Journal of Pharmaceutical and Biomedical Analysis29 (2002) 263–275

Characterization of gamma irradiated petrolatum eyeointment base by headspace–gas chromatography–

mass spectrometry

Lan Hong, Hans Altorfer *Institute of Pharmaceutical Science, Swiss Federal Institute of Technology, ETH Zurich, I17 M48, Winterthurerstrasse 190,

CH-8057 Zurich, Switzerland

Received 5 October 2001; received in revised form 14 January 2002; accepted 11 February 2002

Abstract

The effects of gamma irradiation on petrolatum eye ointment base (EOB) and its ingredients (white petrolatum,liquid paraffin, and wool fat) were studied at different irradiation doses. Forty-one volatile radiolysis products weredetected and identified by a combined system of headspace–gas chromatography–mass spectrometry (HS–GC–MS).The characteristics of the radiolysis products and the degradation pathway were discussed in each case, respectively.GC method demonstrates that the component distribution patterns of eye ointment as well as its individualingredients have no differences before and after gamma irradiation. The influence of gamma treatment on EOB wasquantitatively determined at 15, 25 and 50 kGy. The concentrations of the radiolysis products increase linearly withincreasing doses. Both qualitative and quantitative data show that irradiated eye ointment is safe for human use.© 2002 Elsevier Science B.V. All rights reserved.

Keywords: Headspace–GC–MS; Petrolatum eye ointment; Gamma irradiation

www.elsevier.com/locate/jpba

1. Introduction

Ophthalmic ointment products are among themost successful dosage forms to which radiationsterilization has been applied, mainly due to threefactors. First, the most effective thermal steriliza-tion cannot be applied to ointment products be-cause of their low melting point. Second, theaseptic process, a traditional sterilization method

for eye ointment products, involves the possibilityof secondary contamination and excessive costs.Third, the intrinsic nature of strong penetration ofgamma rays enables gamma sterilization to be theonly method for terminal sterilization of eye oint-ment products.

Eye ointments are normally petrolatum-basedand designed to have a melting point close to thehuman body temperature. The petrolatum base isused as an anhydrous medium to deliver mois-ture-sensitive drugs, like chloramphenicol. Thebase adopted as the excipient of chloramphenicoleye ointment consists of petrolatum, liquid

* Corresponding author. Tel.: +41-1-635-6064; fax: +41-1-635-6885.

E-mail address: altorfer@pharma.ethz.ch (H. Altorfer).

0731-7085/02/$ - see front matter © 2002 Elsevier Science B.V. All rights reserved.

PII: S0 731 -7085 (02 )00017 -1

L. Hong, H. Altorfer / J. Pharm. Biomed. Anal. 29 (2002) 263–275264

paraffin, and wool fat. These ingredients are gen-erally considered to be non-irritant and -toxicmaterials. Therefore, it is important to clarifywhether or not the radiation sterilization processprovokes the formation of unsafe radiolysis prod-ucts for their use in pharmaceuticals.

Earliest investigation of radiated white petrola-tum and liquid paraffin were reported during1960–1963 [1–3]. However, a few cases concernedointments or ointment ingredients, and these lim-ited studies were focused on the changes in pHvalue, physical properties and active ingredientsupon gamma irradiation [4–11]. There has beenno report available on whether or not and howgamma irradiation causes chemical componentchanges in ointment base as well as its relevantingredients although the information about poten-tial decomposition products is required for thevalidation of gamma radiation sterilization.

Gas formation in petrolatum eye ointmentproducts is a phenomenon unavoidable duringgamma irradiation. Thus, the gas compositionneeds to be clarified and untoward activity orcarcinogenic potential of these gases for humanhealth must be assessed. Nevertheless, there hasbeen no report concerning the composition ofthese gases, though the existence of hydrogencarbon dioxide, and methane in the gases wasreported [11].

The present work pursued the chemical impactof gamma radiation on eye ointment base (EOB)as well as its ingredients (white petrolatum, liquidparaffin and wool fat). Organic volatile impuritiesproduced from these materials during irradiation

were clarified, with qualitative and quantitativedata. The research findings enable the elucidationof radiolysis mechanism and the assessment of thetoxic potential of the radiolysis products for hu-man health.

