FINAL DRAFT UGANDA 
STANDARD  
  FDUS EAS 338 
First  Edition  
2013-mm-dd 
 
 Reference number  
FDUS EAS 338: 2013 
© UNBS 201 3 
  
Chemical hair relaxers and hair waving products — 
Specification  
 FDUS EAS 338: 2013 
ii                © UNBS  2013 - All rights reserved 
 Compliance with this standard does not, of itself confer immunity from legal obligations  
A Uganda Standard does not purport to include all necessary provisions of a contract. Users are 
responsible for its correct application  
© UNBS 2013 
All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilised in any 
form or by any means, electronic or mechanical, including photocopying and microfilm, without prior written 
permission from UNBS. 
Requests for permission to reproduce this document should be addressed to 
The Executive Director 
Uganda National Bureau of Standards 
P.O. Box 6329 
Kampala 
Uganda 
Tel: 256 41 505 995 
Fax: 256 41 286 123 
E-mail: unbs@infocom.co.ug  
Web: www.unbs.go.ug  
 FDUS EAS 338: 2013 
© UNBS  2013 - All rights reserved                iii National foreword 
Uganda National Bureau of Standards (UNBS) is a parastatal under the Ministry of Tourism, Trade and 
Industry established under Cap 3 27, of the Laws of Uganda. UNBS is mandated to co-ordinate the elaboration 
of standards and is  
(a) a member of International Organisation for Standardisation (ISO) and  
(b) a contact point for the WHO/FAO Codex Alimentarius Commission on Food Standards, and  
(c) the National Enquiry Point on TBT/SPS Agreements of the World Trade Organisation (WTO). 
The work of preparing Uganda Standards is carried out through Technical Committees. A Technical 
Committee is established to deliberate on standards in a given field or area and consists of representatives of 
consumers, traders, academicians, manufacturers, government and other stakeholders. 
Draft Uganda Standards adopted by the Technical Committee are widely circulated to stakeholders and the 
general public for comments. The committee reviews the comments before recommending the draft standards 
for approval and declaration as Uganda Standards by the National Standards Council. 
This Final Draft Uganda Standard, FDUS EAS 338: 2013, Chemical hair relaxers and hair waving products  — 
Specification , is identical with  and has been r eproduced from an East African  Standard, EAS 338: 2013, 
Chemical hair relaxers and hair waving products  — Specification , and is being proposed for adoption as a 
Uganda Standard.  
Wherever the words, “ East African Standard" appear, they should be replaced by "Uganda Standard."  EAS 338: 2013
ICS 71.100.70  
© EAC 2013                                                                                                                        Second Edition 2013  
 
EAST AFRICAN STANDARD 
 
Chemical hair relaxers and hair wa ving products — Specification 
EAST AFRICAN COMMUNITY  EAS 338: 2013 
ii © EAC  2013  – All rights reserved  
 Copyright notice 
This EAC document is copyright-protected by EAC. While the reproduction of this document by 
participants in the EAC standards development process is permitted without prior permission from EAC, 
neither this document nor any extract from it may be reproduced, stored or transmitted in any form for 
any other purpose without prior written permission from EAC. 
Requests for permission to reproduce this document for the purpose of selling it should be addressed 
as shown below or to EAC’s member body in the country of the requester:  
© East African Community 2013 — All rights reserved 
East African Community  
P.O.Box 1096 
Arusha 
Tanzania 
Tel: 255 27 2504253/8 
Fax: 255 27 2504481/2504255 
E-mail: eac@eachq.org 
       Web: www.eac-quality.net 
Reproduction for sales purposes may be subject to royalty payments or a licensing agreement. 
