Journal of Photochemistry and Photobiology B: Biology 95 (2009) 204212Contents lists available at ScienceDirect

Journal of Photochemistry and Photobiology B: Biology

journal homepage: www.elsevier .com/locate / jphotobiolAn overview of structural features of DNA and RNA complexes with saffroncompounds: Models and antioxidant activity

C.D. Kanakis a, P.A. Tarantilis a, C. Pappas a, J. Bariyanga b, H.A. Tajmir-Riahi c,*, M.G. Polissiou a,*a Laboratory of Chemistry, Department of Science, Agricultural University of Athens, 75 Iera Odos, 118 55 Athens, GreecebDepartment of Chemistry, University of Hawaii-West Oahu, 96-129 Ala Ike, Pearl City, HI 96782, USAcDepartment of ChemistryBiology, University of Qubec at Trois-Rivires, 75 Iera Odos, TR (Qubec), Canada G9A 5H7

a r t i c l e i n f o a b s t r a c tArticle history:Received 23 January 2009Received in revised form 13 March 2009Accepted 20 March 2009Available online 29 March 2009

Keywords:SaffronDNAtRNAAntioxidantConformationFT-IR1011-1344/$ - see front matter 2009 Elsevier B.V. Adoi:10.1016/j.jphotobiol.2009.03.006

* Corresponding authors. Tel.: +1 819 376 5011x331Tajmir-Riahi); Tel.: +30 210 529 4241; fax: +30 210 5

E-mail addresses: heidar-ali.tajmir-riahi@uqtr.cal@aua.gr (M.G. Polissiou).Saffron is the red dried stigmas of Crocus sativus L. flowers and used both as a spice and as a drug in tra-ditional medicine. Its numerous applications as an antioxidant and anticancer agent are due to its second-ary metabolites and their derivatives (safranal, crocetin, dimethylcrocetin). In this work we arecomparing the spectroscopic results and antioxidant activities of saffron components safranal, crocetin(CRT) and dimethylcrocetin (DMCRT) complexes with calf-thymus DNA (ctDNA) and transfer RNA (tRNA)in aqueous solution at physiological conditions Intercalative and external binding modes of saffron com-pounds to DNA and RNA were observed with overall binding constants of Ksafranal = 1.24 103 M1,KCRT = 6.20 103 M1 and KDMCRT = 1.85 105 M1, for DNA adducts and Ksafranal = 6.80 103 M1,KCRT = 1.40 104 M1 and KDMCRT = 3.40 104 M1 for RNA complexes. A partial B- to A-DNA transitionoccurred at high ligand concentrations, while tRNA remained in A-conformation in saffronRNA com-plexes. The antioxidant activity of CRT, DMCRT and safranal was also tested by the DPPH_ (2,2-diphe-nyl-1-picrylhydrazyl) antioxidant activity assay and their IC50 values were compared to that of wellknown antioxidants such as Trolox and Butylated Hydroxy Toluene (BHT). The IC50 values were95 1 lg/mL for safranal and 18 1 lg/mL for crocetin. The inhibition of DMCRT reached a point of38.8%, which corresponds to a concentration of 40 lg/mL.

2009 Elsevier B.V. All rights reserved.1. Introduction

Crocus sativus L. is a plant cultivated in many countries such asGreece, Spain, Italy, Iran and India. Its product is the well knownspice called saffron. Saffron is the dried red stigmas of the flower.Its use as a spice and drug in folk medicine is known since theantiquity [1]. Saffron main substances are safranal, crocins andpicrocrocin. Safranal, the main component of the distilled essentialoil, is a monoterpene aldehyde, responsible for its characteristic ar-oma. Crocins, glucosyl esters of crocetin, are unusual water-solublecarotenoids and are responsible for its characteristic color. Picro-crocin, the glucoside precursor of safranal, is responsible for saffronbitter taste [2]. Dimethylcrocetin is another pure derivative ofcrocins [3]. It has been reported that orally administered crocinsare hydrolyzed to crocetin before or during intestinal absorptionand absorbed crocetin is partly metabolized to mono- and diglu-coronide conjugates [4].ll rights reserved.

