Nakamura, George R.

Shimoda, Satoru C.

English

Northwestern University Illinois, School of Law: Scholarly Commons

Journal of Criminal Law and CriminologyVolume 56Issue 1 March Article 15Spring 1965Examination of Micro-Quantity of Ball Point Inksfrom Documents by Thin-Layer ChromatographyGeorge R. NakamuraSatoru C. ShimodaFollow this and additional works at: https://scholarlycommons.law.northwestern.edu/jclcPart of the Criminal Law Commons, Criminology Commons, and the Criminology and CriminalJustice CommonsThis Criminology is brought to you for free and open access by Northwestern University School of Law Scholarly Commons. It has been accepted forinclusion in Journal of Criminal Law and Criminology by an authorized editor of Northwestern University School of Law Scholarly Commons.Recommended CitationGeorge R. Nakamura, Satoru C. Shimoda, Examination of Micro-Quantity of Ball Point Inks from Documents by Thin-LayerChromatography, 56 J. Crim. L. Criminology & Police Sci. 113 (1965)EXAMINATION OF MICRO-QUANTITY OF BALL POINT INKS FROMDOCUMENTS BY THIN-LAYER CHROMATOGRAPHYGEORGE R. NAKAMURA AND SATORU C. SHIMODAGeorge R. Nakamura is chief chemist, Military Police Crime Laboratory, United States Army,Japan. Dr. Nakamura received his masters degree from the University of Illinois and a doctor ofpharmaceutical sciences from the University of Tokyo in 1963. Prior to his present military assign-ment, he was a biochemist in the Army Medical Laboratory. Dr. Nakamura has published severalarticles in leading scientific journals.Satoru C. Shimoda is a Chief Warrant Officer, Military Police Corps, U. S. Army and documentexaminer with the Military Police Crime Laboratory in Japan. Mr. Shimoda has been previouslyassigned to the Military Police Crime Laboratory at Fort Gordon, Georgia and at their laboratoryin Germany. He has been active in the field of questioned document examination for over ten yearsand has appeared as an expert witness in numerous military courts. He presented a preliminary re-port on this present study at the 1964 annual meeting of the American Society of Questioned Docu-ment Examiners in Denver.-EDIToR.For the document examiner, the crux in anymethod for the examination and identification ofink from a written line has been the limited amountof sample which could be obtained without deface-ment and damage to the document. Documentexaminers have sought a simple method that wouldafford the best possible resolution of componentsfor identification, high sensitivity, accuracy, speed,and convenience.A number of techniques for removing ink fromwritings and separating dye components haveheretofore been described. Paper chromatographicmethods (1, 2, 3, 4) have generally been used forseparating dyes. Although satisfactory resultshave been obtained from paper chromatography,some preliminary results obtained from thin-layerchromatography (TLC) has revealed that theresolution pattern of dye components are defi-nitely superior. It was apparent at the outset inexperimenting with TLC, that this method hadan inherent ability to produce dye spots whichwere more compact and their separation morediscrete than those yielded by paper chroma-tography (Fig. 1).Since the amount of ink which is removed froma writing is extremely small, TLC presented atonce an opportunity for characterizing such amicroquantity of sample. Its possible applicationin ink analysis was described by Doud (5) in1958 and more recently by Witte (6). A solventwas sought in this Laboratory which wouldenable a quick separation of dye components on amicroscope slide layered with silica gel prepara-tion. While no all-purpose solvent mixture couldbe formulated for all ball pen inks, it was decidedto standardize only one solvent mixture to simplifythe procedure and to eliminate haphazard guessesas to which formula to use on a small amount ofunknown ink available for examination. Since theamount of ink sample removed from a writing waslimited, the chromatographic run necessarily hadto be short with the best possible resolution ofdyes at lower Rf values.A number of techniques were tested for collect-ing ink samples from writing including those whichentailed scraping (6), bleeding (3, 5), and punching(7) for application onto a chromatogram. Onemust be reconciled to some damage to the docu-ment upon