Violators may be persecuted EAS 338: 2013 
© EAC 2013  – All rights reserv ed iii 
 Contents Page 
Foreword .............................................................................................................................................................  v 
1 Scope  ................................ ................................ ................................ ................................ ...................... 1 
2 Descrip tion ................................................................ ................................ ................................ ............. 1 
3 Normative references  ................................ ................................ ............................................................  1 
4 Terms and definitions  ................................ ................................ ...........................................................  1 
5 Requ irements  ................................................................ ................................ ................................ ......... 2 
5.1 General requirements  ................................ ................................ ...........................................................  2 
5.2 Specific quality requirements  ................................ ................................ ................................ .............. 2 
6 Packaging and labelling ................................ ................................ ................................ ........................ 3 
6.1 Packaging  ................................ ................................ ................................ ................................ ............... 3 
6.2 Labelling  ................................ ................................ ................................ ................................ ................. 4 
7 Sampling  ................................ ................................ ................................ ................................ ................ 5 
Annex A (normative)  Determination of thermal stability ............................................................................... 6  
A.1 Apparatus  ................................ ................................ ................................ ................................ ............... 6 
A.2 Procedure  ................................ ................................ ................................ ................................ ............... 6 
A.3 Results  ................................ ................................ ................................ ................................ .................... 6 
Annex B (normative)  Determination of total viable count ............................................................................. 7  
B.1 Outline of the method  ................................ ................................ ...........................................................  7 
B.2 Apparatus  ................................ ................................ ................................ ................................ ............... 7 
B.3 Medial buffer  ................................................................ ................................ ................................ .......... 7 
B.4 Sterilization of apparatus  ................................ ................................ ................................ ..................... 8 
B.5 Procedure  ................................ ................................ ................................ ................................ ............... 8 
B.6 Expression of results  ................................ ................................ ............................................................  8 
Annex C (normative)  Determination of Pseudomonas aeruginosa, Staphylococcus  aureus  and 
Candida albicans  in cosmetic products  ................................ ..............................................................  9 
C.1 Introduction  ................................................................ ................................ ................................ ............ 9 
C.2 Principle of the method ................................ ................................ ................................ ......................... 9 
C.3 Apparatus  ................................ ................................ ................................ ................................ ............... 9 
C.4 Reagents  ................................ ................................ ................................ ................................ .............. 10 
C.5 Micro-organisms  ................................ ................................................................ ................................ .. 11 
C.6 Product quality check  ................................ ................................ .........................................................  11 
C.7 Product preparation  ................................ ................................ ............................................................  12 
C.8 Bacterial inoculum preparation  ................................ ................................ ................................ ......... 12 
C.9 Fungal inoculum preparation  ................................ ................................ ................................ ............. 12 
C.10   Inoculum pools  ................................ ................................................................ ................................ ... 12 
C.11  Inoculation  ................................................................ ................................ ................................ ........... 13 
C.12   Sampling intervals  ................................ ................................ ..............................................................  13 
C.13   Aerobic Plate Count (APC)  ................................ ................................ ................................ ................ 13 
C.14   Neutralization check ................................ ................................ ...........................................................  13 
C.15   Data analysis  ................................................................ ................................ ................................ ....... 14 
Annex D (normative)  Determination of pH .................................................................................................... 15 
D.1 Apparatus  ................................ ................................ ................................ ................................ ............. 15 
D.2 Procedure  ................................ ................................ ................................ ................................ ............. 15 
Annex E (normative)  Determination of free alkali content .......................................................................... 16 
E.1 Outline of the method  ................................ ................................ .........................................................  16 
E.2 Reagents  ................................ ................................ ................................ ................................ .............. 16 EAS 338: 2013 
iv © EAC  2013  – All rights reserved  
 E.3 Procedure  ................................ ................................ ................................ ............................................  16 