2. Experimental

2.1. Reagent and materials

Eye ointment base (EOB, a petrolatum oilybase without active ingredient), white petrolatum,liquid paraffin, and wool fat were supplied byCIBA Vision AG (Hettlingen, Switzerland). Ref-erence compounds, solvents and reagents were ofanalytical reagent grade or better from FlukaChemie GmbH (Buchs SG1, Switzerland).

Samples were irradiated with gamma rays from60Co at 15, 25, 50, and 100 kGy respectively, in aradiation sterilization plant, Studer Draht-undKabelwerk AG (Daniken, Switzerland).

2.2. Apparatus

A BGB-Silaren column (10 m×0.32 I.D., 0.12�m, GBG Analytik AG, Adliswil, Switzerland)was used for gas chromatography analysis. ARtx®-624 column (30 m×0.32 I.D., 1.8 �m,Restek, Benner Circle, Bellefonte, PA, USA) wasused for headspace–gas chromatography–massspectrometry (HS–GC–MS) analysis. Operationconditions for gas chromatography (Varian3400cx, Sugar Land, TX, USA) and HS–GC–

Table 1Experimental conditions for GC and HS–GC–MS analyses

GC analysis HS–GC–MS analysis

50 (2 min) to 180 °C at 30 °C min−1 to 280 °C at 45 (5 min) to 95 °C at 2 °C min−1 (25 min) toTemperature5 °C min−1 (25 min) 125 °C at 6 °C min−1 (15 min)

Carrier gas 55 kPa, He (5.0 grade), 1.1 ml min−1He (5.0 grade), 2.7 ml min−1

Detector MSDFID (350 °C)Injection Headspace autosampler (Table 2)SPI, 50 (0.5 min) to 280 °C at 200 °C min−1

(30 min), 1.0 �l4 mg ml−1 in cyclohexane 500 mg original sample per vialSample

L. Hong, H. Altorfer / J. Pharm. Biomed. Anal. 29 (2002) 263–275 265

Table 2Operation conditions of the headspace autousampler

Varian Headspace Autosampler-Tekmar 700080 °CPlaten temperature Loop fill time 1.5 min

Loop equilibrium time50 min 0.2 minSample equilibrium time10 minMixing time Inject time 0.5 min

Valve temperatureStabilising time 200 °C2 minTransfer line temperature8 psi 200 °CVial pressureLoop volumePressurising time 1 ml1.5 minVial size0.20 min 22.5 mlPressurising equilibrium

MS (Varian Headspace Autosampler-Tekmar7000, Varian 3400cx, Saturn 4D/GC/MS/MS,Sugar Land, TX, USA) are summarized in Table1. Conditions for the Headspace Autosamplerare shown in Table 2. Mass spectra were ob-tained at the electron impact of 70 eV and chem-ical ionization (with methane as reagent gas) at15 eV. Chromatograms were recorded by moni-toring the total ion current in the range of 30–400 amu. Transfer line and ion trap manifoldwere maintained at 220 and 170 °C, respectively.

The compound identification was carried outby matching mass spectra with those recorded inthe NIST (National Institute of Standards andTechnology) mass spectral search program aswell as their molecular weights (obtained fromchemical ionization process). Identification re-sults were confirmed by the injection of authenticsubstance as reference, wherever possible (total27 compounds), and by the regular cycle of re-tention time of the homologues.

2.3. Sample preparation

Five hundred milligrams of each ointmentsample was weighed in a headspace vial andcapped air-tightly for the chromatographic pro-cedure.

For the preparation of calibration curves, ref-erence compounds were dissolved in 1,3-dimethyl-2-imidazolidinone (DMI), then spikedin a small volume (10 �l) into EOB-0, whichcontained no detectable VOC compounds underthe headspace conditions. Thus, the matrix ofstandard samples is created to be similar to thatof eye ointment samples and the matrix effect[12] of EOB can be reduced.

