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Journal of Cultural Heritage 13 (2012) 89–97

ase study

dentification of natural dyes in historical textiles from Romanian collections byC-DAD and LC-MS (single stage and tandem MS)

rina Petroviciua,∗,e, Ina Vanden Bergheb, Ileana Cretuc, Florin Albud, Andrei Medvedovici e

National Museum of Romanian History, 030026 Bucharest, RomaniaRoyal Institute for Cultural Heritage, 1000 Brussels, BelgiumNational Museum of Art of Romania, 010063 Bucharest, RomaniaBioanalytical Laboratory, S.C. LaborMed Pharma S.A., 032266 Bucharest, RomaniaDepartment of Analytical Chemistry, Faculty of Chemistry, University of Bucharest, 050663 Bucharest, Romania

r t i c l e i n f o

rticle history:eceived 7 February 2011ccepted 5 May 2011vailable online 14 June 2011

eywords:atural dyesiquid chromatographyiode Array Detectionass spectrometry (single stage MS and

andem MS)omanian historic textiles (religiousmbroideries and brocaded velvets)

a b s t r a c t

In this study, the dyes present in five 17th- to 18th-century textiles from the National Museum of Artof Romania, three religious embroideries and two brocaded velvets, are characterized and discussed,together with earlier results on textiles from Romanian collections obtained by the same research group.Dye analyses were performed using two methods: the well-established liquid chromatography-diodearray detection (LC–DAD) and a recently developed liquid chromatography-mass spectrometry (LC–MS)analytical protocol. The examination of very small historical samples by both techniques allows a betterinsight in the advantages and limitations of the two approaches to real analyses to be obtained. LC–MSdata interpretation is based entirely on the results accumulated for dye standards. Electrospray ionization(ESI) was used in the negative ion mode and an ion trap served as mass analyzer. Both single stage (MS)and tandem (MS/MS) mass spectrometric approaches were considered. The dyes and natural sourcesidentified by both analytical techniques are discussed in the historical context of the textiles, with respectto earlier results collected for similar Romanian objects. The study showed that the dye sources found inthe 17th- and 18th-century Romanian velvets and embroideries were produced using a wide variety of

dye sources, suggesting influences from Europe as well as from Asia Minor. Dye sources imported fromNew World have been also detected. The range of biological sources is in very good correspondence withearlier results obtained from textiles in the Romanian Collections. LC-MS (single stage and tandem MS)approaches have been demonstrated to be valuable tools for dye identification in small-scaled samplesfrom historical textile objects only if sufficient knowledge on the dyes and their biological sources is first

rime

accumulated within expe

. Introduction and research aims

Dyes obtained from naturally occurring biological sources haveeen used in textile dyeing since antiquity. The identification ofheir use in historic pieces may provide useful information abouthere, when and how these objects were created and may also

ontribute to their conservation.

Several studies have been dedicated to the identification of dyes

rom biological sources in various types of textile preserved inestern European collections in the last 50 years [1–6]. In contrast,

bjects in Eastern European museums and monasteries – including

∗ Corresponding author.E-mail addresses: petroviciu@yahoo.com, irinapetroviciu@mnir.ro

I. Petroviciu), ina.vandenberghe@kikirpa.be (I. Vanden Berghe),dileana@yahoo.com (I. Cretu), florin alburo@yahoo.com (F. Albu),vmedved@yahoo.com (A. Medvedovici).

296-2074/$ – see front matter © 2011 Elsevier Masson SAS. All rights reserved.oi:10.1016/j.culher.2011.05.004

nts performed on standard dyes and standard dyed fibers.© 2011 Elsevier Masson SAS. All rights reserved.

studies on dyes in textiles from Romanian collections – have onlyrecently been considered as subjects for characterization of thedyes present [7–11].

In the present work, dye analysis on three religious embroi-deries and two brocaded velvets, dating from the 17th to 18thcenturies, from the National Museum of Art of Romania is pre-sented and discussed, together with earlier results on textiles fromRomanian collections obtained by the same research group. Dataresulting from the liquid chromatography–diode array detection(LC–DAD) method of analysis are compared with those producedusing a newly developed liquid chromatography–mass spectrome-try (LC–MS) analytical protocol, based on the progressive use ofsingle stage (MS) and tandem (MS/MS) mass spectrometry. The

comparison of the resulting pattern of dye constituents obtainedthrough application of the two alternative techniques offers avery good insight into the advantages and limitations of the twoapproaches to real historical problems, where the reality of degra-dation of the textiles and the very limited sample sizes available

9 Cultur

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2

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2

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2

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2

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0 I. Petroviciu et al. / Journal of

ust be faced. This results in a mutual validation of the analyt-cal protocols developed for the identification of dyes in culturaleritage textiles.

The biological sources of dyes identified in the five Romanianextiles using both techniques are evaluated in terms of period,rovenance and technique, and compared with earlier resultsbtained from similar objects [8–10].

