Journal of Criminal Law and Criminology Volume 56 Article 15 Issue 1 March

Spring 1965 Examination of Micro-Quantity of Ball Point Inks from Documents by Thin-Layer Chromatography George R. Nakamura

Satoru C. Shimoda

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Recommended Citation George R. Nakamura, Satoru C. Shimoda, Examination of Micro-Quantity of Ball Point Inks from Documents by Thin-Layer Chromatography, 56 J. Crim. L. Criminology & Police Sci. 113 (1965)

This Criminology is brought to you for free and open access by Northwestern University School of Law Scholarly Commons. It has been accepted for inclusion in Journal of Criminal Law and Criminology by an authorized editor of Northwestern University School of Law Scholarly Commons. EXAMINATION OF MICRO-QUANTITY OF BALL POINT INKS FROM DOCUMENTS BY THIN-LAYER CHROMATOGRAPHY

GEORGE R. NAKAMURA AND SATORU C. SHIMODA

George 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 of pharmaceutical 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 several articles in leading scientific journals. Satoru C. Shimoda is a Chief Warrant Officer, Military Police Corps, U. S. Army and document examiner with the Military Police Crime Laboratory in Japan. Mr. Shimoda has been previously assigned to the Military Police Crime Laboratory at Fort Gordon, Georgia and at their laboratory in Germany. He has been active in the field of questioned document examination for over ten years and 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 any tion. While no all-purpose solvent mixture could method for the examination and identification of be formulated for all ball inks, it was decided ink from a written line has been the limited amount to standardize only one solvent mixture to simplify of sample which could be obtained without deface- the procedure and to eliminate haphazard guesses ment and damage to the document. Document as to which formula to use on a small amount of examiners have sought a simple method that would unknown ink available for examination. Since the afford the best possible resolution of components amount of ink sample removed from a writing was for identification, high sensitivity, accuracy, speed, limited, the chromatographic run necessarily had and convenience. to be short with the best possible resolution of A number of techniques for removing ink from dyes at lower Rf values. writings and separating dye components have A number of techniques were tested for collect- heretofore been described. Paper chromatographic ing ink samples from writing including those which methods (1, 2, 3, 4) have generally been used for entailed scraping (6), bleeding (3, 5), and punching separating dyes. Although satisfactory results (7) for application onto a chromatogram. One have been obtained from paper chromatography, must be reconciled to some damage to the docu- some preliminary results obtained from thin-layer ment upon removal of ink sample, however the chromatography (TLC) has revealed that the latter technique of removing a minute disc from resolution pattern of dye components are defi- a written line with a hypodermic needle left a nitely superior. It was apparent at the outset in clean line without unsightly smears and tears experimenting with TLC, that this method had along the sampling source. The methods for com- an inherent ability to produce dye spots which paring inks as to their chromatographic char- were more compact and their separation more acteristics and for spot testing the component discrete than those yielded by paper chroma- dyes are described in the following paragraphs. tography (Fig. 1). Since the amount of ink which is removed from METHOD a writing is extremely small, TLC presented at once an opportunity for characterizing such a Chromatoslides. Chromatography plates are microquantity of sample. Its possible application prepared from 1" x 3" glass microscope slides (8). in ink analysis was described by Doud (5) in A large number of plates can be layered simul- 1958 and more recently by Witte (6). A solvent taneously using a commercial spreading apparatus was sought in this Laboratory which would and then stored in a desiccator for future use. enable a quick separation of dye components on a However, if only a few slides are required in a microscope slide layered with silica gel prepara- single test, chromatoplates can be made rapidly G. R. NAKAMURA AND S. C. SHIMODA [Vol. 56