3. Results and discussion

3.1. Degradation profiles of EOB andidentification

The most obvious variation of irradiated eyeointment is manifested in the production of bub-bles after irradiation, even at 15 kGy. This indi-cates the formation of volatile radiolysisproducts. HS–GC–MS was therefore employedto analyze these volatile products. A typical totalion chromatogram of an EOB sample irradiatedat 50 kGy, along with the chromatogram of non-irradiated EOB, is presented in Fig. 1. Forty-one volatile compounds were detected andidentified in the irradiated EOB, while onlyfour small peaks were found in the non-irradi-ated sample. The size of the four peaks found inthe non-irradiated EOB increases after irradia-tion, evidencing that all volatile compounds de-tected in the irradiated EOB are radiolysisproducts.

Hydrocarbons are the dominant radiolysisproducts, including 14 alkanes, 18 alkene, andfive cycloparaffin. Alcoholate and ketone werealso observed in radiolysis products. Becausesome alkene compounds have very similar massspectra, the chemical structures of six alkenes(peaks 4, 11, 12, 22, 33 and 34) cannot be as-signed with confidence. The main radiolysisproducts are n-alkanes (peaks 1, 3, 9, 20, 31, and41), except peak 13 (Fig. 1). Hydrocarbon com-pounds in radiolysis products can be divided ap-proximately into six groups sorted by theircarbon numbers, i.e. hydrocarbon compounds inthe same group have the same carbon numbers.Each n-alkane can be a mark of the group to

L. Hong, H. Altorfer / J. Pharm. Biomed. Anal. 29 (2002) 263–275266

which it belongs. As Table 3 shows, the straightand branch chain hydrocarbons of radiolysisproducts appear in a regular order. Whenever thecarbon number allows, certain homologues existin each group.

3.2. Degradation profiles of indi�idual ingredients

The production of bubbles was also observed inwhite petrolatum and wool fat, but not in theliquid paraffin. However, volatile radiolysis com-

Fig. 1. Identification of the volatile radiolysis products of irradiated EOB by HS–GC–MS (EOB-50: irradiated at 50 kGy, EOB-0:non-irradiated EOB, n=carbon number).

L. Hong, H. Altorfer / J. Pharm. Biomed. Anal. 29 (2002) 263–275 267

Table 3Molecule structures and the order of the straight and branch hydrocarbons of the radiolysis products in six groups, n=carbonnumbers, the numbers are scan numbers (the same as in Fig. 1)

pounds were detected in all the three materialsusing HS–GC–MS after irradiation (Fig. 2),while these volatile compounds were not foundin the relevant non-irradiated samples. In addi-tion, the concentration of these volatile radioly-sis products in all samples was directlyproportional to the irradiation dosage (unpub-lished data).

Volatile radiolysis products in liquid paraffinand white petrolatum are the same (Fig. 2B andC), but the radiolysis product profile of wool fatlooks different from those of liquid paraffin andwhite petrolatum. In irradiated wool fat, straightchain alkanes (peaks 1, 3, 9. 20. 31, and 41) existonly in smaller amount (Fig. 2A). Acetone (peak13) is the most predominant radiolysis product in

L. Hong, H. Altorfer / J. Pharm. Biomed. Anal. 29 (2002) 263–275268

Fig. 2. Comparison of volatile radiolysis products at 50 kGy by HS–GC–MS: irradiated wool fat (A), liquid paraffin (B), whitepetrolatum (C), and EOB (D).

L. Hong, H. Altorfer / J. Pharm. Biomed. Anal. 29 (2002) 263–275 269

irradiated wool fat, whereas it is much less inirradiated liquid paraffin and almost disappears inirradiated white petrolatum. It is also true in thecases of 2-propanol (peak 14) and 2-butanone(peak 25). Cycloparaffin products (peaks 17 24, 28and 35) disappear in irradiated wool fat. Al-though there are two peaks at the same retentiontime of peaks 24 and 28 in chromatogram ofirradiated wool fat, their mass spectra prove thatthey are different from the radiolysis products inirradiated white petrolatum or liquid paraffin.

Nevertheless, white petrolatum, liquid paraffinand wool fat produce the same straight andbranch chain hydrocarbons as radiolysis products,though their relative intensities are different.

All volatile radiolysis products that appear inwhite petrolatum, liquid paraffin and wool fat arepresent in irradiated EOB (Fig. 2D). Cyclo-paraffins are the unique radiolysis products ofwhite petrolatum and liquid paraffin, and acetone,2-propanol and 2-butanone are the only radiolysisproducts of wool fat. However, all of these com-pounds appear in irradiated EOB. This meansthat acetone, 2-propanol and 2-butanone in irra-diated EOB originate from wool fat, while cyclo-paraffins come from white petrolatum and liquidparaffin.