. Experimental

.1. Historical samples

Fibers about 0.5 cm long were sampled from three religiousmbroideries and two brocaded velvets dated to the 17th and 18thenturies from the collection of the National Museum of Art, Roma-ia. Sample withdrawal was possible due to the fact that all thebjects had recently passed through the textile restoration work-hop for conservation.

.2. References

For each analytical technique, dedicated libraries of referencesere used. These databases were built up in the laboratories of

hree of the research group partners and consist of UV-visibleUV–vis), single stage and tandem MS data for the dye componentsiscussed [12,13].

.3. Sample preparation

Individual samples (about 0.5 mg) of dyed laboratory stan-ard or historic fibers were extracted with hydrochloric acid37%)/methanol/water (2:1:1, v/v/v) and prepared according tohe procedures described in detail in [12] for LC–DAD and [13]or LC–MS analysis. For green-coloured samples, where the pres-nce of indigoid dyes must be checked, an additional extractiontep was included. For this, after removing the coloured solutionesulting from hydrochloric acid extraction, 200 �L dimethyl for-amide (DMF) was added to the coloured fiber and the mixtureas heated at 140 ◦C for 10 minutes. The solution was then cen-

rifuged at 12,000 rpm for 5 minutes and the supernatant liquid wasransferred to an injection vial.

.4. Instrumentation

.4.1. LC-DADWaters LC–DAD equipment was used, data acquisition and

reatment being made by the Empower Pro 2002 software. Separa-ion was achieved on a LiChrosorb RP-18 column, 125 mm L × 4 mm.d. × 5 �m d.p. The mobile phase consisted of a mixture of methanolsolvent A), methanol in water (1/9, v/v) (solvent B) and an aqueous5%, v/v) solution of phosphoric acid (solvent C). Gradient elutionas applied according to the profile given below, which includes

he re-equilibration step.Time Solvent A (%) Solvent B (%) Solvent C (%)

0 23 67 103 23 67 10

29 90 0 1030 23 67 1035 23 67 10

The flow rate was set at 1.2 mL/min. The volume injected was

0 �L from which 5 �L will pass through the column for analysis.he other 15 �l is sent to the waste. Detection was made within a00–800 nm wavelength range, with a spectral resolution of 1.2 nm.hromatograms were integrated systematically at 254 nm and alsot one or more other wavelengths at which the optimum response

al Heritage 13 (2012) 89–97

of the dye constituent is observed. Results are presented as therelative composition of dyes at the wavelength(s) of integration.

2.4.2. LC-MSDLC–MS and LC–MS/MS experiments were achieved on a sys-

tem constructed from Agilent Series 1100 modules. Detectionwas made through a MS/MS ion trap detector using an electro-spray ionisation (ESI) ion source, operated in negative ion mode.The control of the chromatographic system and data acquisitionwere achieved with the Agilent ChemStation software LC 3D ver-sion 10.02, incorporating the MSD trap control, version 5.2, fromBrucker Daltronics. Separation was achieved on a Zorbax C18 col-umn, 150 mm L × 4.6 mm i.d. × 5 �m d.p., thermostated at 40 ◦C.The mobile phase consisted of a mixture of aqueous 0.2% (v/v)formic acid (solvent A) and methanol/acetonitrile (1:1 v/v, sol-vent B). Gradient elution was applied according to the profile givenbelow, which includes the re-equilibration step.

Time Solvent B (%)

0 155 25

10 5516 10018 10018.01 1522 15

The flow rate was set at 0.8 mL/min. The injected volume was5 �L (from a total of about 200 �L resulting from the samplepreparation). Several injections from the same solution may be per-formed, as described in the Results section below. The DAD detectorwas placed in series between the column and the MS ion source.UV–vis spectra were acquired over the 200–800 nm range with aresolution of 2 nm. MS detection was made in the negative ion modewith the following ESI operational parameters: drying gas tempera-ture 350 ◦C; drying gas flow rate 12 L/min; nebulising gas pressure65 psi; capillary high voltage 2484 V. The ion trap used a maxi-mum accumulation time of 300 ms and a total charge accumulation(ICC) of 30000. The multiplier voltage was set at 2000 V and thedynode potential at 7 kV. When working in the MS/MS mode, thespectral width was 4 a.m.u. and the collisional induced dissociationamplitude 1.6 V.

Automated Mass Spectral Deconvolution and Identification Sys-tem (AMDIS) software was used as a complementary identificationtool. Chromatograms obtained with full scan single stage massspectrometric detection were exported in the Agilent MS Engineformat (.ms) and analyzed with the AMDIS software [13].

3. Results and discussions

Samples described in Table 1 were analyzed by LC–DAD andLC–MS.

Fig. 1 shows the 17th-century Epimanikia (right hand sleeve,after cleaning) described in Table 1.