FIGURE 1 Comparison of a thin-layer chromatoplate (top) and a paper chromatogram (bottom) using the same solvent mixture (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 but they did not afford nearly the separation exhibited by TLC as in Figure lb. Legend: (4) . (2) Crown. (19) KEC. (18) Wearever. (16) Eversharp. (9) Script. (5) . (3) Ballcon. by dipping slides into a small jar containing 10 prodding the discs with the tapered point of a grams of "Wakogel B-5" (Silica gel powder con- drawn glass rod. The paper fragments are moved taining 5% calcium sulfate as binder, Wako Pure to the edge of the well, and the extract is evap- Chemical Industries, Osaka, Japan) suspended in orated to near-dryness. Then the entire ink ex- 38 ml of chloroform. Layer thickness can be meas- tract is removed by a capillary tube having an ured with a micrometer ordinarily used for measur- inside diameter of about 0.5 mm and deposited ing the thickness of papers. Optimum thickness for onto a chromatoslide, effecting a spot about 2 TLC of ink in this method was found to be ap- mm in diameter and about 15 mm from the bottom proximately 250 microns. Typical "microchromato- edge of the slide. The spot should be dried to plates" showing separation of ball point ink dyes remove the pyridine prior to chromatography; are presented in Figure 2. a hot air blower facilitates this step. Sampling. Using a hypodermic syringe needle Chromatography. The chromatoplate bearing with its point removed and resharpened (7), sizes the unknown ink spot and a standard ink spot for 17 to 20, depending on the thickness of the writing comparison is placed in a small jar such as that stroke, punctures are made along the written line used commercially for pickles, jellies, etc. An especially at junctures showing a clot of ink. amount of solvent mixture is placed in this jar to Three or four discs are removed, with the aid of a immerse the end of the chromatoplate without spatula or razor blade if necessary, and then trans- touching the applied ink spot. The solvent mixture ferred to a well of a spot plate. An amount of is comprised of 50 parts of 1-butanol, 10 parts of pyridine sufficient to cover the discs is introduced absolute ethanol, and 15 parts of water. The to extract the ink. The process can be speeded by chromatographic run is completed within twenty THIN-LAYER CHROMATOGRAPHY OF BALL PEN INKS

dyestuffs is not to be construed as being complete for ball pen ink dyes and is presented only to serve as a guide. For presumptive identification, an actual comparison examination with a standard dye sample would be indispensable.

DIscussIoN Ostensibly, it seemed forensically advantageous to identify the dye components of ink instead of characterizing ink by commercial brands, inasmuch as manufacturers change formulation frequently without notice. Some sixty or more commercially prepared 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 shown to be mixtures containing presumably isomers and/or unreacted constituents as has already been suggested by Coldwell (1) in his discussion of dye identification. For this reason, direct spot testing of some of these dyes was valueless for our purpose FrGuR 2 and the principal spot from each separated dye Micro-chromatoplates showing separation of two dif- preparation on our chromatoplates was tested with ferent ball pen inks. The ink was removed from a writ- solvents and reagents listed in (1), (2), and (3). ing on a document by the punch method utilizing a hy- podermic needle. While a presumptive identification can be made of some of the more common dyes used in the minutes. The slides are removed and air dried, manufacture of ball pen ink, the primary value of and may later be photographically recorded. spot testing must remain in the comparison of a Spot Tests. With the aid of a pointed instrument, questioned ink sample from that of a standard. four or more small portions of each separated dye The examination of blue ink samples from popular is removed from the chromatoplate and trans- American and Japanese ball point selected at fered to wells of a spot plate. To the four wells random (e.g.: Playmate, Parker, Script, , containing particles of dye-impregnated silica gel, Mitsubishi, etc.) revealed that the dye formula- tion employed in the manufacture of ball pen ink concentrated HCl, concentrated H2SO 4, NH4OH, and 10% NaOH are introduced in small drops from is not too diverse, and the TLC method and a capillary pipette to the respective wells. Color spot testing confirmed that such dyes as methyl changes are noted, then a drop or more of a satu- violet, rhodamine-B, Victoria blue, phalocyanine rated sodium carbonate solution is added to the blue, and alkali blue are of common occurrence. "HCI" well to observe any further change; many While only a few red dyes exhibited fluorescence dyes undergo an indicator type shift back to the in ultraviolet light, the marked difference particu- original color due to change in pH (2). Solubility larly shown by eosin and rhodamine furnished an of the separated dyes, then, are tested in water, added criterion for their identification. Nearly absolute ethanol, acetone, and chloroform in four all of the dyes were made invisible by infrared additional wells. Agitation is provided with a photography, hence differentiation of these com- thin glass rod or a capillary tube having a sealed pounds by this procedure was not of great value. tip. Observation under a microscope set at four Black ink containing nigrosine or indulin type of to ten power is an aid to these micro-spot tests. dye, was difficult to resolve in any of the many For reference, Table I lists some of the common solvent mixtures tested; the dye manifested it- dyes distributed by major chemical and dye manu- self on the plates as streaks and not as spots. Such facturers in the United States for the manufacture dyes, therefore, were not listed in Table I. The of colored writing inks and their immediate re- formulation of dyes in fluid inks, as they were char- actions to these test reagents and their responses acterized by this TLC method using the standard to ultraviolet and infrared lights. This list of solvent mixture and those using 1-butanol:50, G. R. NAKAMUR-4 AND S. C. SHIMODA [Vol. 56