E.4 Calculation  ................................ ................................ ................................ ...........................................  16 
Annex F (normative)  Determination of total alkali content ......................................................................... 18 
F.1 Outline of the method  ................................................................ ................................ .........................  18 
F.2 Reagents  ................................ ................................ ................................ ..............................................  18 
F.3 Apparatus  ................................ ................................ ................................ ............................................  18 
F.4 Procedure  ................................ ................................ ................................ ............................................  18 
F.5 Calcula tion ................................ ................................ ................................ ...........................................  18 
Annex G (normative)  Determination of thioglycollic acid .......................................................................... 20 
G.1 Reagents  ................................ ................................ ................................ ..............................................  20 
G.2 Procedure  ................................ ................................ ................................ ............................................  20 
G.3 Calculation  ................................ ................................ ................................ ...........................................  20 
Annex H (normative)  Test for lead using atomic absorption spectrophotometer (AAS) ........................  21 
H.1 Scope  ................................ ................................ ................................................................ ...................  21 
H.2 Reagents  ................................ ................................ ................................ ..............................................  21 
H.3 Standard solution s ................................................................ ................................ .............................  21 
H.4 Sample preparation  ................................................................ ................................ ............................  21 
H.5 Instrument conditions  ................................................................ ................................ ........................  22 
H.6 Analysis  ................................ ................................ ................................ ...............................................  23 
H.7 Calculations  ................................ ................................ ................................ ................................ ......... 23 
H.8 Interferences  ................................ ................................ ................................ ................................ ....... 23 
Annex I (normative)  Test for arsenic using atomic absorption spectrophotometer (AAS) ....................  24 
I.1 Scope  ................................ ................................ ................................................................ ...................  24 
I.2 Reagents  ................................ ................................ ................................ ..............................................  24 
I.3 Instrument conditions  ................................................................ ................................ ........................  24 
I.4 Sample preparation  ................................................................ ................................ ............................  25 
I.5 Analysis  ................................ ................................ ................................ ...............................................  25 
I.6 Calculation  ................................ ................................ ................................ ...........................................  25 
Annex J (normative)  Test for mercury using the atomic absorption spectrophotometer (AAS) ...........  26 
J.1 Scope  ................................ ................................ ................................................................ ...................  26 
J.2 Reagents  ................................ ................................ ................................ ..............................................  26 
J.3 Instrument conditions  ................................................................ ................................ ........................  26 
J.4 Sample preparation  ................................................................ ................................ ............................  27 
J.5 Analysis  ................................ ................................ ................................ ...............................................  27 
J.6 Calculations  ................................ ................................ ................................ ................................ ......... 27 
 EAS 338: 2013 
© EAC 2013  – All rights reserv ed v 
 Foreword 
Development of the East African Standards has been necessitated by the need for harmonizing requirements 
governing quality of products and services in the East African Community. It is envisaged that through 
harmonized standardization, trade barriers that are encountered when goods and services are exchanged 
within the Community will be removed.  
In order to achieve this objective, the Community established an East African Standards Committee mandated 
to develop and issue East African Standards.   
The Committee is composed of representatives of the National Standards Bodies in Partner States, together 
with the representatives from the private sectors and consumer organizations. Draft East African Standards 
are circulated to stakeholders through the National Standards Bodies in the Partner States. The comments 
received are discussed and incorporated before finalization of standards, in accordance with the procedures 
of the Community. 
East African Standards are subject to review, to keep pace with technological advances. Users of the East 
African Standards are therefore expected to ensure that they always have the latest versions of the standards 
they are implementing .  
EAS 338 was prepared by Technical Committee EAS/TC 071, Cosmetics and cosmetic products . 
This second edition cancels and replaces the first edition (EAS 338:2004), which has been technically revised  
 
 EAST AFRICAN STANDARD  EAS 338:2013  
 
© EAC 2013  – All rights reserved  1 
 Chemical hair relaxers and hair waving products — Specification 
1 Scope 
This East African Standard specifies requirements and methods of sampling and test for chemical hair 
relaxers and hair waving products. 
This East African Standard applies to chemical cream hair relaxers based on alkalis or thioglycollates, as well 
as hair waving (curling) products based on thioglycollates. 
2 Description 
Chemical hair relaxers and waving products are grouped into three categories, namely: 
a) lye-based;  
b) no lye; and 
c) thioglycollate-based. 
3 Normative references 
The following referenced documents are indispensable for the application of this document. For dated 
references, only the edition cited applies. For undated references, the latest edition of the referenced 
document (including any amendments) applies. 