Table 2 summarizes DAD and MS data obtained for the sam-ples analyzed. For MS analysis, the detection of dyes was madeaccording to a dedicated analytical protocol, described in detail inan earlier publication [13]. It includes chromatographic separationand single MS full scan (FS) detection, followed by data process-ing through ion extraction chromatograms (IEC) according to m/zvalues of the molecular ions of dyes in the database. Results are cor-related with UV–vis spectral data. For minor compounds the samplewas re-injected using single stage MS detection in the selected ion

monitoring/multiple ion monitoring modes (SIM/MID) as well asby using tandem MS, product ion scan/single reaction monitoring(SRM)/multiple reaction monitoring (MRM), for unambiguous con-firmation of dye components. The confirmation procedure for thedye constituents (single MS-FS, single MS-SIM or tandem MS) may

I. Petroviciu et al. / Journal of Cultural Heritage 13 (2012) 89–97 91

Table 1Characterization of samples discussed in the present study.

Textile object (period) Function of the threads Color Sample code

Bedernita,Religious embroidery (1746)

Lining Green A1Sewing thread Pink yellow A12Embroidery thread Green A13Thread sewing the lining Kaki A14

Bedernita(lining), religious embroidery(1746)

Embroidery thread Kaki B15Sewing thread Green B17Sewing thread Brown B20

Epimanikia, Religious embroidery(17th c.)

Embroidery support (edge) Green D23aEmbroidery support (edge) Pink D23bEmbroidery support (edge) Pink yellow D23cLining Yellow D24Embroidery thread Orange D26Embroidery thread Red D27Embroidery thread Green D29

Sakkos,Brocaded velvet(17th c.)

Embroidery thread Pink yellow F37aEmbroidery thread Red F37bSilk core metallic embroidery thread Pink yellow F38

bt

dscueiMcd

3

ewrtcPI

FiM

Lining

Lining

Robe of Princess, brocaded velvet (16–17th c.) Weft

e directly correlated with the occurrence of the specific analyte inhe sample (Table 2).

When the semi-quantitative evaluation made using diode arrayetection (indicated through normalisation of a peak area to theum of the peak areas in the chromatogram) gives a result for theonstituent of interest greater than 10%, the FS operating mode issually sufficient for the MS detection. If the semi-quantitative DADvaluation for the constituent is between 1 and 10%, single stage MSn the SIM mode or MS/MS approaches are necessary for successful

S detection. When DAD detection indicates the occurrence of aompound at a level below 0.5%, it may be necessary to use AMDISeconvolution software for MS detection.

.1. Red dyes

Red dyes were detected in eight samples, four from religiousmbroideries and four from brocaded velvets. Anthraquinone dyesere present in all the cases where samples still have a visibly

ed color (3/8 samples). In one of these samples carminic acid,he main dye component in Mexican cochineal (Dactylopius coc-us Costa), Armenian cochineal (Porphyrophora hameli Brandt) andolish cochineal (Porphyrophora polonica L.), was detected by DAD.ts presence was also confirmed by MS in both cases. Three minor

ig. 1. Epimanikia described in Table 1. Religious embroidery (17th century) workedn the Byzantine tradition, preserved in the Art Collection Museum, National

useum of Art of Romania, inv. 88371.

Green F39Red F40

Green E34

compounds, dcII (the C-glucoside of flavokermesic acid [14,15]),kermesic and flavokermesic acids are also present in fibers dyedwith cochineal. In the present work, only dcII was detected by thepresence of its molecular ion (m/z = 475), according to the FS-IonExtracted Chromatogram (IEC). The presence of kermesic and fla-vokermesic acids was confirmed by the profiles of their fragmentsafter spectral deconvolution with AMDIS (Fig. 2).

Based on the calculation between the ratio of the minor com-pounds and carminic acid in HPLC-DAD analysis [16–18], thebiological source in F37b was established as Mexican cochineal(D. coccus Costa).

For two other red samples alizarin and purpurin, the mainanthraquinone constituents in madder (Rubia tinctorum L.),were detected by both DAD and MSD. The presence of minoranthraquinone compounds from madder, anthragallol, munjistin,xanthopurpurin and rubiadin was also established.

In five cases described as “pink” or “pink yellow” a marker com-pound for redwood (Caesalpinia spp.) dyeings, called “srw–solubleredwood” according to Wouters [4] or “type C” by Nowik [19] wasidentified by both DAD and MSD. For the latter, identification wasmade by FS-single stage MS followed by IEC of m/z = 243 a.m.u., themajor ion of “srw” according to the literature [13,20]. Confirmationof this identification was achieved by detection by single stage MSin the SIM mode and by MS/MS.

3.2. Yellow dyes

Flavonoid yellow dyes were detected in 15 out of a total of 20analyzed samples.

Dyer’s broom (Genista tinctoria L.) was identified as the mainbiological source in five samples and as a second biological source intwo other samples. In all the green samples blue “indigo” dyes (fromIsatis tinctoria L., Indigofera spp. or other species, discussed below)were also present. The identification of dyer’s broom was based onthe detection of luteolin, genistein and apigenin, by both LC–DADand MS detection techniques. In almost all cases, chrysoeriol – aminor compound recently identified in both weld and dyer’s broom

[13,15,21,22] – was also detected by single stage MS in SIM modeor by MS/MS.

