j. Soc. Cosmet. Chem.

, 39, 383-392 (November/December 1988)

Methodto determine silicones on humanhair by atomic absorption spectroscopy

EMMETT G. GOOCH and GRETCHEN S. KOHL, Dow Corning

Corporation, 2200 West Salzburg Road, Midland,MI 48686-0994.

Received June4, 1987.
Synopsis

This method consists of an enzyme digestion of hair samples to whichsilicone-containing hairproducts havebeenapplied. The released organosiloxane is recovered by liquid-liquid extraction into an organic
solvent. The silicon content of the solvent extractis measured by atomicabsorption spectroscopy (AAS) to determine the total mass of deposited silicone on the hair.

INTRODUCTION

Hair doesnot containnaturallyoccurring amounts of organosilicon (silicone) species (1). Generally,naturallyoccurring siliceous moietiesare in the form of silica, silicates, or substituted silicates. Unlike most commonly found siliceous materials,organosi1oxane polymers, provided theyarenot crosslinked, aresoluble in organic solvents. This organosiloxane solubilityin organicsolvents formsthe basicconcept for a selective solventextraction approach. However,somesiloxanes suchasaminofunctional siloxanes bind so tightly to hair that meresolventextraction is not sufficient for quantitative
removal.

Therefore, the first stepin the development of thismethod wasto find a wayto physicallyrelease the siloxanes fromthe hair. The nextstepwasan organic solvent extraction of the siloxane, followed by atomicabsorption spectroscopy analysis of the extract.
EXPERIMENTAL
MATERIALS

Test tresses weremadeof virgin European, naturalbrownhair obtained from DeMeo Brothers,Inc. The siloxanes usedare illustratedin Figure 1 and wereappliedas emulsions, dilutedto 0.25 or 1.0 weight% silicone activein deionized water,asspecified for eachexperiment. A 200-g treatmentbath of diluted emulsion wasusedfor each 12-g hair tressprepared. Eachtresswaspre-washed with a non-silicone-containing shampoo, and thendippedinto a treatment bathfor 30 seconds. The tress wasrinsed under 40Crunning tapwater for 30 seconds, andthenair driedpriorto sample preparation.

The dimethylpolysiloxane fluids (dimethicones) usedfor AAS calibration were Dow Corning 200 fluids,350 and 1000centistoke viscosity.
383

384

JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS

Me

OH

I o-)-(-si HO(-Si---o 3;-H

Me R

H(CH2)2NH2

M (-Si--O-) (-Si--O--)--Si eSI-OI ' [ ' VMe

Me Me R Me

Me.

Me

Me

Me

H(CHz)zNHz
LYLAIODnTCOI

Me

Me

Me

li__O_ )--Si X/ Me'Si-O(-S '-----Me

Me Me Me
Figure 1. Siloxane structures.

Papainenzyme is isolated from the latexof the Carica papaya plant. Suitable sources of

this enzyme included Pfaltz& Bauer(item #PO1100) and SigmaChemical (catalog #P-3375). Methyl isobutylketone,concentrated hydrochloric acid (36-37%), and sodium sulfitewerereagent-grade available fromanybulk supplier. Kimble 4.0-dramvials(catalog #60957) with polyethylene protected caps wereused for the sample preparation, and extraction wascompleted with 5-ml plastictransfer pipets(Fisher catalog # 13-711-9A).

METHOD

FOR SILICONES ON HAIR

385

INSTRUMENTATION

The atomic absorption assay was performed ona PerkinElmerModel380 spectrometer with a siliconhollowcathode lamp and a 5-cm slot nitrousoxide-acetylene burner.
Samples were prepared with the aid of a Burrell wrist-actionshakerand an IEC Clin-

ical model 428 centrifuge, equipped witha 6 X 52-mlangle rotor and50-mlshields. Photos weretakenon Polaroid Polapan Type-52film, through theobjective of a JEOL
T300 scanning electron microscope. Hair samples weremounted on an aluminumtab with double-sided tape and then gold-sputtered for 50 seconds. The tab was then mounted in the microscope and the samples viewedwithouttilting the stage.No difference in imageresolution wasdetectable by rotatingthe angleof the stage.The photos weretakenat a magnification of 2000 x.