EAS 346, Labelling of cosmetics — General requirements  
EAS 377-1 , Cosmetics and cosmetic products  Part 1: List of substances prohibited in cosmetic products 
EAS 377-2 , Cosmetics and cosmetic products  Part 2: List of substances which cosmetic products must not 
contain except subject to the restrictions laid down 
EAS 377- 3, Cosmetics and cosmetic products  Part 3: List of colorants allowed in cosmetic products  
EAS 377- 4, Cosmetics and cosmetic products  Part 4: List of preservatives allowed in cosmetic products  
EAS 377- 5, Cosmetics and cosmetic products  Part 5: Use of UV filters in cosmetic products 
ISO 6887- 1, Microbiology of food and animal feeding stuffs — Preparation of test samples, initial suspension 
and decimal dilutions for microbiological examination — Part 1: General rules for preparation of the initial 
suspensions and decimal dilutions  
4 Terms and definitions 
For the purposes of this East African Standard, the following terms and definitions shall apply. EAS 338: 2013 
2 © EAC  2013  – All rights reserved  
 4.1 
lye based 
chemical hair relaxers that contain sodium hydroxide 
4.2 
no lye 
chemical hair relaxers that contain alkalis other than sodium hydroxide such as potassium hydroxide, lithium 
hydroxide, calcium hydroxide 
5 Requirements 
5.1 General requirements 
5.1.1  Chemical hair relaxers and waving products shall comply with the requirements specified in all parts 
of EAS 377.  
5.1.2  The products shall be in the form of emulsions or gels .  
5.1.3  The se products shall be homogenous and of uniform colour, with no visible signs of separation nor 
visible impurities.  
5.2 Specific quality requirements 
5.2.1  Hair relaxers and hair waving products shall pass the test for thermal stability when tested in 
accordance with Annex A. 
5.2.2  The hair relaxers and hair waving products shall also comply with the microbiological limits given in 
Table 1 when tested in accordance with the methods indicated therein. 
Table 1   Microbiological limits  
S/N Micro -organisms  Limits, max.  
Cfu/g  Method of test  
i.  Total viable count  100 in 0.5 g  Annex B  
ii.  Pseudomonas aeruginosa  
Not detect ed in 0.5 g  Annex C  
iii.  Staphylococcus aureus  
iv.  Candida albicans  
 
 
5.2.3  The lye and no lye relaxers (Type 1 and Type 2) shall, in addition, comply with the requirements given 
in Table 2 when tested in accordance with the methods indicated therein.  EAS 338: 2013 
© EAC 2013  – All rights reserv ed 3 
 Table 2 — Chemical requirements for lye and no lye chemical hair relaxers  
S/N Characteristic  Requirements  Method of test  
i.  pH range  11- 13 Annex D 
ii.  Free alkali content, % by mass, max . 2.5 Annex E 
iii.  Total alkali content:  
 Potassium hydroxide , KOH , % by 
mass, max . 
 Sodium hydroxide , NaOH , % by mass, 
max. 
 Lithium hydroxide , LiOH , % by mass, 
max. 
 Calcium hydroxide , Ca(OH)2, % by 
mass, max . 
 Ammonium hydroxide , NH 4OH, % by 
mass, max . 2.5 
2.5 
2.5 
7.0 
6.0 Annex F 
 
5.2.4  The thioglycollate based hair relaxers and hair waving products shall, in addition, comply with the 
requirements given in Table 3 when tested in accordance with the methods indicated therein.  
Table 3 — Requirements for thioglycollate based hair relaxers and hair waving products 
S/N Characteristic  Requirements  
Method of test  
Thioglycollic 
acid and its salts  Thioglycollic 
acid esters  
i.  pH range  7- 9.5 6-9.5 Annex D 
ii.  Thioglycollic acid, %, max . 7- 11 7-11 Annex  G 
 
5.2.5  The products shall comply with the limits for heavy metal contaminants in accordance with Table 4. 
Table 4 — Limits for heavy metal contaminants 
S/N Characteristic  Limits  Method of test  
i.  Lead, ppm, max . 20 Annex H 
ii.  Arsenic, ppm, max . 2 Annex I 
iii.  Mercury, ppm, max . 2 Annex J 
NOTE  The total amount of heavy metals as lead, mercury and arsenic, in combination, in the finished 
product should not exceed 20 ppm.  