Luteolin and apigenin – without the presence of genistein – wereidentified by both DAD and MSD in five samples, only one beingfrom brocaded velvet, a dyestuff constituent profile suggesting

92

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et al.

/ Journal

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13 (2012)

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Table 2Results obtained by alternative diode array and mass spectrometric (MS or MS/MS) detection modes.

Sample code Visual color Results Biological source

Dye constituentsDADa

Dye constituents MSDb Main source(s) Traces

A 1 Green 51 lu, 29 ge, 8 ap, 2 chry, 10 in lu(1), ge(1), ap(1), chry(2), in(2) Dyer’s broom and “indigo” –A 12 Yellow pink 77 srw, 7 qu, 1 kf,

7 rht, 2 rhz, 6 easrw(1), qu(2), kf(2), ea(2) Redwood and buckthorn berries Tannin plantc

A 13 Green 90 lu, 6 ap, 2 chry, 2 in lu(1), ap(1), in(2) Weld and “indigo”d

A 14 Kaki(yellow/green)

32 dat, 5 ge + lu, 4 kf, 1 irht, 1 ap, 23 in, 28 em, 5 chrys lu(1), ge(2), ap(2), dat(1,3), em(1), in(2) Bastard hemp, rhubarbe and “indigo” Dyer’s broom

B 15 Kaki 18 dat, 25 lu,1 ap, 19 in, 32 em,1 kf, 4 chrys

lu(1), ap(2), dat(1,3), em(1), kf(2), in(2) Bastard hemp, rhubarb, weld or eq. and “indigo” –

B 17 Green 43 lu, 43 ge, 10 ap, 1 chry, 3 in lu(1), ge(1), ap(1), chry(2), in(2), Dyer’s broom and “indigo” –B 20 Brown 92 lu, 8 ap lu(2) Weld (or another flavone-containg plant) –D 23 a Green 32 lu, 66 ge, 1 ap, +chry, 1 in lu(1), ge(1), ap(2), chry(3), in(2) Dyer’s broom and “indigo”D 23 b Pink 64.5 ca, 12 srw,

23 ea, +fk, 0.5 kasrw(1), ca(1,3), ea(2), fk(*), ka(*) (Mexican) cochineal, redwood and tannin plant –

D 23 c Pink yellow 90 srw, 1 lu, 9 ge srw(1), lu(2), ge(2), ap(3) Redwood Dyer’s broomD 24 Yellow 54 lu, 36 ge, 2 ap,

1 chry, 5 ea, 2 qu,+fi, +sul

lu(1), ge(1), ap(2), chry(2), fi(3), sul(3), ea(2,3), qu(3), Dyer’s broom and redwood Young fustic and qu based dye

D 26 Orange 35 fi, 56 sul, 8 ea, + kf fi(1,3), sul(1,3), ea(2) Young fusticD 27 Red 69 al, +xp, 25 pu, 1 ru, +ag, pu(1), ru(1), al(1), mu(1), xp(1), ag(2) Madder –D 29 Green 46 lu, 4 ap, 2 chry, 43 in, 4 ea, 0.5 fi, 0.5 sul lu(1), ap(2),

chry(2),fi(2), sul(2), in(2)

Weld and “indigo” Young fustic

F 37 a Yellow pink 68 srw, 14 fi, 15 sul, 3 ea srw(1,2), fi(2), sul(2), ea(2) Redwood, young fusticF 37 b Red 71.5 ca, 1.5 dcII, 1

fk, 0.5 ka, 25.5 ea,[288 nm: 0.8 dcII,97.7 ca, 1.5 fk + ka]

ca(1), dcII(2), fk(*), ka(*), ea(2) (Mexican) Cochineal and tannin plant –

F 38 Pink yellow 81 srw, 10 fi, 9 sul, +ea srw(1,3), fi(2),sul(2),ea(2)

Redwood, young fustic

F 39 Green 39 lu, 46 ge, 12 ap, 1 chry, 2 in lu(1),ge(1), ap(1), chry(2), in(2) Dyer’s broom and “indigo” –F 40 Red 87 al, 12 pu, +ag al(1), pu(1), ru(1), ag(2) Madder –E 34 Green 57 lu, 5 ap, 1 chry, 37 in lu(1), ap(2), chry(2), in(2) Weld and “indigo” –

The following abbreviations were used: al: alizarin; ag: anthragallol; ap: apigenin; ca: carminic acid; chrys: chrysophanic acid; chry: chrysoeriol; dat: datiscetin; ea: ellagic acid; em: emodin; fi: fisetin; fk: flavokermesic acid(also called laccaic acid D); dcII: flavokermesic acid, C-glucoside; ge: genistein; in: indigotin; irht: isorhamnetin; kf: kaempferol; ka: kermesic acid; laA: laccaic acid A; lu: luteolin; mu: munjistin; pu: purpurin; qu: quercetin;rht: rhamnetin; rhz: rhamnazin; ru: rubiadin; srw: soluble redwood; sul: sulfuretin; xp: xanthopurpurin.