0; fax: +1 819 376 5084 (H.A.29 4265 (M.G. Polissiou).(H.A. Tajmir-Riahi), mopo-Metabolic processes produce free radicals [5,6]. These mole-cules function as physiological signals. However, free radicals arehighly reactive and unstable [7]. Characterized by unpaired elec-trons in their outer orbit, free radicals also can cause oxidativedamage to cells and tissues [8]. Carotenoids play an important rolein human health by acting as biological antioxidants, protectingcells and tissues from damaging effects of free radicals and singletoxygen [9,10]. The antioxidant properties of crocins have beenstudied by several laboratories [1114]. Diets rich in antioxidantscontribute to a lower incidence of several major chronic diseases.In particular, cancer development or growth is inhibited by antiox-idants. Despite saffron use in traditional medicine, the biologicalactivity and preventing effect in anticancer research is in develop-ment. The effect of crocetin on intracellular nucleic acids and pro-tein synthesis in malignant cells has been examined [15]. Crocetinhad a dose-dependent inhibitory effect on DNA and RNA synthesisin isolated nuclei and suppressed the activity of purified RNA poly-merase II. Crocetin and dimethylcrocetin are highly effective,inhibiting the proliferation and inducing differentiation of HL-60leukemic cells and their action was compared with that of all-transretinoic acid [16]. Also, crocetin and dimethylcrocetin are not pro-vitamin A precursors and could therefore be less toxic than reti-noids. The anticarcinogenic properties of saffron were also

C.D. Kanakis et al. / Journal of Photochemistry and Photobiology B: Biology 95 (2009) 204212 205investigated by the effects of saffron carotenoids on histone H1structure and H1DNA interaction [17]. It was reported that croc-ins and dimethylcrocetin isolated form saffron were non-muta-genic and nontoxic [1,18,19]. Recent studies present theprotective effect of safranal, C. sativus stigma extract and trans-cro-cin 4 against methyl methanesulfonate (MMS)-induced DNA dam-age in mice organs [20,21]. In recent reports the complexation ofsafranal, crocetin (CRT) and dimethylcrocetin (DMCRT) with DNAand tRNA were discussed [22,23].

We report a comparison of the interaction of safranal, crocetinand dimethylcrocetin (Fig. 1) with DNA and tRNA in vitro with re-gard to their binding modes, stability and structural aspects of theligandnucleic acids complexes. The various possibilities of ligandbinding modes with regard to antioxidative activity and the way inwhich ligand interaction can protect DNA and tRNA from damageby free radicals are discussed here.

2. Materials and methods

2.1. Materials

Highly polymerised type I calf-thymus DNA sodium salt (7% Nacontent) and yeast transfer RNA sodium salt were purchased fromSigma Chemical Co., and used as supplied. The absorbance at 260and 280 nm was recorded, in order to check the protein contentof DNA and tRNA solution. The A260/A280 ratios were 1.85 forDNA and 2.2 for tRNA showing that polynucleotides are sufficientlyfree from protein. Safranal (75%) was supplied from Fluka. 2,2-di-phenyl-1-picrylhydrazyl (DPPH_) (90%) were purchased from SIG-MA. Trolox (97%) was purchased from Aldrich and ButylatedHydroxy Toluene (BHT) from BDH. Other chemicals were of re-agent grade and used without further purification. Stigmas of pureCH3H3C

CH3

CHO

CH3 CH3

O

Safranal

OH

CH3 CH3

O

Crocetin (CRT)

OCH3

Dimethylcrocetin (DMCRT)

Fig. 1. Chemical structures of safranared Greek saffron were kindly supplied by the Cooperative of Saf-fron, Krokos Kozanis.

2.2. Preparation of crocetin (CRT) and dimethylcrocetin (DMCRT)

The preparation of crocetin and dimethylcrocetin was per-formed according to the method described in previous work [24].

2.3. DPPH antioxidant capacity assay

The antioxidant activity of safranal, crocetin and dimethylcroce-tin was performed according to the methods already described byKanakis et al. [24].

2.4. Preparation of stock solutions

DNA or tRNA sodium salt (5 mg/mL) was dissolved in Tris/HClbuffer (pH7.0) at 5 C for 24 h with occasional stirring to ensurethe formation of a homogeneous solution. The final concentrationof the stock tRNA solution was determined spectrophotometricallyat 260 nm using molar extinction coefficient e260 = 9250 cm1 M1

(expressed as molarity of phosphate groups) [25]. The final concen-tration of the calf-thymus DNA solution was determined spectro-photometrically at 260 nm using molar extinction coefficiente260 = 6600 cm1 M1 (expressed as molarity of phosphate groups)[26]. The average length of the DNA molecules, estimated by gelelectrophoresis was 9000 base pairs (molecular weight 6 106 Da). In the case of the DNA stock solutions, the appropriateamounts of safranal, crocetin (CRT) (0.266.25 mM) and dim-ethylcrocetin (DMCRT) (0.261.56 mM) were prepared in distilledwaterethanol (50/50%) and added dropwise to DNA solution inorder to attain the desired ligand/DNA (P) molar ratios (r) of 1/48CH3CH3

O

OH

CH3CH3

O

OCH3

l, crocetin and dimethylcrocetin.