6 Packaging and labelling 
6.1 Packaging 
The product shall be pack aged in suitable well-sealed containers that shall protect the contents and shall not 
cause any contamination or react with the product. EAS 338: 2013 
4 © EAC  2013  – All rights reserved  
 6.2 Labelling 
6.2.1 The labelling shall be in English, Kiswahili or French or in combination as agreed between the 
manufacturer and supplier. 
6.2.2 The labelling shall comply with the requirements of EAS 346 
6.2.3 In addition, each package shall be legibly and indelibly marked with the following (an enclosed leaflet 
may also be used): 
a) type of product as follows: 
i. mild — for fine textured or processed hair; 
ii. regular — For medium textured hair; 
iii. super — For coarse hair;  
NOTE This does apply in the case of thioglycollic based formulations. 
b) type of alkali used; 
c) instructions for use including the neutralising shampoo and conditioner which are recommended for 
use with the product; 
d) precautions/warnings: 
i. for products containing thioglycollic acid, its salts and its esters, the warnings include: 
 ―For professional use only‖ ; 
 ―Contains thioglycollate ‖; 
 ―Follow the instructions ‖; 
 ―Avoid contact with eyes ‖; 
 ―In the event of contact with eyes, rinse immediately with plenty of water and seek medical 
advice ‖; 
 ―Wear suitable gloves ‖; and  
 ―Keep out of reach of children ‖. 
ii. for lye-based and no lye relaxers, the  warnings include: 
 ―For professional use only ‖; 
 ―Contains alkali ‖; 
 ―Keep out of reach of children ‖; 
 ―Follow the instructions ‖; 
 ―Wear suitable gloves ‖; 
 ―Avoid contact with eyes as the product can cause blindness ‖; and  EAS 338: 2013 
© EAC 2013  – All rights reserv ed 5 
  ―In the event of contact with eyes, rinse immediately with plenty of water and seek medical 
advice ‖. 
7 Sampling 
Random samples of the product shall be drawn for test in accordance with ISO 24153 from the market, factory 
or anywhere else.  
 EAS 338: 2013 
6 © EAC  2013  – All rights reserved  
 Annex A 
(normative)  
 
Determination of thermal stability 
A.1 Apparatus 
Thermostatically controlled oven , capable of maintaining a temperature of 37 C ± 1 C 
A.2 Procedure 
Place a fresh, unopened sample of the cream in its original container into a thermostatically controlled oven at 
37 C ±1 C for 48 h, making sure that the sample is securely sealed. If the product is packed in an opaque 
container (for example a tube), remove 50 g of the sample and place into an effectively sealed glass tube or 
vial, and test as above. 
A.3 Results 
The products shall be taken to have passed the test if, on removal from the oven, the following indications of 
instability are not observed: 
a) change of colour; 
b) change of smell or odour; 
c) phase separation; 
d) formation of granules or crystal growth; and 
e) shrinkage due to evaporation of water. EAS 338: 2013 
© EAC 2013  – All rights reserv ed 7 
 Annex B 
(normative) 
 
Determination of total viable count 
B.1 Outline of the method 
The test consists of plating a known dilution of the sample or any digest agar medium (soya bean casein is 
recommended) suitable for the total count of bacteria and fungi after incubating them for a specified period to 
permit the development of visual colonies.  
IMPORTANT  Take precaution in ascertaining that only fresh samples, from carefully sealed containers that 
had not been opened before, are used for this test.  This is very necessary for getting accurate results. 
B.2 Apparatus 
B.2.1 Tubes, of resistant glass, provided with closely fitting metal caps 
B.2.2 Autoclaves , of sufficient size. They shall keep uniform temperature within the chamber up to and 
including the sterilizing temperature of 122 C. They shall be equipped with an accurate thermometer, located 
so as to register the minimum temperature within the sterilizing chamber, a pressure gauge and properly 
adjusted safety valves. 
B.2.3 Petri dishes , of 100 mm diameter and 15 mm depth. The bottom of the dishes shall be free from 
bubbles and scratches and shall be flat so that the medium is of uniform thickness throughout the plate.  