3.3. Component distribution pattern

The distribution pattern of hydrocarbons wasidentifying by means of GC method (Fig. 3). Thecomponents of EOB, liquid paraffin and whitepetrolatum are so complex that the GC methodwith a direct injection does not achieve a completeseparation even with a capillary column. How-ever, the slow and drifted baseline suggests thepresence of isoparaffins, cycloparaffins, etc. andthe normal alkanes show appreciable peaks [13].The gas chromatograms of non-irradiated andirradiated samples overlap perfectly in each case,even at very high irradiation dose of 100 kGy(Fig. 3). Although the GC method could notprovide an enough separation capability, it hintsthat the amounts of the main composition ofEOB changes little after irradiation.

Furthermore, normal alkanes in the GC chro-matograms provide information on the distribu-

tion of carbon numbers. It was reported that forliquid or solid hydrocarbons, an important conse-quence of radiation effect was the formation ofradiolysis products of higher weight molecules[14], and under prolonged irradiation, a sufficientamount of the molecule became linked together toform a gel extending throughout the system [15].Present GC results show that the distributionpatterns of n-alkanes remain unchanged aftergamma processing and no peaks appear at theend part of the chromatogram. This providesstrong evidences that there are no detectableproducts of condensation reaction in the petrola-tum eye ointment.

In conclusion, the GC results demonstrate thatthere are no significant chemical changes in themain composition of the petrolatum ointmentbase as well as its individual ingredients aftergamma irradiation. The distribution of hydrocar-bons in EOB does not change significantly afterirradiation although volatile radiolysis productsare produced during irradiation.

3.4. Degradation pathways

3.4.1. Long carbon chain degradationParaffin differs from petrolatum in their carbon

chains. Wool fat differs from paraffin and petro-latum in molecular structure. EOB differs fromparaffin and petrolatum in composition. How-ever, headspace chromatograms (Fig. 2) showthat the radiolysis products of these different ma-terials include the same straight and branch chainhydrocarbons. This indicates that the radiolysisproducts of white petrolatum, liquid paraffin, andwool fat are probably produced by the similardegradation pathway, possibly the same func-tional group in their molecules degrades in allthese materials.

What all these materials have in common is thelong carbon chains in their molecular structure.Since the C�C bonds in large straight moleculesare of similar strength, and can be broken equallyunder irradiation [15], the initial ionization canoccur virtually anywhere along the chain. Thus,the radiolysis products are independent of theoriginal chain structure and its length after theinitial ionization. Accordingly, petrolatum and

L. Hong, H. Altorfer / J. Pharm. Biomed. Anal. 29 (2002) 263–275270

Fig. 3. Profiles by gas chromatograms: (A) irradiated and non-irradiated EOB (peak 1=cetyl alcohol, peak 2=stearyl alcohol, theyare also the ingredients of the present EOB); (B) irradiated and non-irradiated liquid paraffin; (C) irradiated and non-irradiatedpetrolatum.

L. Hong, H. Altorfer / J. Pharm. Biomed. Anal. 29 (2002) 263–275 271

liquid paraffin give the same degradationproducts.

In the case of wool fat, as long as the carbonyland long straight chain radicals are formed bybond rupture reactions, further bond scission ofthe C�C chains will proceed by the same path-ways as in white petrolatum or liquid paraffin.Eventually wool fat has the radiolysis productssimilar to those of liquid paraffin and whitepetrolatum.

As unique radiolysis products of wool fat, onthe other hands, the appearance of acetone, 2-propanol and 2-butanone reflects the chemicalfeature of wool fat. The presence of cycloparaffinsin irradiated white petrolatum and liquid paraffincould be due to the existence of cycloparaffingroups in molecular structures of the originalsamples.

The radiolysis product profile of EOB showsthat the radiolysis products of EOB are a totalcontribution of the radiolysis products of whitepetrolatum, liquid paraffin and wool fat. Thisindicates that the matrix changes have no influ-ence upon the radiolysis pathway of thesematerials.