SPECIFIC

SILICON

ASSAY TEST PROCEDURE

[Special note:As thisis a traceelemental analysis technique, caremustbe takenduring sample preparation to avoidinadvertent contamination of the samples and reagents. Latexgloves should be worn if the analyst hasusedsilicone-containing cosmetics, to
avoidsamplecontamination.]

Sample preparation procedure. Using solvent-cleaned scissors, about 50-75 hair fibers werecut fromeach previously treated,driedhair tress to be evaluated. The fibersfrom the tress werecut just belowthe (carnauba-waxed) boundendof the tress, clippingoff anddiscarding the upper2.5 cm of these hairs(rootend).The remaining fiberswere cut into approximately 0.3-cm length clippingswith the cleanscissors.

Individual samples wereprepared by weighing 0.10 _+0.02 g of theclipped hairinto a taredvial, usingsolvent-cleaned tweezers. The hair sample weightwasrecorded to use for calculating mg/kg Si fromthe reported AAS results. Samples werethendilutedwith 10.0 _+ 0.10 g of enzyme solution,asprepared in the formulabelow:
Papainenzyme Sodium sulfite Distilled water 0.13 -+ 0.02 g 2.00 _+ 0.10 g 100.00 _+ 0.10 g

[Shaken until dissolved, then adjusted to pH 6.8 with HC1 (conc.).This recipeis enough for nine samples and a blank. Freshenzyme solutions should be prepared for eachday'stesting.]
Each vial was then cappedand shakenlightly to wet the hair clippings. A blank sample, of just 10.0 g of enzyme solution, wasalsoprepared. All of the samples were

thenplaced in a blockheater or in an oven setat 63C_+ 3 for threedays,swirling

eachsample at leastonceduring the heatingcycleto make surethat all of the hair sample is in the solutionand not adhering to the sides of the vial. To eachsample,seven milliliters of methyl isobutylketone (MIBK) and five hundred microliters (0.5 ml) of concentrated reagent hydrochloric acidwereadded. The samples were then placedin a Burrell mechanical shakerand agitatedfor 30 minutes.The extraction vialswerethenspunin an IEC Clinical centrifuge at moderate speed for 20-30 minutesto permit phaseseparation. The 4.0-dram vials will fit into 50-ml centrifuge shields. Following separation, the top (solvent) layerof each sample is drawn off with a transfer pipet andreserved in a newvial for AAS analysis.

386

JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS

Instrument calibration andsample analysis. Silicon wasdetermined in the extracts by AAS, usinga nitrous oxide-acetylene flame.The atomicabsorption spectrometer initialization procedure followedthe manufacturer's recommended values(i.e., for gas flows and
lamp current). The 251.6-nm silicon wavelengthwas used. After the instrument warm-upperiod, the gasflow, nebulizer uptake, and burnerheadalignmentadjust-

mentsweremadewhile aspirating a Dow Corning 200 fluid-MIBK solution containing60 to 70 xgSi/ml. The finaloptimization adjustments for bestsensitivity and
signal-to-noise ratio weremadewhile aspirating the water-saturated blank solventand calibratingsolutions.

Calibration in bothwet anddry solvent solutions wasnecessary because silicon is very

sensitive to small changes in flame temperature. The water which saturates the MIBK duringthe solvent extraction stepalters the temperature andfuel-oxidant stoichiometry of the N20-C2H2 flame. The atomicabsorption response for siliconin wet MIBK is perhaps reduced to 70% ascompared to the samequantity of siliconin dry MIBK.

Dry MIBK silicon calibration solutions wereprepared from Dow Corning 200 fluid, which hasa polymerrepeating unit molecular weight of 74. ! g/unit of Me2SiO. It contains 37.9 percentsilicon.Viscosity grades between100 centistokes and 1000 centistokes arepermissible for useasstandards. For a typicalstandard, 50 mg of DC 200 fluid wasweighed into a 250-ml flaskpartiallyfilled with MIBK. DC 200 fluid tends
to adhere to dry glasssurfaces and may requirea long contacttime with the solventto ensurecompletesolubility of the entire sample.An exactweight of 50 mg is not essential, but that weight in 250 ml will resultin a siliconconcentration of 75 }zg of

silicon per milliliter of solvent, or 75 mg/kg Si on a wt/volume basis.