B.2.4 Colony counter , an approved counting aid, such as Quebec colony counter. If such a counter is not 
available, counting may be done with a lens giving a magnification of 1.5 diameter. In order to ensure 
uniformity of conditions during counting, illumination equivalent to that provided by Quebec colony counter 
shall be employed.  
B.2.5 Balance 
B.2.6 pH meter 
B.2.7 Water bath 
B.3 Medial buffer 
B.3.1 Soya bean casein digest agar medium  
Dissolve 15 g of pancreatic digest of casein 5 g of papaic digest of soya bean meal, and 5 g of sodium 
chloride in 100 mL of distilled water contained in a 2-litre beaker by heating in a water bath. Add 15 g of 
powdered agar and continue boiling until the agar is completely digested. Adjust the pH to 7.5 with sodium 
hydroxide solution. Distribute in 20-mL quantities, close the tubes with metal caps and autoclave at 123 C for 
20 min. After autoclaving, store the tubes in a cool place and use them within three weeks.  EAS 338: 2013 
8 © EAC  2013  – All rights reserved  
 B.3.2 Stock solution pH 7.2 phosphate buffer 
Dissolve 34 g of monobasic potassium phosphate in about 100 mL of water contained in a 500-mL volumetric 
flask. Adjust the pH 7.2 ± 0.1 by the addition of 4 % sodium hydroxide solution. Add water to volume and mix. 
Sterilize at 122 C for 20 min. Store under refrigeration. 
B.3.3 Dilute phosphate buffer solution pH 7.2  
Dilute 1 mL of stock solution with distilled water in the ratio of 1:800. Fill 50 mL in each of the conical flask of 
100 mL capacity. Plug the flasks with cotton and sterilize at 122 C for 20 min. 
B.4 Sterilization of apparatus 
B.4.1 Tubes 
These shall be sterilized in the autoclave at 122 C temperature and 1.05 kg/cm2 pressure for 20 min or in a 
hot oven at 160 C for one hour.  
B.4.2 Petri dishes  
These shall be packed in drums and autoclaved at 122 C temperature and 1.05 kg/cm2 pressure for 20 min or 
individually wrapped in kraft paper and sterilized  in a hot air oven at 160 C for one hour.  
B.4.3 Pipettes  
These shall be placed in pipette cones (copper, stainless steel or aluminium) after plugging the breeder end 
with cotton and sterilized in the autoclave at 122 C temperature and 1.05 kg/cm2 pressure for 20 min, or at 
160 C for one hour in the air oven. 
B.5 Procedure 
B.5.1 Melt a sufficient number of soya bean casein digest agar medium tube in a hot water bath and 
transfer while hot in to a constant temperature water bath maintained at 48 C ± 20 C.  
B.5.2 Weigh and transfer aseptically 1 g of the sample to a conical flask containing sterile 50 mL or any 
suitable dilution factors of dilute phosphate buffer at pH 7.2. Shake well. Pipette out in 1 mL portions into three 
sterile petri dishes. Pour melted and cooled (at 45 C) soya bean casein digest agar medium over it, and 
rotate to mix thoroughly. Incubate the plate at 32 C for 72 h in an inverted position.  
B.6 Expression of results 
Get the average number of colonies on soya bean casein digest agar medium plates and determine the 
number of micro-organisms per gram of the sample. If no colony is recovered from any of the plates micro-
organisms can be stated as being less than 50 per gram. For calculations, refer to ISO 6887- 1. EAS 338: 2013 
© EAC 2013  – All rights reserv ed 9 
 Annex C 
(normative) 
 
Determination of Pseudomonas aeruginosa, Staphylococcus  aureus  and Candida 
albicans  in cosmetic products  
C.1 Introduction 
A general aseptic and safety procedures should be followed. Table C.1 gives the results of the interlaboratory 
study supporting the acceptance of the method. 