206 C.D. Kanakis et al. / Journal of Photochemistry and Photobiology B: Biology 95 (2009) 204212to 1/2 for safranal and crocetin/DNA and 1/48 to 1/8 for dim-ethylcrocetin/DNA, with a final DNA concentration of 6.25 mM.In the case of tRNA stock solutions, the appropriate amounts ofsafranal, crocetin and dimethylcrocetin (0.261.56 mM) were pre-pared in distilled waterethanol (50/50%) and added dropwise totRNA solution, in order to attain the desired ligand/tRNA (P) molarratios (r) of 1/48 to 1/8 with a final tRNA concentration of 6.25 mM.

2.5. FT-IR spectroscopic measurements

Infrared spectra were recorded on a Nicolet Magna 750 FT-IRspectrophotometer (DTGS detector, Ni-chrome source and KBrbeam splitter) with 100 scans and resolution of 4 cm1. Spectrawere collected and manipulated using the OMNIC (ver. 3.1) soft-ware supplied by the manufacturer of the spectrophotometer.Spectra were recorded after 1 h of incubation, using AgBr windows.The difference spectra [(polynucleotide solution + ligand solu-tion) (polynucleotide)] were generated using bands at968 cm1 (DNA) and 867 cm1 (RNA) as internal standard[27,28]. These vibrations are due to sugar CC stretching modesFig. 2. Infrared absorption spectra of the free DNA (first curve) and difference spectra600 cm1.and exhibit no spectral changes upon ligandpolynucleotideinteraction.

2.6. UVVis absorption spectroscopy

The UVVis spectra were recorded on a Jasco UVVis, V-550spectrophotometer with a slit of 2 nm and scan speed of400 nmmin1. Quartz cuvettes of 1 cm were used. The absorbanceassessments were performed at pH7.0 by keeping the concentra-tion of DNA or tRNA constant (0.25 mM), while varying the concen-tration of the ligands 0.0025 mM0.625 mM).

The values of the binding constants were obtained according tothe method described by Connors [29]. It is assumed that the inter-action between the ligand L and the substrate S is 1:1; for this rea-son a single complex SL (1:1) is formed. It was also assumed thatthe sites (and all the binding acts) are independent and finallythe Beers law is followed by all species. A wavelength is selectedat which the molar absorptivities eS (molar absorptivity of the sub-strate) and e11 (molar absorptivity of the complex) are different.Then at total concentration St of the substrate, in the absence of li-of saffron compounds (six curves) in aqueous solution at pH7 in the region 1800

C.D. Kanakis et al. / Journal of Photochemistry and Photobiology B: Biology 95 (2009) 204212 207gand and the light path length is b = 1 cm, the solution absorbanceis

Ao eSbSt 1

In the presence of ligand at total concentration Lt, the absor-bance of a solution containing the same total substrate concentra-tion is

AL eSbS eLbL e11bSL 2

(where [S] is the concentration of the uncomplexed substrate, [L]the concentration of the uncomplexed ligand and [SL] is the concen-tration of the complex) which, combined with the mass balance onS and L, gives

AL eSbSt eLbLt De11bSL 3

where De11 = e11 eS eL (eL molar absorptivity of the ligand). Bymeasuring the solution absorbance against a reference containingligand at the same total concentration Lt, the measured absorbancebecomes

A eSbSt De11bSL 4Fig. 3. Infrared absorption spectra of the free tRNA (first curve) and difference spectra600 cm1.Combining Eq. (4) with the stability constant definitionK11 = [SL]/[S][L], gives

DA K11De11bSL 5

where DA = A Ao. From the mass balance expression St = [S] + [SL]we get [S] = St/(1 + K11[L]), which is Eq. (5), giving Eq. (6) at the rela-tionship between the observed absorbance change per centimeterand the system variables and parameters.

DAb

StK11De11L1 K11L

6

Eq. (6) is the binding isotherm, which shows the hyperbolicdependence on free ligand concentration.

