DORMANCY AND CHANGES IN PHENOLIC COMPOUNDS OF ARTEMESIA SCOPARIA SEEDS

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1 Annals f Arid Zne 27 (3&4) : ,1988 DORMANCY AND CHANGES IN PHENOLIC COMPOUNDS OF ARTEMESIA SCOPARIA SEEDS FAWZIA AL-CHARCHAFCHI, FAWZIA M.J. REDHA AND WIDAD M. KAMEL Scientific Research Cuncil, P.O. Bx 2371, JadiTiyah, Baghdad, Iraq ABSTRACT Pr germinatin f fresh seeds f Arttmesi scpria (30% in husked and 5% in unhusked seeds) was fund t be due t the relatively high percentage f phenlic cnstituents f the seeds (9.75) and their bracts (6.50). After ne year f dry strage, when the ttal phenlic cmpunds decreased t t.45% in seeds and 3.95% in bracts, the germinatin f bth husked and unhusked seeds imprved t 98% and 30%, respectively. INTRODUCTION Artemisia scparia Waldstetkit (Cmpsitae), is a widely scattered and useful range shrub in the arid regins f Iraq (A l-charchafchi et al., 1986). Sme aspects f seed germinatin f this species have been studied (AI-Ani et ai., 1971, AI-Charchafchi et ai., 1986), but nthing is knwn abut the bserved seed drmancy and the pssible rle f the bracts surrunding the seeds n inhibitin f germinatin. AI-Charchafchi et ai. (1987) indicated that phenlic cmpunds might be invlved in the germinatin inhibitin f Artemesia herba-alba. The studies were, therefre, undertaken t find ut the endgenus phenlic cmpunds f A. scparia, the pssible changes during the dry strage and their effects n seed germinatin. MATERIAL AND METHODS Husked (bracts remved) and unhusked fresh seeds f A. scparia were kept in paper bags and stred at rm temperature in December Germinatin tests were carried fr tw years in glass petri dishes (9 cm dia) lined with a layer f Whatman N.3 filter paper, and mistened with 5 ml f distilled water r test slutins. Fifty seeds per dish were replicated fur times fr each treatment. All tests were dne in an incubatr (Precisin Mdel 806) set at 20 C and under cntinuus illuminatin. Cunts f germinatin were made every ther day fr a perid f 12 days. The phenlic cnstituents were studied fr the husked seeds and bracts at 1, 6 and 12 mnths after harvest, fllwing AI-Charchafchi et al. (1987). Gradient

2 172 : CHARCHAFCHI et ai. cncentratins were prepared frm certain amunt f ttal phenlic cnstituents and tested fr their inhibitry actin n nn-drmant husked seeds f A. scparia. The phenlic cmpunds were separated by silica thin layer chrmatgraphic plates (20 x 20 cm, 2 mm and 5 mm thickness fur preparative scale) develped with 15% acetic acid/water mixture. The spts were detected by ammnia vapurs, ferric,chlride slutin, and ethan!ic slutin f ptassium hydrxide. Fr the identificatin f the separated phenlic cmpunds tw dimensinal paper chrmatgraphy using Whatman 3MM (Mabry et ai., 1970) and develped by the descending technique, was carried ut with tertiary butanl/acetic acid/water mixture (TBA, 3:1:1) as the first and water/acetic acid mixture (15:85) as the secnd slvent. The UV-spectra were recrded with SP8-250 UV/Vis Pye-Unicam Spectrpht meter in ethanl; A max fr the islated cmpund frm the seeds were 320, 299, 280, 246 nm and A max fr the islated cmpund frm the bracts were 399 and 246 nm. Accurately weighed amunts f these purified cnstituents were separately disslved in distilled water and gradient cncentratins were prepared t test th.;:ir inhibitry actin n the husked seeds f A. scparia. RESULTS AND DISCUSSION Effects f the bracts and seed age n germinatin Fresh husked seeds exhibited lw percentage f germinatin in winter (Fig. 1). Hwever, in tbe fllwing spring and summer, the germinatin capacity imprved prgresively t a maximum f 98% afler ne year. Similar resulls were reprted fr A. herba-alba by AI-Charchafchi and Jawad (1982). In unhusked seeds f A. scparia germinatin did nt exceed 30% thrughut the tw year strage perid, clearly indicating the inhibitry effects f bracts (Fig. I). The pssibility f mechanical resistance t germinatin is ruled ut by the fact that tbe bracts are pen at the tp. AI-Charchafchi et al. (I986) shwed that as the seeds becme lder (98% germinatin) respiratin significantly increases. Thus certain changes in the seed t take place which result in their better germinatin. Changes in ttal phenlic cntents during the dry swrage The Rf values and clur reactions f the separated spts are given in Table t. The Rf values f the majr spt f ne mnth ld seeds detected by this prcedure were 82 and 65 in TBA and 15% acetic acid, respectively. Frm the seeds, 227 mg (Rf = 85) f the majr f1avnidal cnstituents was separated and 25 mg