removal of ink sample, however thelatter technique of removing a minute disc froma written line with a hypodermic needle left aclean line without unsightly smears and tearsalong the sampling source. The methods for com-paring inks as to their chromatographic char-acteristics and for spot testing the componentdyes are described in the following paragraphs.METHODChromatoslides. Chromatography plates areprepared from 1" x 3" glass microscope slides (8).A large number of plates can be layered simul-taneously using a commercial spreading apparatusand then stored in a desiccator for future use.However, if only a few slides are required in asingle test, chromatoplates can be made rapidlyG. R. NAKAMURA AND S. C. SHIMODAFIGURE 1Comparison of a thin-layer chromatoplate (top) and a paper chromatogram (bottom) using the same solventmixture (1-butanol: 50, ethanol: 10, water: 15) on the same group of blue ball pen inks.Other solvent systems described in (1, 2, 3, 4) improved the resolution of dyes by paper chromatography butthey did not afford nearly the separation exhibited by TLC as in Figure lb. Legend: (4) Zebra. (2) Tombow Crown.(19) Eversharp KEC. (18) Wearever. (16) Eversharp. (9) Script. (5) Ballograf. (3) Ballcon.by dipping slides into a small jar containing 10grams of "Wakogel B-5" (Silica gel powder con-taining 5% calcium sulfate as binder, Wako PureChemical Industries, Osaka, Japan) suspended in38 ml of chloroform. Layer thickness can be meas-ured with a micrometer ordinarily used for measur-ing the thickness of papers. Optimum thickness forTLC of ink in this method was found to be ap-proximately 250 microns. Typical "microchromato-plates" showing separation of ball point ink dyesare presented in Figure 2.Sampling. Using a hypodermic syringe needlewith its point removed and resharpened (7), sizes17 to 20, depending on the thickness of the writingstroke, punctures are made along the written lineespecially at junctures showing a clot of ink.Three or four discs are removed, with the aid of aspatula or razor blade if necessary, and then trans-ferred to a well of a spot plate. An amount ofpyridine sufficient to cover the discs is introducedto extract the ink. The process can be speeded byprodding the discs with the tapered point of adrawn glass rod. The paper fragments are movedto the edge of the well, and the extract is evap-orated to near-dryness. Then the entire ink ex-tract is removed by a capillary tube having aninside diameter of about 0.5 mm and depositedonto a chromatoslide, effecting a spot about 2mm in diameter and about 15 mm from the bottomedge of the slide. The spot should be dried toremove the pyridine prior to chromatography;a hot air blower facilitates this step.Chromatography. The chromatoplate bearingthe unknown ink spot and a standard ink spot forcomparison is placed in a small jar such as thatused commercially for pickles, jellies, etc. Anamount of solvent mixture is placed in this jar toimmerse the end of the chromatoplate withouttouching the applied ink spot. The solvent mixtureis comprised of 50 parts of 1-butanol, 10 parts ofabsolute ethanol, and 15 parts of water. Thechromatographic run is completed within twenty[Vol. 56THIN-LAYER CHROMATOGRAPHY OF BALL PEN INKSFrGuR 2Micro-chromatoplates showing separation of two dif-ferent ball pen inks. The ink was removed from a writ-ing on a document by the punch method utilizing a hy-podermic needle.minutes. The slides are removed and air dried,and may later be photographically recorded.Spot Tests. With the aid of a pointed instrument,four or more small portions of each separated dyeis removed from the chromatoplate and trans-fered to wells of a spot plate. To the four wellscontaining particles of dye-impregnated silica gel,concentrated HCl, concentrated H2SO 4, NH4OH,and 10% NaOH are introduced in small drops froma capillary pipette to the respective wells. Colorchanges are noted, then a drop or more of a satu-rated sodium carbonate solution is added to the"HCI" well to observe any further change; manydyes undergo an indicator type shift back to theoriginal color due to change in pH (2). Solubilityof the separated dyes, then, are tested in water,absolute ethanol, acetone, and chloroform in fouradditional wells. Agitation is provided with athin glass rod or a capillary tube having a