The double-reciprocal form of plotting the rectangular hyper-bola 1y

fd 1x ed, is based on the linearization of Eq. (6) according

to the following equation,

bDA

1StK11De11L

1StDe11

7

Thus the double reciprocal plot of 1/DA versus 1/[L] is linear and thebinding constant can be estimated from the following equationof saffron compounds (six curves) in aqueous solution at pH7 in the region 1800

208 C.D. Kanakis et al. / Journal of Photochemistry and Photobiology B: Biology 95 (2009) 204212K11 interceptslope

83. Results and discussion

3.1. Saffron componentsDNA complexes

Safranal, crocetin (CRT) and dimethylcrocetin (DMCRT) bindexternally to DNA. Evidence for external binding comes from shift-ing of the guanine band at 1710 cm1 to 17081707 cm1 (spectranot shown). External binding of safranal, CRT and DMCRT was alsoobserved by Bathaie et al. [30] and Hoshyar et al. [31]. The ob-served shifting was accompanied by an increase in the intensityof the guanine vibration. The positive features at 1696, 1672 and1685 cm1 are due to the increase of the intensity of the guanineFig. 4. UVVisible spectra of (A) safranal (0.08 mM) at 314 nm, (B) crocetin (0.08 mM) awith final DNA concentration 0.25 mM. Plot of 1/(A A0) versus 1/C for DNA and its drugabsorption at different drug concentrations (0.00250.625 mM) and final DNA concentrband (Fig. 2). The observed spectral changes are due to an indirectinteraction of the ligands with guanine N-7. No major spectralshifting was observed for the backbone phosphate group at1225 cm1. However positive peaks at 1233, 1220 and 1221 cm1

(Fig. 2) are due to an increase in the intensity of the phosphatestretching vibrations as a result of ligandphosphate binding. Theincrease in the intensity of DNA vibrations at high ligand contentcan also be attributed to some degree of helix destabilization. Sim-ilar intensity increase was also observed for DNA vibrations in thepresence of high copper and flavonoid concentration [32,33]. How-ever some degree of drug intercalation occurs with DNA duplex.Evidence for this comes from a reduction in the intensity of theUVVis bands characteristic of saffrons components upon DNAintercalation. The decrease in absorbance of characteristic UVVisband at 314 nm (safranal), 424 and 449 nm (CRT) and 427 and453 nm (DMCRT) is indicative of some degree of drug intercalationt 424 nm and 449 nm, and (C) dimethylcrocetin (0.04 mM) at 427 nm and 453 nm,complexes, where A0 is the initial absorption of DNA (260 nm) and A is the recordedation of 0.25 mM.

C.D. Kanakis et al. / Journal of Photochemistry and Photobiology B: Biology 95 (2009) 204212 209via DNA duplex (Fig. 4). Intercalation was greater for DMCRT com-plexes than safranal and CRT complexes. The limitation of molecu-lar movements of safranal, CRT and DMCRT causes a decrease intheir ability to absorb light energy [34].

3.2. Saffron componentstRNA complexes

Safranal, CRT and DMCRT bind externally to tRNA. Evidence forexternal binding comes from infrared and UVVis spectroscopy.The band at 1700 cm1 in the free tRNA spectrum (Fig. 3) relatedto C@O stretching vibrations of guanine and uracil bases exhibitedshifting towards lower frequency at 16941690 cm1 (spectra notshown) upon drug complexation. The observed shifting wasFig. 5. UVVisible spectra of (A) safranal (0.08 mM) at 314 nm, (B) crocetin (0.02 mM) awith final tRNA concentration of 0.1 mM. Plot of 1/(A A0) versus 1/C for tRNA and itsrecorded absorption at different drug concentrations (0.010.625 mM) and final tRNA caccompanied by an increase in the intensity of the guanine vibra-tion. The positive features at 1695 and 1691 cm1 are due to theincrease of the intensity of the guanine band (Fig. 3). The spectralchanges observed for the band at 1700 cm1 are due to some de-gree of drug interaction with guanine bases. The backbone PO2asymmetric stretching band at 1240 cm1 showed alteration inthe spectra of safranaltRNA adducts and CRTtRNA adducts. Inthe case of safranaltRNA adducts the band at 1240 cm1 exhibitedshifting to a lower frequency at 1237 cm1 (spectrum not shown)and the positive peak at 1235 (Fig. 3) is due to an increase in thephosphate stretching vibrations upon safranal binding. The samewas observed for CRTtRNA adducts where the asymmetricstretching band at 1240 cm1 shifting to a lower frequency att 424 nm and 449 nm, and (C) dimethylcrocetin (0.04 mM) at 427 nm and 453 nm,drug complexes, where A0 is the initial absorption of tRNA (260 nm) and A is theoncentration of 0.1 mM.