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10% oxalic acid:15, absolute ethanol:10; and dye spots for micro-tests. Also, this procedure 1-butanol:50, 6 N ammonium hydroxide:15, precludes having to bleed the ink writing with absolute ethanol:t0, clearly revealed that the acid directly on the document for dye identifi- more popular blue "" inks such as cation such as done by Crown, et al (2). This will Parker, Script, Pilot, etc., contained seven or prevent further damage to the writing and hence more blue dye components and in some cases red to the document. and green dyes. Cartridge inks of various popular American and Japanese brands were found to contain, by the A thin-layer chromatography system is de- present TLC method, the same dyes as their scribed for characterizing small amounts of blue bottled ink counterparts. For example, Parker blue and blue-black ball point inks collected from Quink in both bottle and cartridge form had the writings. A procedure is described for removing same dye components. portions of the separated dyes for further char- In testing fluid inks by TLC, there was some acterization by spot testing. A table listing com- difficulty in completely removing the ink from mercial dyestuffs used in the manufacture of discs punctured from an ink line. A number of colored writing inks, their Rf values, solubility solvents including adds, alkali, and salts including and color reaction in acids and bases are reported. potassium binoxalate (4) were tested, but none Acknowledgment: The authors are indebted to afforded complete removal; some residual ink, Sp/7 Gerald A. Boardman for his assistance in presumably pigments, remained in the fibers. color, ultraviolet and infrared photography and Fluid ink also had a tendency of penetrating into to Koichi Funakoshi and Seisaburo Miyamoto for the silica gel layer leaving only a light spot on the their technical assistance in chromatography and surface. These losses seriously hampered micro- spot testing. To General Aniline and Film Corp., application such as described here for ball pen National Aniline Division, Allied Chemical Corp., ink. Further study is being conducted to establish Dupont DeNemour Co., and Pilot Pen Co., Japan, a micro-application method for fluid inks. for their aid, co-operation and generous contribu- A number of commercial silica gel preparations tion of their dyes. were tested, and it was found that the amount of binder substance, calcium sulfate, had an ap- REFERENCES preciable effect on the resolution of ink dyes. The 1. CoLrwELL, B. B., "The Comparison of Inks and more commonly used "Silica Gel-G" containing Writings by Paper Chromatography". THE ANA- LYST, 80, 68 (1955). 13 % calcium sulfate, although offering comparable 2. Caoww, D. A., CONWAY, J. V. P., AND Krnx, P. L., results, did not resolve some of the ball point ink "Differentiation of Blue Ball Point Pen Ink". samples in the manner effected by "Wakoge! B-5" J. CRIMINAL LAW, CRIMNOL., AND POLICE SCL, 52, 338 (1961). which has a 5% calcium sulfate inclusion. A silica 3. B Arcxxrr, JR. J. W., AND BAA'oRn, L. W., "Com- gel preparation containing no binder, "Wakogel parison of Ink Writing On Documents By Means B-0", offered results comparable to those of of Paper Chromatography". J. CRaIM LAW, CRImnNOL., AND POLICE Sci., 43, 530 (1952). "Wakogel B-5" indicating that the amount of 4. MATHY.R, J., "Comparison of Inks by Paper Chro- calcium sulfate does affect resolution of ink dyes matography, Practical Methods". INTERNATIONAL CR AI. POLICE RE iEW, 148, 130 (1961). on TLC. 5. DOUD, D., "Chromatographic Analysis of Inks". J. The advantage of the TLC method is in the oF FoRsENSIC SciEicEs, 3, 486 (1958). high resolution of dyestuffs as compared with that 6. Wirr, A. H., "The Examination and Identification of Inks". METHODs OF FORENSIC SCIENCE, Vol.. of paper chromatography, for example, methyl vio- II, 35, (1963), Interscience Publishers, London; let will resolve characteristically into a formation New York. of three or four spots (Fig. 2) strung like violet 7. HARRISON, W. R., SUSPECT DocumaT'rs, THEIR ScIENTIFIC EX.AmINATfON. Frederick A. Praeger, beads. None of the paper chromatography systems Publisher, New York, 1958. (1, 3, 4, 8) which we have tested make this char- 8. Pairxa, J. J., "Rapid and Simplified Method of Analysis By Thin-Layer Chromatography Using acterization: The separation is sufficiently discrete Michrochromatoplates". MICROCHIMICA AcrA, 3, to allow examiners to remove small portions from 529 (1962).