sealedtip. Observation under a microscope set at fourto ten power is an aid to these micro-spot tests.For reference, Table I lists some of the commondyes distributed by major chemical and dye manu-facturers in the United States for the manufactureof colored writing inks and their immediate re-actions to these test reagents and their responsesto ultraviolet and infrared lights. This list ofdyestuffs is not to be construed as being completefor ball pen ink dyes and is presented only toserve as a guide. For presumptive identification,an actual comparison examination with a standarddye sample would be indispensable.DIscussIoNOstensibly, it seemed forensically advantageousto identify the dye components of ink instead ofcharacterizing ink by commercial brands, inasmuchas manufacturers change formulation frequentlywithout notice. Some sixty or more commerciallyprepared dyestuffs which are used in the manu-facture of colored inks were tested on our chromato-plates. While most of them were chromatographi-cally pure compounds, some of them were shownto be mixtures containing presumably isomersand/or unreacted constituents as has already beensuggested by Coldwell (1) in his discussion of dyeidentification. For this reason, direct spot testingof some of these dyes was valueless for our purposeand the principal spot from each separated dyepreparation on our chromatoplates was tested withsolvents and reagents listed in (1), (2), and (3).While a presumptive identification can be madeof some of the more common dyes used in themanufacture of ball pen ink, the primary value ofspot testing must remain in the comparison of aquestioned ink sample from that of a standard.The examination of blue ink samples from popularAmerican and Japanese ball point pens selected atrandom (e.g.: Playmate, Parker, Script, Pilot,Mitsubishi, etc.) revealed that the dye formula-tion employed in the manufacture of ball pen inkis not too diverse, and the TLC method andspot testing confirmed that such dyes as methylviolet, rhodamine-B, Victoria blue, phalocyanineblue, and alkali blue are of common occurrence.While only a few red dyes exhibited fluorescencein ultraviolet light, the marked difference particu-larly shown by eosin and rhodamine furnished anadded criterion for their identification. Nearlyall of the dyes were made invisible by infraredphotography, hence differentiation of these com-pounds by this procedure was not of great value.Black ink containing nigrosine or indulin type ofdye, was difficult to resolve in any of the manysolvent mixtures tested; the dye manifested it-self on the plates as streaks and not as spots. Suchdyes, therefore, were not listed in Table I. Theformulation of dyes in fluid inks, as they were char-acterized by this TLC method using the standardsolvent mixture and those using 1-butanol:50,G. R. NAKAMUR-4 AND S. C. SHIMODALU)t4,to too PCto" , .0 .0 ' .0W to0o >,. :a 4.; to to to> > ~. > > I ->..0.0.w0'A 'n0. 0 0 W>>.-; .CD Q '0 o- V n00' 4 0a4-. m, 9-p 490 -0 -Z "- ~ ~ do/ -q 9z Q'4: : : : 0U *c : :* :Z "0'44 : : : :zo==z :. :_' o :U)U U) 0 . U 2 -- .~000ZZ t'~U,)2-=• = = = c. s: -r .0;4 .g -P4 t Z u.4 w E[Vol. 56.x 0 0 opq . 1-.2-M M00Q0*" A :4,4.4 L.1965] THIN-LA YER CHROMATOGRAPHY OF BALL PEN IANKS0 ~0 0 0 r.4; ~ ~ b 0 00 00 1: . 2 2 0 0 I I II I --'--" -'1- I I I I I I 11 I 1 I I I I II I o--lI I I1 I II I I IIIIIII I I Ii l l l i i, I IlI I II I+..0 -3-.-- --. --, - -- -  - -....o00o .. . 00.0 00 .. o o o o o ... .. 0. 0. 0 o BE0)P4=-3-. : .-. . 3..: : : : : : : : :'n±a maw.: ... : ,. a .3m m a m ...... . . . .. .. .-.0'a~~~~~~. to t .ib a 0-am .. . ... a a ,a- a.. . a :a ma ao a m. ao a a ma0~~~~''3 >- -0 .0t -00 0-- t 1))- . - fJ.ovJ) '- - ; " - Z .= , = . . -, , 'to w w w Pto to to to -o 0 > >- > > >>.>>.00 ~ 00 0 " 0 0. 0)00M0.0'0 0 M) M w0 In -n w ._ w In) -- w 0 0 w 0 t 0 wVr- t- .. t- t- aa '0 '...0 C4. - ...0. 