Saffron's Components-DPPH assay

0

20

40

60

80

100

0 20 40 60 80 100 120 140 160 180 200

Safranal-DPPHassay

DMCRT-DPPHassay

CRT-DPPH assay

Concentration g/mL

Inhi

biti

on %

Fig. 6. Inhibition (%) against various concentrations of safranal, DMCRT and CRT used in the DPPH test. Results are means of three different measurements.

Table 1Antioxidant activity of saffron compounds.

Compound DPPH method IC50 value (lg/mL)

Safranal 95 1Crocetin (CRT) 18 1Trolox 5.2 1BHT 5.3 1

Dimethylcrocetin (DMCRT) 40 1a

a At this concentration DMCRT has the maximum inhibition of 38.8 %.

210 C.D. Kanakis et al. / Journal of Photochemistry and Photobiology B: Biology 95 (2009) 2042121232 cm1 (spectrum not shown) and the positive peak at 1228(Fig. 3) is due to an increase in the phosphate stretching vibrationsupon safranal binding. In the case of DMCRTtRNA adducts, no ma-jor spectral shifting was observed for the backbone phosphategroup at 1240 cm1. However the positive peak at 1241 cm1 isdue to an increase in the intensity of the PO2 asymmetric stretch-ing band. Additional evidence for the external binding of the saf-frons components comes also from UVVis spectroscopy. Theintensity increase of characteristic UVVis band at 314 nm (safr-anal), 424 and 449 nm (CRT) and 427 and 453 nm (DMCRT) isindicative of external binding (Fig. 5). The increase in the intensityof characteristic UVVis bands is due to ligandtRNA interaction attRNA surface, which does not limit the mobility of the ligandsaround tRNA molecule [35].

3.3. DNA and RNA conformations

A partial B- to A-DNA transition occurred upon carotenoids andsafranal adducts formation at high drug concentrations. Evidencefor this comes from the shift of the sugarphosphate band at837 cm1 (B-DNAmarker) towards a lower frequency. In the differ-ence spectra of carotenoids and safranalDNA complexes (Diff. = 1/2 for safranal and CRT, Diff. = 1/8 for DMCRT, Fig. 2), the emergenceof a new peak at about 820 cm1 accompanied by loss of the inten-sity of the band at 837 cm1 (B-DNA marker) [36]. Similarly, theother B-DNA marker band at 1710 cm1 shifted to 17081707 cm1 upon carotenoids and safranal complexation. However,other B-DNA marker band at 1225 cm1 showed no major shiftingin the spectra of the carotenoids and safranalDNA adducts. In acomplete B to A transition, the B-DNA marker bands are observedat 17101700 cm1, 12251240 cm1, 825800 cm1, respectively,and a new band appears at about 870860 cm1 [3743]. The ob-served shifting for the bands at 837 cm1 and 1710 cm1, is due toa partial transition of the B-DNA to A-DNA upon carotenoids andsafranal complexation. However, CD spectroscopic studies of theinteraction of saffron carotenoids and safranal with DNA indicatedB to C-DNA transition [30,31].

The free tRNA is in A-conformation with characteristic infraredbands at 1700 cm1 (guanine), 1240 cm1 (phosphate), 868 and815 cm1 (phosphodiester) (Fig. 3). The presence of the majorbands at 16941690 cm1 (guanine), 12391232 cm1 (phos-phate), 867865 cm1 (ribosephosphate) and 815 cm1 (phospho-diester) are indicative of tRNA remaining in the A-conformationupon ligand complexation [34].

3.4. Stability of ligandDNA and ligandtRNA adducts

The binding constants estimated for the ligandDNA complexeswere Ksafranal = 1.24 103 M1, KCRT = 6.20 103 M1 and KDMCRT= 1.85 105 M1 (Fig. 4). Similarly one binding constant was calcu-lated for the ligandtRNA adducts with Ksafranal = 6.80 103 M1,KCRT = 1.40 104 M1 and KDMCRT = 3.40 104 M1 (Fig. 5). Thestability of adduct formation in both cases is DMCRT > CRT > safr-anal. It should be noted that even though the calculated K valuesfor ligandDNA and ligandtRNA interactions are small, majorDNA and tRNA structural changes occurred upon carotenoids andsafranal binding.