The calibrationplot of Si concentration vs. absorbance was linear from the zero intercept througha siliconconcentration of 75 mg/kg providedthat the atomicabsorption spectrometer wasset up underoptimum conditions.The exactsiliconconcentration was calculated usingthe followingequation:

Sample weight(milligrams+ 0.1 mg) x 28.1 x 1000

74.1 x 250ml

= micrograms of Si per ml

This weight/volume siliconstandard may be serially dilutedwith MIBK to produce calibrating solutions lowerin silicon content asrequired.

To makethe wet solvent standard calibration solutions, an aliquot of thestandard was shaken with an excess of water,which wascentrifuged out, priorto use.Forexample,
to a 25-ml aliquotof calibrating solution, five ml of waterand0.25 ml of concentrated HCI reagentwere added. Dry standards can be storedand will remain constant,but wet standards deteriorate on standing andsoweremadeanewfor each day's testing.In additionto the standards, the background solvent whichis aspirated into the instrument between samples mustalsobe water-saturated. A supply of MIBK stored over water wasmaintainedto usefor background solvent.
Samples wereaspirated into the AA spectrometer, and the absorbance values at 251.6 nm wererecorded alongwith the absorbance values for the wet calibrating solutions. A calibration plot of absorbance vs. siliconconcentration wasconstructed. The concentration of siliconin the sample extracts wasrecorded from the calibration plot, and then multipliedby the extractsolvent volumeof 7 ml to obtainthe total weightof silicon

METHOD

FOR SILICONES

ON HAIR

387

recovered from the sample. The total weight of silicon,dividedby the hair sample weight,gavethe concentration of silicon on the hair. The quantityof siloxane polymer treatment on the hair wasdetermined by multiplyingthe silicon valueby the proper gravimetric factorfor the specific polymer.

A carefully alignedand adjusted atomicabsorption spectrometer typicallyrecords an absorbance of 0.050 absorbance unitsfor 10 xgof silicon permilliliter (10 ppm) in wet MIBK. For AAS, sensitivity is generally definedasthat quantityof analyteper milliliter whichwill produce anabsorption signal of onepercent, or an absorbance of 0.0044
absorbance units. The sensitivity for organosilicon in wet MIBK is 0.88 xgof silicone per milliliter. The instrument should be expected to reproduce an absorbance measurement of 0.050 within _+ 0.002 absorbance units. The absolute detection limit for

siliconefor theseexperiments was 0.002 absorbance units. An absorbance of 0.002 units may not be readilydiscernible from the normalbackground noisewith instrumentsequipped only with light-emittingdiodearray(LED) readouts, but may be seen on a stripchartrecorder. The detection limit for silicon,based on a total solvent volume of 7 ml, is about3 xg, with a precision of -+ 3 }xg. If the spectrometer features scale

expansion capability, moderate scale expansion of 2 X or 3 X mayprovidesome sensitivity improvement.Scale expansion is an aid when one is attemptingto discernsmall differences in absorbance signals between samples.

RESULTS

AND

DISCUSSION

METHOD

SELECTION

Siloxanes, whenplacedin contact with porous surfaces suchashair, tend to be adsorbed

intothesubstrate andcanonlybepartially extracted with organic solvents. Treated hair must be decomposed prior to extraction. Several reagents weretried for preparing treated hair samples for AAS assay, including caustic (KOH andNaOH), phosphoric acid,andtetramethylguanidine. While thehairprotein structure could bedecomposed with sodium or potassium hydroxides, those materials werenotselected, astheycaused cleavage of the organosiloxane polymer into cyclics, silanols, andsilicates. Those reaction products wereeithervolatileor non-extractable in organic solvents. Phosphoric acidandtetramethylguanidine did notsufficiently decompose thehairat roomtemperature,even afterseveral weeks, andwhenheated also caused siloxane polymer degradation and generation of non-extractable species. As an alternative,an enzyme digestion methodwasdiscovered in a literaturesearch (2)

that employed papain (an enzyme foundin papaya). This enzyme is a proteolytic enzymethat is active with freesulfhydryl groups. An aqueous solution containing 0.13% papain and 2% sodium sulfiteasa catalyst (with the pH adjusted to 6.8) washeated with a hair sample at 65Cfor threedays.Samples prepared by this method were furtherextracted with methylisobutyl ketone (MIBK) anda smallquantity of hydrochloric acidbefore AAS analysis. The HCI served to precipitate otherwise soluble proteins andaided in phase separation laterin theprocedure. Theextraction was completed byshaking thevialandcontents ona Burrell mechanical shaker, andcompleting phase
separation in a centrifuge. The top (solvent) layerwasdrawnoff and the siliconcontent

was assayed by atomic absorption spectroscopy utilizinga nitrous oxide-acetylene flame.