5 - t- .0q c-* r- C-eq 0oG. R. NAKAMURA AND S. C. SHIMODA10% oxalic acid:15, absolute ethanol:10; and1-butanol:50, 6 N ammonium hydroxide:15,absolute ethanol:t0, clearly revealed that themore popular blue "fountain pen" inks such asParker, Script, Pilot, etc., contained seven ormore blue dye components and in some cases redand green dyes.Cartridge inks of various popular American andJapanese brands were found to contain, by thepresent TLC method, the same dyes as theirbottled ink counterparts. For example, Parker blueQuink in both bottle and cartridge form had thesame dye components.In testing fluid inks by TLC, there was somedifficulty in completely removing the ink fromdiscs punctured from an ink line. A number ofsolvents including adds, alkali, and salts includingpotassium binoxalate (4) were tested, but noneafforded complete removal; some residual ink,presumably pigments, remained in the fibers.Fluid ink also had a tendency of penetrating intothe silica gel layer leaving only a light spot on thesurface. These losses seriously hampered micro-application such as described here for ball penink. Further study is being conducted to establisha micro-application method for fluid inks.A number of commercial silica gel preparationswere tested, and it was found that the amount ofbinder substance, calcium sulfate, had an ap-preciable effect on the resolution of ink dyes. Themore commonly used "Silica Gel-G" containing13 % calcium sulfate, although offering comparableresults, did not resolve some of the ball point inksamples in the manner effected by "Wakoge! B-5"which has a 5% calcium sulfate inclusion. A silicagel preparation containing no binder, "WakogelB-0", offered results comparable to those of"Wakogel B-5" indicating that the amount ofcalcium sulfate does affect resolution of ink dyeson TLC.The advantage of the TLC method is in thehigh resolution of dyestuffs as compared with thatof paper chromatography, for example, methyl vio-let will resolve characteristically into a formationof three or four spots (Fig. 2) strung like violetbeads. None of the paper chromatography systems(1, 3, 4, 8) which we have tested make this char-acterization: The separation is sufficiently discreteto allow examiners to remove small portions fromdye spots for micro-tests. Also, this procedureprecludes having to bleed the ink writing withacid directly on the document for dye identifi-cation such as done by Crown, et al (2). This willprevent further damage to the writing and henceto the document.A thin-layer chromatography system is de-scribed for characterizing small amounts of blueand blue-black ball point inks collected fromwritings. A procedure is described for removingportions of the separated dyes for further char-acterization by spot testing. A table listing com-mercial dyestuffs used in the manufacture ofcolored writing inks, their Rf values, solubilityand color reaction in acids and bases are reported.Acknowledgment: The authors are indebted toSp/7 Gerald A. Boardman for his assistance incolor, ultraviolet and infrared photography andto Koichi Funakoshi and Seisaburo Miyamoto fortheir technical assistance in chromatography andspot testing. To General Aniline and Film Corp.,National Aniline Division, Allied Chemical Corp.,Dupont DeNemour Co., and Pilot Pen Co., Japan,for their aid, co-operation and generous contribu-tion of their dyes.REFERENCES1. CoLrwELL, B. B., "The Comparison of Inks andWritings by Paper Chromatography". THE ANA-LYST, 80, 68 (1955).2. Caoww, D. A., CONWAY, J. V. P., AND Krnx, P. L.,"Differentiation of Blue Ball Point Pen Ink".J. CRIMINAL LAW, CRIMNOL., AND POLICE SCL,52, 338 (1961).3. B Arcxxrr, JR. J. W., AND BA A'oRn, L. W., "Com-parison of Ink Writing On Documents By Meansof Paper Chromatography". J. CRaIM LAW,CRImnNOL., AND POLICE Sci., 43, 530 (1952).4. MATHY.R, J., "Comparison of Inks by Paper Chro-matography, Practical Methods". INTERNATIONALCR AI. POLICE RE iEW, 148, 130 (1961).5. DOUD, D., "Chromatographic Analysis of Inks". J.oF FoRsENSIC SciEicEs, 3, 486 (1958).6. Wirr, A. H., "The Examination and Identificationof Inks". METHODs OF FORENSIC SCIENCE, Vol..II, 35, (1963), Interscience Publishers, London;New York.7. HARRISON, W. R., SUSPECT DocumaT'rs, THEIRScIENTI FIC EX.AmINATfON. Frederick A. Praeger,Publisher, New York, 1958.8. Pairxa, J. J., "Rapid and Simplified Method ofAnalysis By Thin-Layer Chromatography UsingMichrochromatoplates". MICROCHIMICA AcrA, 3,529 (1962).[Vol. 56

Examination of Micro-Quantity of Ball Point Inks from Documents by Thin-Layer Chromatography

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Examination of Micro-Quantity of Ball Point Inks from Documents by Thin-Layer Chromatography

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