a Numbers before the abbreviation for a dye represent the relative composition (%) corresponding to peak areas integrated in the chromatogram monitored at 255 nm (unless specified in the column); “ + ” indicates valueslower than 0.5%.

b Numbers between brackets have the following meaning: (1) detected through single stage MS–FS/IEC; (2) detected through MS–SIM/MID modes; (3) detected through MS/MS (MRM, product ion scan); (*) is used for dyesevidenced through deconvolution by AMDIS software.

c The term “tannin plant” is used as a shorter name for “tannin-producing plant” and is not indicative of a particular dye source.d “Indigo” refers to several plants that produce indigo.e The term “rhubarb” should be read as “rhubarb or another emodin-containing plant”.

I. Petroviciu et al. / Journal of Cultural Heritage 13 (2012) 89–97 93

Fig. 2. LC–MS and LC–MS/MS of sample F37b (red), where (Mexican) cochineal was identified. Upper image, from top to bottom, the UV–vis chromatogram; MS–FS, IEC forcarminic acid (molecular ion m/z = 491 and ion produced by decarboxylation in the source m/z = 447); IEC for dcII (molecular ion m/z = 475 and ion produced by decarboxylationin the source m/z = 431); IEC for ellagic acid. Lower images: detail of ion profiles after AMDIS deconvolution for flavokermesic (molecular ion m/z = 313 and ion produced bydecarboxylation in the source m/z = 269) and kermesic (molecular ion m/z = 329 and ion produced by decarboxylation in the source m/z = 285) acids. The figure illustrates theflexibility in use of mass spectrometry which allows the gradual detection of minor compounds.

94 I. Petroviciu et al. / Journal of Cultural Heritage 13 (2012) 89–97

F pectiva ne-conc ention

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ig. 3. LC–DAD chromatograms for sample B15, integrated at 255 and 350 nm, resnd chrysophanic acid, suggesting the use of bastard hemp, weld (or another flavoompound marked with “?” has similar UV spectrum to datiscetin, but different ret

he use of weld (Reseda luteola L.). However, recent studies haveemonstrated that other biological sources containing luteolin andpigenin, of which sawwort (Serratula tinctoria L.) is the mostell known [14,15], also exist and could have been used as tex-

ile dyes. As a consequence, only those samples where chrysoeriolas detected together with luteolin and apigenin are likely toave been dyed using weld; in the other cases the use of anotheravone-containing plant may not be excluded. When the pres-nce of weld (or another flavone-containing plant) correspondedo green-coloured fibers, “indigo” dyes were also detected, whilen another sample a combination of rhubarb (or another emodin-ontaining plant), bastard hemp and “indigo” dyes were found toave been used.

Fisetin and sulphuretin, the main dye components in young fus-ic (Cotinus coggygria L.) were detected in three samples withinhe present study. Young fustic was detected as a single biolog-cal source in one case and was identified together with solubleedwood in pink-yellow samples. Traces of young fustic were alsoetected in two additional samples where fisetin and sulphuretinere detected by single stage MS in SIM mode or by MS/MS product

on scan.Datiscetin, the main dye component in bastard hemp

Datisca cannabina L.) was identified by DAD in two kaki-oloured samples, in both cases together mainly with emodinnd with chrysophanic acid, kaempferol and isorhamnetin asinor constituents, as well as flavonoids from either dyer’s

room or weld (or another flavone-containing plant) andindigo” (Fig. 3). Poor chromatographic resolution is obtainedetween datiscetin and luteolin. Additionally, another com-ound exhibiting a very similar UV–vis spectrum to datiscetin,ut having increased retention, could be observed in thehromatogram.

Evidence for the presence of datiscetin (m/z = 285) in both sam-

les was based on the MS information collected from a standardample of wool dyed with bastard hemp. As no other dye waseported in this source, the identification of datiscetin by MS waslso confirmed by MS/MS analysis, based on the fragmentation pat-ern obtained through product ion scan. In LC–MS separations, as

ely, with detection of datiscetin, emodin, indigotin, luteolin, apigenin, kaempferoltaining plant; indigo/woad and rhubarb (or another emodin containing plant) the

time.

illustrated in Fig. 4, datiscetin co-elutes with kaempferol; however,comparison of the MS/MS product ion scan spectra allowed iden-tification of both datiscetin and kaempferol in the kaki-colouredsample B15.