3.4.2. Mass spectrum degradation modelThe earlier investigation evidenced that dissoci-

ation patterns obtained by mass spectrometry areessentially independent of the ionizing potentialemployed, so that mass spectral data could beapplied to radiation–chemical systems irradiatedwith radiation of much higher energy. This meansthat the dissociation pattern of a compound in themass spectrometer could be a useful source ofinformation in establishing the radiolysis mecha-nism [16].

It was found in the present work that theradiolysis product profile of irradiated EOB de-tected by HS–GC was quite similar to a typicalmass spectrum of long carbon chain n-alkane.The Fig. 4 shows that the radiolysis products ofirradiated EOB exhibit the followingcharacteristics:– An abundance of low mass.– Characteristic peaks of n-alkanes.– Independent on the chain length.

These are features of mass spectra of longcarbon chain n-alkane, i.e. n-octadecane. Thecharacteristic ions in mass spectrum are [(CnH2n+

2)−1]+ and [(CnH2n)−1]+, and the characteris-tic peaks in irradiated EOB profile are CnH2n+2

and CnH2n. The series of intensive ions at m/z 43,57, 71, 85, 99 and so forth are highly characteris-tic of saturated aliphatic hydrocarbons of n-pro-pane (44), n-butane (58), n-pentane (72), n-hexane(86), n-heptane (100), etc.

This indicates that alkane chain in both massspectrometer and radiolysis process degrades by asimilar pathway. As the same in mass spectrome-ter, the secondary and especially tertiary C�Cbonds are particularly liable to fission duringionization process with high energy [14], accord-ingly, the proportion of fragmentation productsdecreases in the order of n-alkane� iso-alkane�neo-alkane, and n-alkanes become the featuredradiolysis products. And the main rupture of C�Cbond tends to continue from the chain end andresults in secondary or further fragmentation inthe following way:

The driving force for this fragmentation is theformation of the new �-bond in the extrudedmolecule of ethylene, which lowers the activationenergy toward the secondary fragmentation ofhigh mass ions [17]. As a result, intensive peaks oflow mass radiolysis compounds were observedand material with different chain length gave sim-ilar radiolysis products.

This result suggests by means of molecule rap-ture pattern in mass spectrum a rupture model ofradiolysis for long carbon chain hydrocarbonscould be created or confirmed without a greatdeal of laborious work.

3.5. Quantitati�e analysis and e�aluation

The quantitative determination of radiolysisproducts of the EOB is summarized in Table 4.The concentrations of all the radiolysis productsof EOB increase linearly with increasing irradia-tion doses. Most of the correlation coefficients (r)

L. Hong, H. Altorfer / J. Pharm. Biomed. Anal. 29 (2002) 263–275272

Fig. 4. Relationship between n-alkane degradations in MS process and gamma radiation process: (A) a mass spectrum ofn-octadecane from NIST database, in which the numbers represent ion fragment m/z of [M−1]+; (B) a total ion chromatogramof radiolysis products in irradiated eye ointment by HS–GC–MS, in which the numbers represent molecular weight of radiolysisproducts, n-propane (44), n-butane (58), 1-pentene (70), n-pentane (72), 1-hexene (84), n-hexane (86), 1-heptene (98), n-heptane(100), 1-octene (112), n-octane (114), 1-nonene (126) and n-nonane (128).

L. Hong, H. Altorfer / J. Pharm. Biomed. Anal. 29 (2002) 263–275 273

Tab

le4

Qua

ntit

ativ

ean

alys

isof

the

radi

olys

ispr

oduc

tsof

irra

diat

edE

OB

No.

Scan

no.

Con

cent

rati

on,

ppm

(RSD

%)

rbSc

anno

.N

o.a

Con

cent

rati

on,

ppm

(RSD

%)

rb

50kG

y25

kGy

15kG

y15

kGy

25kG

y50

kGy

7.0

(2.4

)0.

985

2210

801.

64(2

.4)

0.91

(6.9

)0.

79(5

.2)

0.98

81

396

26.4

(2.3

)15

.5(1

.3)

2311

387.

3(6

.7)

6.6

(5.7

)6.

4(6

.6)

0.99

90.

995

436

21.

6(3

.1)

2.5

(0.6

)5.

1(6

.8)

0.99

946

924

1222

2.4

(6.7

)1.