388
METHOD

JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS

ACCURACY DETERMINATION

The siliconcontentof organosilicon materials wasdetermined by AAS afterdilution of the siloxane of interest in an organic solvent. However,the silicon absorption signal,or signalintensity,canvary, depending on the chemical structure of the siloxane. Calibration error is minimized if a siloxane of known purity and having the samechemical structure and vaporpressure as the materialto be assayed is foundfrom which to prepare calibrationstandards. In actualpractice,it wasmore convenient to compare the signal intensity of the compounds of interestto calibratingsolutionspreparedfrom known purity standard materialsand then to decideif the signal response differences betweenthe materialsintroduced an acceptable level of error. If the errorwasonly a percentor so relativeto the actualsiliconcontentof the material, that error is negligible when measuring siliconlevelsof a few partsper million. Normal instrumental signal-to-noise ratios will impart far greater measurement error, especially with readings close to the detection limit. Standard deviation of the measurement is givenin
all of the data tables.

To test the accuracy of the methodfor detectingthe properratio of Si polymeremulsion, several samples were assayed. Table I contains the results from samples of three differentaminofunctional siloxane emulsions: amodimethicone and two trimethylsilylamodimethicone (TSA) polymers[seeFigure 1] with differentpolymerchainlengths and percentamine functionality. The assayed Si valuesare within about 10% of the
calculated values.

The calibratingstandardused in the previousexamplewas a well-characterized dimethyl fluid with a viscosity of 1000 centistokes. To checkthe correlation of various neat organofunctional siloxane fluids with this standard,several samples of different functionalitysiloxane fluids wereanalyzed. Thosedata are reportedin Table II. The Si assay of thesesiloxane polymers matched veryclosely to the calculated % Si values also, exceptfor one polymer (siloxane E) which contained a very long dimethyl siloxane chain, and which because of its high molecular weight is lesssolublein the organic solvent.As a resultof thesetests,it wasdecided to usethe 1000-centistoke dimethyl fluid as the primarystandard for all of the futuresilicondeterminations.
PRECISION OF METHOD

The methodwastestedfor reproducibility (precision) by analyzingduplicatesamples

Table I

Quantitationof % Si in Aminofunctional Siloxane Fluids

Calculated Calculated Calculated

Sampleemulsion

molecularwt

% Si

% Si in

% Si

(35% silicone solids)

A TSA, x = 96, y = 2
B Amodimethicone

of polymer
7624
38038

in polymer
36.7
36.8

emulsion
12.9
12.9

by AAS (+ SD)*
12.4 _ .2
12.1 --- .2

C TSA, x = 45.8 y= 2.2

3939

35.5

12.8

11.5 --- .2

* Meansrepresent averages of two determinations, and standard deviations (SD) represent only instrumental variation.

METHOD

FOR SILICONES

ON HAIR

389

Table

II

Accuracy of UsingDimethylSiloxane Standards for Quantitation of % Si in Various Organofunctional Siloxanes

%Sias

Sample (neatfluids)
Siloxane A SiloxaneB SiloxaneC SiloxaneD Siloxane E SiloxaneF

Calculated MW of polymer
8005 7980 7698 8059 30480 7630

Calculated % Si
35.0 35.1 36.4 34.7 41.3 36.7

determined by AAS ( ___ SD)*

35.6 36.2 36.3 33.4 35.3 36.8 + .3 + .3 + .3 -4_-_ .3 + .3 -4_-_ .3

* Meansrepresent averages of two determinations, and standard deviations (SD) represent only instrumental variation.