Rhamnetin was identified by DAD together with quercetin,kaempferol and rhamnazin in an ochre-yellow sample (A12), sug-gesting the use of berries from a species of buckthorn (Rhamnusspp.). Analyses of two wool references, dyed with either the barkor the fruits (berries) from alder buckthorn (Rhamnus frangula L.)resulted in the detection of mainly emodin and minor amountsof rhamnetin and quercetin for the former, bark-dyedsample, andmainly rhamnetin, isorhamnetin and quercetin for the latter. Thecomponents detected in sample A12 by MS thus confirm the use ofberries from a Rhamnus species as the source of dye.

Emodin, the main dye component in alder buckthorn bark,rhubarb, yellow dock and other biological sources was also detectedin two samples. It was not possible to evidence the presence ofchrysophanic acid (m/z = 253) by MS, based on the IEC, nor by theidentification of the fragment m/z = 239, which would correspondto the fragment induced by de-methylation. A more detailed studyon a pure standard of chrysophanic acid (not available at the timeof the present experiments) is needed in order to be able to iden-tify it in historical samples. The limited information on these twoanthraquinone dyes of vegetable origin, emodin and chrysophanicacid, as textile dyes may be explained by their rare identification inhistorical textiles. Like emodin, the presence of chrysophanic acidmay indicate the use of alder buckthorn bark (Rhamnus frangulaL.), or the roots of rhubarb (Rheum sp.) or dock (Rumex sp.) [23–25].However, no systematic study in which the biological source of thedye has been identified unambiguously as one or other of theseplants has so far been reported.

3.3. Blue dyes

Although blue-coloured samples were not selected for analy-sis, indigotin was detected in eight green-coloured samples. Formass spectrometric detection, FS-single stage MS in SIM modeaccording to the molecular ion of indigotin (m/z = 261) was used.

I. Petroviciu et al. / Journal of Cultural Heritage 13 (2012) 89–97 95

F of dan C–MS)

Iam[p

ig. 4. LC–MS and LC–MS/MS data from sample B15, supporting the identificationot given; chrysophanic acid, although detected by LC–DAD, was not detected by L

ndigotin is the main dye component in woad, Isatis tinctoria L.nd in the indigo plant itself, Indigofera spp. but no analyticalethod has been reported to distinguish between these species

26]. The term “indigo” is thus used to refer to indigotin-producinglants.

tiscetin, emodin, luteolin, apigenin, and kaempferol (identification for indigotin is.

3.4. Tannin

Ellagic acid was detected in seven samples with both detec-tion techniques. The presence of ellagic acid indicates the use ofa tannin-containing plant material either for textile dyeing or for

96 I. Petroviciu et al. / Journal of Cultural Heritage 13 (2012) 89–97

Table 3Biological sources attributed to samples analyzed throughout the present study, compared to other results for samples from religious embroideries and brocaded velvetsfrom Romanian collections previously identified through the LC–DAD method by the same research group [8–10].

Religious embroideries Brocaded velvets

15th-16th c. 17th–18th c. 19th c. 15th–16th c. 17th c.

Present Previous Present Previous

CochinealAll 6 1 3 1 2 1 0(DC) (3)–16th c. (1) (1) (0) (0) (1) 0

Kermes 3 0 0 0 1 0 0Lac dye 21 0 0 0 0 0 0Madder 29 1 3 0 0 1 0Safflower 2 0 1 0 0 0 0Redwood 24 3 4 3 4 2 0Young fustic 24 1 1 3 3 2 1Weld (or another flavone-containing plant) 26 4 3 4 7 1 0Dyer’s broom 8 4 0 1 0 1 2Buckthorn berries 0 1 3 2 0 0 0Rhubarb (or another emodin-containing plant) 2 2 2 1 0 0 0Bastard hemp 1 2 2 0 0 0 0Tannins 39 1 5 2 6 0 3

wetlwiiTi

3rR

fisfirsa1

hr(tpwdwliiuf

caamt

Logwood 0 0

Indigoid 40 5

eighting silk [4,24]. In the present study, a high proportion ofllagic acid from a tannin-producing plant source was detectedwice in red samples in combination with cochineal while muchower proportions were detected in a yellow pink sample together

ith redwood and buckthorn berries. Ellagic acid was also detectedn all the five cases when fisetin and sulphuretin were identified,ndicating the use of the dye from the heartwood of young fustic.his is probably due to the presence of a trace of the tannins presentn young fustic (primarily in the leaves and twigs).

.5. Discussion on the biological sources together with previousesults on religious embroideries and brocaded velvets fromomanian collections

Attribution of the biological sources of dyes identified and con-rmed in the analyzed samples is summarized in Table 2. All theources detected in the present series of analyses were also identi-ed in one or more groups of samples analyzed before by the sameesearch team. According to the studies performed until now, sixources of red dye have been identified in religious embroideriesnd brocaded velvets from Romanian collections dating from the5th to the 19th centuries (Table 3).