3(2

.4)

0.8

(3.5

)0.

999

40.3

(2.9

)21

.6(1

.6)

13.3

(2.0

)3

1.3

(3.3

)0.

999

2513

037.

8(0

.8)

5.1

(5.5

)3.

5(3

.0)

0.99

54

491

3.9

(3.4

)2.

0(3

.6)

2613

560.

96(4

.4)

0.41

(5.6

)0.

28(8

.7)

0.99

90.

995

2.1

(3.9

)1.

6(5

.3)

551

13.

4(2

.5)

0.99

756

327

1420

3.8

(3.2

)2.

1(7

.2)

1.2

(8.8

)0.

997

2.0

(6.3

)1.

2(1

.6)

0.7

(5.3

)6

2814

495.

3(1

.5)

2.7

(1.6

)1.

8(2

.0)

71.

000

589

7.7

(1.9

)3.

9(4

.9)

2.3

(4.5

)1.

000

2914

864.

6(4

.1)

2.3

(4.7

)1.

5(5

.1)

0.99

90.

999

639

82.

2(6

.3)

3.8

(1.6

)7.

2(4

.5)

1.00

065

030

1623

4.9

(7.2

)2.

4(2

.6)

1.8

(2.5

)0.

997

21.5

(1.7

)10

.9(4

.7)

6.6

(3.2

)9

0.6

(11.

2)0.

999

3116

5012

.0(0

.4)

5.8

(1.9

)3.

9(1

.8)

0.99

810

2.2

(6.2

)68

61.

1(1

6.4)

3216

680.

43(5

.6)

0.21

(2.4

)0.

17(4

.2)

0.98

50.

990

0.48

(10.

2)11

713

1.05

(14.

7)0.

55(5

.2)

3317

153.

0(1

.8)

1.6

(8.2

)1.

1(1

0.8)

121.

000

728

0.90

(0.5

)0.

54(3

.0)

0.49

(5.1

)0.

984

3417

280.

00.

00.

00.

999

0.00

013

6.3

(2.1

)8.

2(0

.2)

12.1

(3.3

)79

50.

982

821

3518

843.

9(2

.0)

2.1

(8.0

)1.

4(4

.2)

1.00

02.

1(3

.4)

1.2

(6.9

)1.

1(4

.5)

1436

2125

2.3

(4.1

)1.

3(1

.5)

0.8

(2.6

)0.

992

0.99

585

915

1.3

(6.8

)1.

6(5

.3)

3.2

(3.6

)0.

998

883

3721

533.

9(5

.2)

2.0

(2.6

)1.

1(3

.0)

0.99

910

.2(1

.0)

5.2

(3.2

)3.

7(2

.1)

160.

5(8

.3)

0.99

438

2170

0.75

(5.7

)0.

38(1

3.5)

0.20

(8.8

)0.

999

904

172.

3(4

.2)

1.2

(6.3

)39

2223

2.0

(5.7

)1.

0(5

.6)

0.6

(6.6

)1.

000

0.99

91.

6(9

.8)

1.0

(5.5

)18

953

3.2

(5.5

)0.

999

1009

4024

003.

1(1

.4)

1.6

(9.5

)0.

9(1

.8)

0.99

88.

3(1

.2)

4.3

(1.5

)2.

6(1

.8)

1941

2024

3010

286.

8(1

.7)

3.7

(0.6

)2.

2(1

.8)

0.99

920

.3(2

.8)

10.3

(2.1

)6.

9(2

.0)

0.99

90.

986

0.28

(8.7

)0.

34(1

0.5)

2110

630.

76(8

.0)

The

data

inpa

rent

hese

spr

esen

tre

lati

vest

anda

rdde

viat

ion,

n=

3.a

Pea

knu

mbe

rin

Fig

.1.

br 0

.05,1

=0.

997.

L. Hong, H. Altorfer / J. Pharm. Biomed. Anal. 29 (2002) 263–275274

of linear regression between concentrations of theradiolysis products and the irradiation doses arehigher than 0.997 (r0.05,1, the critical value at 95%confidence interval), especially for the main radi-olysis products. The main radiolysis products ofthe EOB irradiated at 25 kGy (Table 4) exist atthe levels of about 10 ppm. An immediate ques-tion is whether or not these radiolysis productswill cause unacceptable toxicity at such a concen-tration level.