Table C .1 — Inter-laboratory study results for determination of preservation of non-eye area water-
miscible cosmetic and toiletry formulations 
 
Incidence of false - negatives among total positive samplesa) Incidence of false -positives among total 
negative samplesb) 
Product name  Number  Percentage  Sensitivity 
rate Number  Percentage  Sensitivity 
rate 
Shampoo  2/49 4 96 0/53 0 100 
Conditioner  3/48 6 94 0/54 0 100 
Water -in-oil emulsion  0/52 0 100 1/50 2 98 
Oil-in-water emu lsion 0/51 0 100 0/51 0 100 
All combined  5/200  2 98 1/208  0.5 99.5 
a) False -negative analysis indicates a sample is adequately preserved.  
b) False -positive analysis indicates a sample is not adequately preserved.  
 
C.2 Principle of the method  
Bacteria yeast and mould grown on laboratory media, harvested, calibrated, and inoculated into test products.  
Using serial dilutions and plate counts; the numbers of organisms surviving in the test products are 
determined over time.  Products meeting the specified criteria are considered adequately preserved for 
manufacture and consumer use.  Products not meeting criteria are considered inadequately preserved. 
C.3 Apparatus  
C.3.1 Jars, 2-4 oz wide-mouth, straight-side flint glass ointment jars with linerless metal, polypropylene or 
teflon lined screw caps 
C.3.2 Disposable borosilicate glass culture tubes , 16 mm x 125 mm, with caps 
C.3.3 Disposable borosilicate glass culture tubes, 20 mm x 150 mm, with screw caps 
C.3.4 Petri plates Z , 100 mm x 15 mm 
C.3.5 Sterile 2.2 mL pipettes EAS 338: 2013 
10 © EAC  2013  – All rights reserved  
 C.3.6 Sterile swabs 
C.3.7 Glass beads 
C.3.8 Sterile gauze 
C.3.9 10 l -20 l inoculating loops 
C.3.10  Vortex mixer 
C.4 Reagents  
C.4.1 Preparation 
For convenience, dehydrated media of any brand equivalent in function may be used. Test each lot of medium 
for sterility and growth-promotion using suitable organisms. 
C.4.2 List of reagents 
C.4.2.1 Letheen agar, contains 5.0 g pancreatic digest of casein 1.0 g dextrose, 3.0 g beef extract, 1.0 g 
lecithin, 7.0 g polysorbate 80 g, and 15.0 g agar per L.   Prepare according to manufacturer’s directions.  
Dispense into suitable containers and sterilize by autoclaving at 121 C for 15 min. Final pH should be 7.0  
0.2 at 25 C . Place in 45 C water bath until agar is 45 C  2 C.  Use for pour plates. 
C.4.2.2 D/E Neutralizing broth (Dey/Engley), contains 5.0 g pancreatic digest of casein, 2.5 g yeast 
extract, 10 g dextrose, 1.0 g sodium thiogycollate, 6.0g Na 2 S2O3 . 5H2O, 2.5 g NaHSO 3, 7.0 g lecithin, 5.0 g 
polysobate 80 g, and 0.02 g bromcresol purple per L. 
Prepare according to manufacturer’s directions. Dispense 9  mL or 9.9 mL aliquot into tubes and sterilize by 
autoclaving at 121 C for 15 min.  Final pH should be 7.6  0.2 at 25 C.  Use for aerobic plate count, L, 
dilutions. 
C.4.2.3 Nutrient agar,  contains 5.0 g pancreatic digest of gelatin 3.0 g beef extract, and 15.0 g agar per 
L. Prepare according to manufacturer’s directions. Dispense into tubes and sterilize by autoclaving at 121 C 
for 15 min. Final pH should be 6.8  0.2 at 25 C. Cool in inclined position to form a slant. Use for bacterial 
culture maintenance and inoculum preparation. 
C.4.2.4 Y/M agar (yeast/malt extract),  contains 3.0 yeast extract, 3.0 g malt extract, 5.0 g peptone.10.0 
g dextrose and 20.0 g agar per L. Prepare according to ma nufacturer’s directions.  Dispense into tubes and 
sterilize by autoclaving at 121 C for 15 min.  Final pH should be 6.2  0.2 at 25 C. Cool in slanted position.  