Half of these are of animal origin (cochineal, kermes and lac) andalf derive from plant sources, and from various parts of the plants:oots (madder), petals (safflower) and wood (redwood). Cochinealboth Old and New World) and redwood were detected in all theextile groups studied; lac dye, madder and safflower were onlyresent in embroideries. The combination of lac dye and madderas the main source of red used in the support fabric for embroi-eries in the 15th and 16th centuries. According to literature, lacas hardly used in Europe for textile dyeing, but only for dyeing

eather and as an organic pigment. It was mentioned as importednto the Ottoman Empire as early as the 15th century and, accord-ng to literature, it has been detected in Ottoman textiles [27,28]. Itsse in religious embroideries would thus suggest an Oriental originor these materials.

Kermes was only identified in religious embroideries and bro-

aded velvets dated to the 15th and 16th centuries, which is inccordance with literature mentioning that, due to its lower costnd ease of use Mexican cochineal eventually became the only ani-al source of red dye used in Europe, soon after the discovery of

he New World [25].

1 1 0 0 07 5 3 2 1

Six sources of yellow flavonoids were identified in the series,weld (or another flavone-containing plant), young fustic and dyer’sbroom being the most widely used. Except for the latter that wasnot present in 15th–16th century embroideries, the others weredetected in all the groups considered. Bastard hemp, buckthornberries and rhubarb (or another emodin-containing plant) wereonly detected in embroideries, bastard hemp up to the 18th cen-tury, buckthorn berries not before the 17th century and rhubarb (oranother emodin-containing plant source) in textiles from the 15thto the 18th century. However, both bastard hemp and rhubarb (oranother emodin-containing plant) were only detected in Ottomantextiles [28], which also supports the suggestion of an Orientalorigin for the materials.

As far as the blue dyes are concerned, “indigo” was identified inreligious embroideries and brocaded velvets from the 15th to the19th century, while logwood was only present in 17th- to 19th-century embroideries. This is no surprise when we remember thatthis source was only available after the discovery of the New World[29].

4. Conclusions

All the dyes identified on the 17th- and 18th-century Roma-nian velvets and embroideries are in very good correlation with theexisting knowledge on dyes and biological sources used in Europeand Asia Minor during this period [1,2,5,6,24,25,28]. The resultsmay be also correlated in terms of period and fiber function withprevious data obtained for similar textiles by the same researchteam [8–10]. From the whole group of analyses performed up tillthe present on textiles from Romanian collections, it may be con-cluded that the combination of lac dye and madder was the mainsource of red in the 15th- to 16th-century embroideries, while ker-mes was used when a more precious textile was intended. Mexicancochineal was preferred in later textiles. Lac dye, bastard hemp andrhubarb (or another emodin-containing plant) were only detectedin embroideries. Based on this fact and considering that these dyesources were not identified in textiles from Europe, but only in

Ottoman pieces, it may be stated that at least part of the materialsused in embroideries dating from the 15th to the 18th centurieshave an Oriental origin.

As far as the techniques applied are concerned, it can be con-cluded that a good correlation between the results produced

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y LC–DAD, LC–MS and LC–MS/MS was found, not only for theajor dye components, but also for the minor accompanying con-

tituents. For some of the minor components co-eluting under thehromatographic conditions of the LC–MS method, spectral decon-olution with AMDIS software was established. The study showedhat LC–MS and LC–MS/MS approaches were confirmed as versa-ile tools for the identification and confirmation of dyes in historicextiles, if consistent work is first accumulated on a collection oftandard dyes and dyed fibers for construction of suitable spectralibraries.

cknowledgements

The authors would like to thank the National Museum of Art ofomania and the Putna Monastery, Romania for access to their col-

ections. They are also grateful to LaborMed Pharma, who offeredn open access to the LC–MS/MS analytical instrumentation and tos Marie-Christine Maquoi from the KIK/IRPA laboratory for the

xpert assistance in LC–DAD dye analysis. Professor Recep Karadagrom Marmara University, Istanbul, Turkey, who offered a bastardemp-dyed fibre, is also acknowledged. The authors are also grate-

ul to Ms. Jo Kirby, National Gallery London Scientific Departmentretired), for carefully reading and improving the English text.

eferences

[1] J.H. Hofenk de Graaff, W.G. Th Roelofs, On the occurrence of red dyestuffs intextile materials from the period 1450–1600, in: 4th Meeting ICOM Committeefor Conservation, Madrid, 1972.

[2] J.H. Hofenk de Graaff, T.W.G. Roelofs, The analysis of flavonoids in natural yel-low dyestuffs occuring in ancient textiles, in: 5th Meeting ICOM Committee forConservation, Zagreb, 1978, p. 1–15.

[3] J. Wouters, Analyse des colorants des tapisseries brugeoises, in: Bruges et latapisserie des xvie et xviie siècles, Mouscron, Bruges, 1987, p. 515–526.

[4] J. Wouters, Dye analysis of Florentine borders of the 14th to 16th centuries,Dyes in History and Archaeology 14 (1995) 48–58.