All the radiolysis products of EOB detected inthe present work are volatile organic compounds.They can be regarded as volatile residual solventsin pharmaceuticals. According to the ImpuritiesGuideline for Residual Sol�ents developed by In-ternational Conference on Harmonization (ICH)[18], safety risks of residual solvents to humanhealth are placed into three classes. Class 1 resid-ual solvents should not be contained in pharma-ceuticals, and classes 2 and 3 residual solvents inpharmaceuticals should be controlled at the levelslower than those required in the guideline. Thelimits of some compound, which are involved inthe present work, are excerpted from the guidelinein Table 5.

The present result shows that none of radiolysisproducts from EOB belongs to the compounds inclasses 1 and 9 of them are listed in classes 2 and3. Comparison of Tables 4 and 5 shows that theconcentrations of the nine radiolysis compoundsat 25 kGy (the reference dose for pharmaceuticalsterilization), or even at a higher dose of 50 kGy,are much lower than their relevant thresholds in

ICH guideline. This means that the nine radiolysisproducts are at safety levels for human health.

The rest 32 radiolysis products are hydrocarboncompounds, which not only belong to a lowertoxicological class, but also are at safety concen-tration levels for pharmaceutical products. Thehighest concentration among the 32 radiolysisproducts is �5 ppm at 25 kGy. According toICH guideline, the concentration of 5 ppm ismuch lower than the safety limits for classes 2 and3, even for class 1. For example, the very toxiccompounds in class 1, benzene (carcinogen) and1,2-dichloromethene (toxic and environmentalhazard) have the concentration limits of 2 and 5ppm, respectively [18].

Therefore, the both qualitative and quantitativedata ensure that all these volatile radiolysis prod-ucts have a low toxic potential, and are at thesafety levels acceptable for human use at thereference radiation dose of 25 kGy.

4. Conclusion

HS–GC is an appropriate technique to deter-mine trace volatile radiolysis products in irradi-ated eye ointment. Forty-one radiolysis products,14 alkanes, 18 alkene, five cycloparaffin, and fourothers, are found. Acetone, 2-propanol and 2-bu-tanone are the unique radiolysis products of woolfat, and cycloparaffins are the unique radiolysisproducts of white petrolatum and liquid paraffin.The common radiolysis products of the three

Table 5Safety limits of residual solvents in pharmaceuticals for human use by International Conference on Harmonization (ICH) guideline

Peak no. Name Class PDEa (mg/day) Concentration limit (ppm)

9 n-Pentane 3 50 5000Acetone 313 50 5000

50002-Propanol 5014 3n-Hexane 220 2.9 290

23 1-Propanol 3 50 500025 2-Butanone 3 50 5000

Cyclohexane 228 38.8 3880n-Heptane 3 50 500031Methylcyclohxane 2 11.8 118035

a Permitted daily exposure.

L. Hong, H. Altorfer / J. Pharm. Biomed. Anal. 29 (2002) 263–275 275

materials are straight and branch chain hydrocar-bon, which appear regularly as homologues in sixgroups. The radiolysis products of EOB are atotal contribution of the radiolysis products of itsingredients (white petrolatum, liquid paraffin andwool fat).

Ruptures of the long carbon chains are themain degradation pathways of EOB as well as itsingredients. Variation in molecular structure andmatrix composition has little influence on theradiolysis behavior of these materials.

The radiolysis product profile of irradiatedEOB determined by HS–GC is quite similar to atypical mass spectrum of long carbon chain n-alkane, suggesting that the radiolysis mechanismapproximates the degradation mechanism in massspectrometer. The fragment pattern of n-alkane inmass spectrometer can be used as a theoreticalprediction for the radiolysis product study.

Gamma radiation treatment imposes no majorinfluence on the composition of petrolatum eyeointment, and no significant condensation reac-tion can be observed. Both qualitative and quanti-tative data demonstrate that the volatile radiolysisproducts in irradiated petrolatum EOB are safefor human health at the standard irradiation doseof 25 kGy. Thus, it is feasible to perform gammasterilization on petrolatum eye ointment, whitepetrolatum, liquid paraffin and wool fat.

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