3 PHENOLS IN ARTEMESIA SEEDS: ! z ē- «Ẕ f5 e r 20 r I 4 8 SEED AGE (MONTH} Figure 1. Effect f time lapse and the presence r absence f bracts arrund the seeds n their germinatin. Husked, Unhusked. (Rf = 05) frm the bracts. The ttal phenlic cnstituents decreased frm the initial levels (ne mnth after harvest) f 6.50% t 5.30% at 6 mnths and t 3.95% at ] 2 mnths after harvest. In the seeds, initial level f phenlic cmpunds ne mnth after harvest (9.75%) drpped t ].45% at 12 mnths after harvest. Higher percentage f the ttal phenlic cmpunds left in the bracts even after ne year explains the lwer germinatin percentage f the unhusked seeds. Lwer phenlase activity might be related t the lwer activity f gr_wth prmting substances in the drmant seeds. Effects f the islated phenlic cmpunds n germinatin and seedling develpment The inhibitry effects f the phenlic cmpunds islated frm the seeds and the bracts at tw stages (after harvest time) n germinatin f husked seeds and cnsequently n seedling develpment are shwn in Figure 2. It is seen that germinatin sharply decreased as the applied cncentratin f the ttal phenlic cmpunds islated frm hacts increased (Fig. 2A). Hwever, the ttal phenlic cmpunds islated frm ]-mnth ld bracts decreased the germinatin significantly mre (P=O.OI) then phenlic cmpunds islated frm 12-mnth ld bracts. In ther wrds, ttal phenlic cmpunds islated frm the bracts f drmant seeds prbably cntained mre inhibitry fractins. Cmparable effects were bserved with regards t seedling develpment (Figure 28).

4 174: AL CHARCHAFCHI et al. :.2. c z 0. "0 '" C '" E. '0 c..c. '" bl/ C "0 C '"C.2 E '" c 0 = > '"c E '" c "0 0 '" z ' lr) CCl M II') LO 0 I,(') S 0 <5 0-- r> V).,.. V) 0 0- N M '" "0 Vl '". -c._ '" bl/-0> L.l.,cn..

5 PHENOLS IN ARTEMESIA SEEDS: !00,.: "i"s 60 z 40 a: :520 'A I fv"onth '2 MONTH '00 c.e " '2.8 CONCENTATO" 1/11 05,.() , " IzB OS, SO CONCENTA TION I 'Ill ) Figure 2. Effect f different cncentratins r islated endgenus phenlic cmpunds frm the bracts (A,B) and husked seeds (C,DI n germinatin and seedling develpment f the nn-drmant husked seeds f A. scparia Similar inhibitry effects f the phenlic cmpunds islated frm husked seeds n germinatin were lwer as cmpared t the inhibitry effect f such cmpunds islated frm bracts, at bth the stages f islatin time (Fig. 2, C.D). Identificatin f the phenlic cmpunds The TLC f the phenlic fractins btained frm 1- and 12- mnth ld seeds revealed the presence f similar cmpunds f f1avnidal type, but the quantitatively majr cmpund detected in the 1- mnth ld seeds was fund as the minr cmpund in the 12 -mnth ld seeds (similar Rf values in different slvent systems and the same clur reactins) and vice - versa (Table 1). This was pssibly due t the enzymatic reactins during the dry strage perid. UV spectral analyses and clur reactins f the majr phenlic cmpund f the 1- mnth ld seeds cnfirmed it t be a 3-methxy flavne derivative and pssibly. Quercetin-3-methyl ether. The TLC analyses and the clurs bserved under lng UV shwed that the ther (flavnidal) cnstituent f the seeds was different frm that separated frm the bracts (Table I).

6 176: AL-CHARCHAFCHI et al. Fr the islated flavnidal glycside f the bracts, n specific structure culd be suggested fr the aglycn part because f scarcity f mateiial, nt enugh fr the hydrlysis and chemical identiflcatin prcedures. All chrmatgraphic results btained by TLC and tw dimensinal paper chrmatgraphic techiques fr the majr phenlic cnstituent were cnsistent with the suggested chemical structure. In the drmant husked seeds, the quantitatively majr phenlic cmpund separated by preparative TLC (Quercetin-3-methyl ether) was tested (r its inhibitry actin n germinatin and seedling develpment as shwn in Figure 3. It shws that significant reductin in the germinatin percentages and seedling grwth (P=O.OJ) was btained by all cncentratins used. It is quite reasnable t cnclude that Quercetin- 3-methyl ether was the dminant phenlic inhibitr cntained in the drmant husked seeds f A. scparia. J 2': ::; ::; J:.- l 100 ē') Z Ib z 10 w Q..J bo - 12 e') II : J II> b CONCENTPATION l/!i Figure 3. Effect f different cncentratin f islated majr phenlic cmpund frm the drmant husked seeds n germinatin and seedling develpment f the nn-drmant husked seeds.) Germinatin (%), 0 Seedling length (mm) REFERENCES AI-Ani, T.A., AI-Mufti, M.M. and AI-Charchafchi, F.M.R Range Resurces f Iraq. IV. Germinability f sme native range species f Iraq. Institute f Applied Research f Natural Resurces, Technical Reprt N.3, Abu-Ghraib, Iraq. AI-Charchafchi, F., Clr, M.A. and Jawad, M Seed characteristics and germinatin f Artemesia scparia. Annals f Arid Zne 26 (2) : AI-Charchafchi, F., and Jawad, M Effect f light, temperature, strage cnditins and seeds age n germinatin f desert seeds. I. Artemesia herba-alba.

7 FHENOLS IN ARTEMESIA SEEDS: 177 Jurnal f Agriculture and Water Resurces Research, Iraq. I : AI-Charchafchi, F., Redha, F.M.J., Kamel, W.M. and AI-Faky, S.M Drmancy f Artemesia herba-alba seeds in relatin t endgenus chemical cnstituents. Jurnal f Bilgical Sciences Research. 18 (Suppl): Mabry, T" Markham, K. and Thmas, M. 197 The Systematic Identificatin f Flavnids. Springer-Verlag, Berlin.