prepared by eachof two differentresearchers. Table III contains the results from three differenttreatedhair samples as determined by two operators. The hair samples included untreatedhair and two increasingly higher levelsof siliconetreatment (with TSA). Variationin the mg/kg Si as determined by the differentoperators waswithin experimentalerror (standarddeviationof +/30 mg/kg Si), as was the variation between replicate samples. It should be notedthat an accurate weighingandrecording of the hair sample weight is importantin controllingthe accuracy of the calculated Si mg/kg. The Si values for untreated hair should be close to the detection limit or the test shouldbe repeated.With valuescloseto the detectionlimit, the standard deviation (SD) of the measurement often approximates the magnitudeof the readingitself. The blank should measure close to or belowthe detection limit, meaningthat thereis no Si present. Sample readings that arein thislow rangeof detection should alsobe suspected of havingno Si present.
APPLICATIONS OF METHOD

A series of siloxane emulsions was used to treat hair. The emulsions were all diluted

to

0.25 wt % solids in deionized water, andtwo TSA fluidsof varyingpolymerlength and % aminofunctionality wereevaluated (represented by x and y, respectively, in Figure
Table III

Precision of Method by Two Operators

mg/kg Si by A.A. (+ SD)*

Tress # K13-3 Treatment Low conc. TSA (1) 150 + 30
170 --- 30

(2) 140 --- 30

150 + 30 +
380 +

30 30
30

K13-U

Untreated

70 +- 30
70 -+ 30

60 --- 30

K14-2

High conc.TSA

370 + 30

380 + 30

* Meansrepresent averages of two determinations, and standard deviations (SD) represent only instrumental variation.

390

JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS

Table
First Series of Aminofunctional

IV
Siloxane*-Treated Hair

Treatment (. 25% Emulsion) A TSA, x = 96, y = 2 B TSA, x = 298, y = 2

C Dimethicone D Blank

mg/kg Si by AA (+ SD)** 1170 + 50 4220 + 100

190 + 20 30 + 30

* Trimethylsilylamodimethicone (TSA). ** Meansrepresent averages of duplicatedeterminations.

1). The deposition levelsof thesepolymers were compared to that of a dimethicone emulsion. Table IV contains the results,that showthat the higher-molecular-weight amine-containing polymer deposits the most(4220 mg/kg Si), followed by the lowermolecular-weightamine-containingpolymer (1170 mg/kg Si). The dimethicone showed only 190 mg/kg Si, andthe blankcontained <30 mg/kg Si. Scanning electron microscopy (SEM) visuallyconfirmed this progression of deposition. Figures 2- 5 showSEM photos takenat a magnification of 2000 x. The untreated hair (Figure2) shows highly definedcuticlescales. No gross surface changes areapparent for the dimethicone emulsion-treated hair (Figure 3). However, the lower-molecularweight amine-containing polymertreatmentshows a full coatingof the cuticle, with loss of cuticledefinition(Figure4). The photoof the higher-molecular-weight aminecontaining polymertreatment(Figure5) shows a heavy,"wrinkled"coatingof silicone, obscuring the cuticledefinition.

Figure 2. SEM photograph of untreated hair.

METHOD

FOR SILICONES

ON HAIR

391

/-

Figure3. SEMphotograph of hairtreated with a dimethicone emulsion.

CONCLUSIONS

A methodis presented for bulk analysis of the Si contentof hair fibers. The method consists of enzymedigestingsilicone-treated hair, and extractingthe organosiloxanes with an acidifiedorganicsolvent.The detection limit with currentinstrumentation is 3

Figure4. SEMphotograph of hairtreated with low-molecular-weight TSA emulsion.

392

JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS

Figure 5. SEM photograph of hair treated with high-molecular-weight TSA emulsion.

xgof Si. Reproducibility of mg/kgdata,between replicate samples andbetween operators during samplepreparation,was within instrumentalmeasurement error when

careful attention waspaidto accurately weighing the hair sample. Applications of this method show a trendbetween atomic absorption-determined Si deposition anddeposition asvisuallydetermined by SEM.
REFERENCES

(1) C. R. Robbins,Chemical and Physical Behavior of HumanHair (Van NostrandReinholdand Co., 1979), p. 39(2) A. W. Holmes,Degradation of human hairby papain,PartI: The pattern of degradation, Text.Res. J., 34, 706-712 (1964).