortex; 3. Vascular bundle; 4. Pith; 5. Leaf wings;6. Phloem; 7. Vessel; 8. Libriform fiber.
Y.-f. Huang et al. / Pharmacognosy Journal 5 (2013) 30e4038
Morphologic structure of M. macrophylla (Willd.) O. Kuntze ispeculiar compared to the other species. Pod of M. macrophylla(Willd.) O. Kuntze is elliptic with 1e1.6 cm long and 7e9 mmwide, brown, covered short soft hair, while that of M. latifoliaBenth. is covered ferrugineous short soft hair and M. glutiaosa(Prain) Y.T.Wei. is covered light golden base-expanded glan-dular hair. Common pedicel of M. macrophylla (Willd.)O. Kuntze and M. latifolia Benth. are almost no, while that ofM. glutiaosa (Prain) Y.T.Wei. is from 1 to 10 cm. All of thesemicroscopic characteristics made of Moghania easily recog-nizable. Root of M. macrophylla (Willd.) O. Kuntze is large andthick, and petiole has narrow wings, but others are not; thelower side of leaf of M. latifolia Benth. and M. glutiaosa (Prain)Y.T.Wei is whiteegrey color, but that of M. macrophylla (Willd.)O. Kuntze is not.
2. Compared with the M. macrophylla (Willd.) O. Kuntze, thehistological structures of M. Latifolia Benth. and M. glutiaosa(Prain) Y.T.Wei. were similar to some extent. Twig and antho-taxy of M. glutiaosa (Prain) Y.T.Wei is densely covered withbasal expanded golden long glandular hair and grey slenderhair, and epidermis of stem and leaf is covered glandular hairand non-glandular hair, while that of other species just onlyhave soft hair and non-glandular hair. M. macrophylla (Willd.)O. Kuntze andM. glutiaosa (Prain) Y.T.Wei. have a large numberof calcium oxalate crystal, brown-red or brown inclusion andstone cell in parenchyma cell of cortex, phloem and pith, whileM. Latifolia Benth. has a little that of them.
3. According to the morphologic and histological structures re-sults of this article, we have found that M. glutiaosa (Prain)Y.T.Wei. has almost the same large number of calcium oxalatecrystal, brown-red or brown inclusion and stone cell in pa-renchyma cell of cortex, phloem and pith as M. macrophylla(Willd.) O. Kuntze, which were recorded in appendix of“Pharmacopoeia of the People’s Republic of China” (2005 and2010 edition), but M. latifolia Benth. has a little that of them.
Though there are some reports,12e20 (The State PharmacopoeiaCommittee of China, 2010) of M. we didn’t find systematic infor-mation of microscopic authentication and morphoanatomy aboutthree species in this area. What’s more, for the sake of offeringa scientific database to ensure their safety and efficacy, it will beworthwhile doing further correlative studies. We can still authen-ticate and distinguish every species by the microscopic featuresrevealed in our study. We constructed a key to the three species toallow convenient classification.
4. To some extent, authentication by microscopy has advantagescompared with chemical analysis. Fewer samples are required,at lower cost, with easier operation, and greater accuracy. Thisis especially true when the materials must be stored beforeauthentication, due to chemical changes that may occurdepending on the storage environment. Authentication bymicroscopy has a greater reliability, since the features observedare more stable
Fig. 6. Dissociation and powder of root, stem and leaf of three species of M.; A. Root: M. macrophylla (Willd.) O. Kuntze 1. Cork cell; 2. Parenchyma cell; 3. Calcium oxalate crystal inthe parenchyma cell; 4. Brown inclusion; 5. Stone cell; 6. Spiral vessel; 7. Pitted vessel; 8. Fiber; 9. Starch; 10. Calcium oxalate crystal. M. Latifolia Benth 1. Cork cell; 2. Fiber; 3.Parenchyma cell; 4. Brown inclusion; 5. Stone cell; 6. Pitted vessel; 7. Starch; 8. Calcium oxalate crystal. M. glutiaosa (Prain) Y.T.Wei 1. Cork cell; 2. Parenchyma cell; 3. Reticulatevessel; 4. Fiber; 5. Annular vessel; 6. Crystal fiber; 7. Calcium oxalate crystal in the parenchyma cell; 8. Stone cell; 9. Brown inclusion; 10. Starch; 11. Calcium oxalate crystal. B. Stem:M. macrophylla (Willd.) O. Kuntze. 1. Cork cell; 2. Parenchyma cell; 3. Pitted vessel; 4. Fiber; 5. Crystal fiber; 6. Non-glandular hair; 7. Brown inclusion; 8. Starch; 9. Calcium oxalatecrystal. M. Latifolia Benth. 1. Cork cell; 2. Parenchyma cell; 3. Reticulate vessel; 4. Spiral vessel; 5. Stone cell; 6. Fiber; 7. Brown inclusion; 8. Starch; 9. Calcium oxalate crystal.M. glutiaosa (Prain) Y.T.Wei. 1. Cork cell; 2. Parenchyma cell; 3. Annular vessel; 4. Fiber; 5. Brown inclusion; 6. Pitted vessel; 7. Non-glandular hair; 8. Starch; 9. Calcium oxalatecrystal. C. Leaf: M. macrophylla (Willd.) O. Kuntze. 1. Epidermis cell and stoma; 2. Non-glandular hair; 3. Epidermis and veius; 4. Palisade tissue; 5. Non-glandular hair trace; 6. Fiber;7. Crystal fiber; 8. Vessel; 9. Calcium oxalate crystal. M. Latifolia Benth. 1. Epidermis cell and stoma; 2. Annular vessel; 3. Fiber; 4. Non-glandular hair trace; 5. Non-glandular hair; 6.Brown inclusion; 7. Epidermis and veius; 8. Palisade tissue and spomgy tissue; 9. Calcium oxalate crystal.M. glutiaosa (Prain) Y.T.Wei. 1. Epidermis; 2. Basal expanded non-glandularhair and glandular trace; 3. Non-glandular hair; 4. Scalariform vessel; 5. Annular vessel.
In brief, several species of M. are available or presented asmixtures in the herbal markets for use in TCM. Some of them can bedistinguished based on their macroscopic characteristics, but somecannot, especially when the herbs are in a dried and matted con-dition. In such cases, discrimination can be very difficult. Lightmicroscopy should be of great use in identifying these materials.
Acknowledgments
The authors appreciated the National Major Scientific andTechnological Special Project of the State Ministry of Science andTechnology (2011ZX09307-002-01), the Pillar Program of Ministryof Science and Technology of the People’s Republic of China(2012BAI27B07), the China Natural Science Foundation (No.81173653), the Science & Technology Department of Pillar ProgramSichuan Province (2011SZ0233), and the Follow-up Plan for theOutstanding Youth Academic and Technical Pacemaker of Sichuan
Province (2011JQ0051), and the Programming Plan of Aba TeachersCollege (ASB-17) for financial support.
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