[5] I. Karapanagiotis, L. Valianou, Y. Sister Daniila, Chryssoulakis, Organic dyes inByzantine and post-Byzantine icons from Chalkidiki (Greece), Journal of Cul-tural Heritage 8 (2007) 294–298.

[6] M. Van Bommel, J.H. Hofenk de Graaff, Master dyers to the court of Sicily, Dyesin History and Archaeology, 22/23, publication due 2012.

[7] I. Petroviciu, J. Wouters, Analysis of natural dyes from Romanian 19th and 20th

century ethnographical textiles by DAD-HPLC, Dyes in History and Archaeology18 (2002) 57–62.

[8] I. Petroviciu, J. Wouters, I. Vanden Berghe, I. Cretu, Dyes in some textiles fromthe Romanian Medieval Art Gallery, Dyes in History and Archaeology 22/23,publication due 2012.

[

[

al Heritage 13 (2012) 89–97 97

[9] I. Petroviciu, J. Wouters, I. Vanden Berghe, I. Cretu, Dye analysis on some 15thCentury byzantine embroideries, Dyes in History and Archaeology. 22/23, pub-lication due 2012.

10] I. Petroviciu, I. Vanden Berghe, I. Cretu, J. Wouters, Analysis of Dyestuffs in15th–17th Century byzantine embroideries from Putna Monastery, Romania,Dyes in History and Archaeology, 24/25, publication due 2012.

11] M. Trojanowicz, J. Orska-Gawrys, I. Surowiec, B. Szostek, Chromatographicinvestigation of dyes extracted from Coptic texiles from the National Museumin Warsaw, Studies in Conservation 49 (2004) 115–130.

12] J. Wouters, N. Rosario-Chirinos, Dye analysis of pre-combian peruvian textileswith HPLC and DAD, Journal of the American Institute for Conservation 31(1992) 237–255.

13] I. Petroviciu, F. Albu, A. Medvedovici, LC/MS and LC/MS/MS based protocolfor identification of dyes in historic textiles, Microchemical Journal 95 (2010)247–254.

14] D. Peggie, The development and application of analytical methods for the iden-tification of dyes on historical textiles, PhD thesis, University of Edinburgh(2006).

15] D. Peggie, A. Hulme, Mc. Nab, H.A. Quye, Towards the identification of charac-teristic minor components from textiles dyed with weld (Reseda luteola L.) andthose dyed with Mexican cochineal (Dactylopius coccus Costa), MicrochemicaActa 162 (2008) 371–380.

16] J. Wouters, A. Verhecken, The Coccid insect dyes: HPLC and computerized anal-ysis of dyed yarns, Studies in Conservation 34 (1989) 189–200.

17] J. Wouters, A. Verhecken, The scale insect dyes (Homoptera:Coccoidea) speciesrecognition by HPLC and Diode-Array analysis of the dyestuffs, Annales de laSociété entomologique de France 25 (1989) 393–410.

18] I. Vanden Berghe, Investigation of cochineal dyeing parameters and refinementof the cochineal identification system’, Dyes in History and Archaeology22/23,publication due 2012.

19] W. Nowik, The possibility of differentiation and identification of red and blue‘soluble’ dyewoods: determination of species used in dyeing and chemistry oftheir dyestuffs, Dyes in History and Archaeology 16/17 (2001) 129–144.

20] I. Karapanagiotis, E. Minopoulou, L. Valianou, Y. Sister Daniilia, Chryssoulakis,Investigation of the colourants used in icons of the Cretan school of iconogra-phy, Analytica Chimica Acta 647 (2009) 231–242.

21] E. Ferreira, New approaches towards the identification of yellow dyes in 463ancient textiles, PhD thesis, University of Edinburgh (2001).

22] http://www.organic-colorants.org.23] H. Schweppe, Handbuch der Naturfarbstoffe, Vorkommen, Verwendung, Nach-

weis, Nikol Verlagsgesellschaft mbH&Co, KG, Hamburg, 1993, p. 224–228.24] J.H. Hofenk de Graaff, The colourful past. Origins chemistry and identification

of natural dyestuffs, Abegg Stiftung & Archetype Publications Ltd, Riggisbergand London, 2004.

25] D. Cardon, Natural dyes–sources, tradition, technology science, Archetype Pub-lications, London, 2007.

26] J. Wouters, Possible future developments in the analysis of organic dyes, Dyesin History and Archaeology 20 (2005) 23–29.

27] I. Vanden Berghe, M.C. Maquoi, J. Wouters, Dye analysis of Ottoman silk, in: M.van Raemdonck (Ed.), The Ottoman silk textiles from the Royal Museums of Art

and History in Brussels, Turhout Brepols, Brussels, 2004, pp. 49–60.

28] N. Enez, H. Böhmer, Ottoman textiles: dye analysis, results and interpretation,Dyes in History and Archaeology 14 (1995) 39–44.

29] F. Brunello, The art of dyeing in the History of Mankind, Vicenza, 1973,p. 175–221.