3.5. Antioxidant activity

The results of the antioxidant activity of safranal, CRT andDMCRT tested by the DPPH assay and shown in Fig. 6 and Table1 are related to the IC50 values of safranal and CRT.

Safranal, the main component of saffrons essential oil, is amonoterpene aldehyde; its free radical scavenging activity isshown in Fig. 6. From its IC50 value (IC50 = 95 1 lg/mL, Table 1)it can be observed that it has lower antioxidant activity than CRT(IC50 = 18 1 lg/mL, Table 1).

The antioxidant activity of dimethylcrocetin is shown in Fig. 6. Itis shown that for concentrations up to 40 lg/mL, the antioxidantactivity is increasing (inhibition percentage 38.8%, Table 1), whilefor concentrations higher than 40 lg/mL it is decreasing. Thisshould be explained by the fact that at higher concentrationsDMCRT shows pro-oxidant effect since theoretically it could gener-ate more radicals than it consumes [44,45]. The same resulted forcrocins when they were tested for their antioxidant activity bythe DPPH assay [46]; crocins showed an IC50 value of 44 1 lg/mL and for a concentration higher than 50 lg/mL their antioxidantactivity started to decrease. These results are in agreement withthose reported by Pham et al. [12]. Comparing the antioxidantactivity of DMCRT with that of safranal it can be observed fromFig. 6 that for concentrations up to 40 lg/mL, DMCRT has higherantioxidant activity (inhibition% = 38.8%) than that of safranal(inhibition% = 20%).

Free radical scavenging activity of CRT is higher than that ofDMCRT (CRT IC50 = 18 1 lg/mL). The structural differences be-tween the two carotenoids are the reason for that behavior as is

C.D. Kanakis et al. / Journal of Photochemistry and Photobiology B: Biology 95 (2009) 204212 211described by Jimnez-Escrib et al. [47]. In both carotenoids thelength of the conjugated double bond system is the same; the dif-ference between them is the presence of the hydroxyl moiety ofthe carboxylic group on each of the terminal of the unsaturatedhydrocarbon chain in the case of CRT and the presence of onemethyl ester group on each terminal of the unsaturated hydrocar-bon chain in the case of DMCRT. The capability of the stable freeradical DPPH to react with H-donors such as OH of carboxylicgroups, is the mechanism of action of this antioxidant activity as-say. The presence of such H-donors in CRT makes it more effectiveto react with DPPH.

The antioxidant activity of CRT and safranal was compared tothat of Trolox and BHT which are well known antioxidants (Table1). In general, it can be observed that saffrons components showedlower antioxidant activity than that of Trolox and BHT and espe-cially safranal; but the antioxidant activity of CRT is closer toBHT and Trolox. Thus CRT is more effective than safranal. Howeverit can be assumed that the synergistic effect of all the bioactiveconstituents gives to saffron spice a significant antioxidant activity.

4. Summary

Safranal, CRT and DMCRT bind DNA via external binding and insome degree by intercalation while they bind tRNA only externally.The stabilities of DNA complexes formed are Ksafranal = 1.24 103 M1, KCRT = 6.20 103 M1 and KDMCRT = 1.85 105 M1 andfor tRNA adducts are Ksafranal = 6.80 103 M1, KCRT = 1.40 104 M1 and KDMCRT = 3.40 104 M1. The stability of adduct for-mation in both cases is DMCRT > CRT > safranal. The complexationof safranal, CRT and DMCRT leads to a partial B to A-DNA transitionwhile tRNA remains in A-conformation. The antioxidant activity ofsaffron carotenoids is more effective than safranal. However thesynergistic effect of all the bioactive constituents gives to saffronspice a significant antioxidant activity. The antioxidant activity ofsaffron compounds can protect DNA and tRNA from harmful chem-ical reaction in these ligandpolynucleotide complexes but in or-der to have a solid proof for that, further experimental work hasto be performed in order to prove that the oxidation products ofthe saffron compounds does not harm DNA/tRNA.

5. Abbreviations

CRT crocetinDMCRT dimethylcrocetinctDNA calf-thymus DNAFT-IR Fourier transform infraredUVVis UltravioletVisible

Acknowledgments

This work was supported by grants from Natural Sciences andEngineering Research Council of Canada (NSERC) and the Agricul-tural University of Athens.

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