Use for yeast culture maintenance and inoculum preparation. 
C.4.2.5 Potato dextrose agar (PDA), contains 200 g potato infusion, 20.0 g dextrose, and 15.0 g agar 
per L. Prepare according to manufacturer’s directions. Dispense into tubes and sterile by autoclaving at 121 
C for 15 min. Final pH should be 5.6  0.2 at 25 C.  Cool in slanted position. Use for mould culture 
maintenance and inoculum preparation. 
C.4.2.6 0.85 % Saline, dissolve 8.50 g NaCI in water and dilute to 1 L.  Dispense into flasks or bottles 
and sterilize by autoclaving at 121 C for 15 min. 
C.4.2.7 0.85 % Saline with 0.05 % Polysorbate 80,  dissolve 8.5 g NaCI in water, mix in 0.50 g 
polysorbate 80 g, and dilute to 1 L.  Dispense into suitable containers and sterilize by autoclaving at 121 C 
for 15 min. 
C.4.2.8 Barium sulphate standard No. 2   EAS 338: 2013 
© EAC 2013  – All rights reserv ed 11 
 C.4.2.8.1  Prepare a 1.0 % BaCI 2 solution by dissolving 1.0 g BaCI 2. 2H 2O in 100 mL water. Let this be 
referred to as solution 1. 
C.4.2.8.2  Prepare a 1.0 % H 2SO 4  solution by mixing 1.0 mL H 2SO 4 in 100 mL water. Let this be referred to 
as solution 2. 
C.4.2.8.3  Mix 0.2 mL of solution (1) with9.8 mL solution (2), in screw-capped test tube. Cap tightly and store 
in the dark at room temperature. 
C.4.2.9 Barium sulphate standard No. 7, use solutions from C .4.2.8. Mix 0.7 mL of solution C.4.2.8.1, 
with 9.3 mL of solution C .4.2.8.2, in a screw-capped test tube. Cap tightly and store in the dark at room 
temperature. 
C.5 Micro-organisms  
C.5.1 Staphylococcus aureus  — ATCC 6538 
C.5.2 Staphycloccuss epidermidis  — ATCC12228 
C.5.3 Klebsiella pneumoniae   — ATCC10031 
C.5.4 Escherichia coli    — ATCC 8739 
C.5.5 Enterobacter gergoviae  — ATCC 33028 
C.5.6 Pseudomonas aeruginosa  — ATCC 9027 
C.5.7 Burkholderia cepacia   — ATCC25416 
C.5.8 Acinetobacter baumannii  — ATCC 19606 
C.5.9 Candida albicans    — ATCC10231 
C.5.10 Aspergillus niger    — ATCC 16404 
NOTE Environmental micro organisms (s) likely to be contaminants of concern during product manufacture or use be 
included as a separate inoculum. Predominant environmental microbes isolated during manufacturing, equipment 
cleaning, and sanitizing, or from related deionized water systems are used as supplemental test inocula). 
C.6 Product quality check  
C.6.1 Weigh 1.0 g product into a screw-capped culture tube containing 9.0 mL sterile neutralizing broth to 
make a 1:10 dilution.  If necessary to disperse product, add ten to twenty 3-mm diameter glass beads to tube. 
Mix on Vortex mixer until homogeneous.  
C.6.2 Pipette 1.0 mL of the 1:10 dilution into each of four sterile petri plates. Pour 15 mL- 20 mL sterile 
molten Letheen agar (45 C  2 C) into each plate . Mix by rotating plates to disperse the dilution thoroughly. 
Let solidify. 
C.6.3 Invert and incubate 2 plates at 35 C  2 C for 48 h and two plates at 25 C  2 C for five days. 
C.6.4 Count the number of colonies on all plates, add, and multiply by 2.5 to determine the number of 
colony forming units per gram (cfu./g) in the product. 
C.6.5 Save plates to be used for the neutralization validation by refrigerating. EAS 338: 2013 
12 © EAC  2013  – All rights reserved  
 C.7 Product preparation  
C.7.1 Measure 20 mL sterile saline into four sterile jars, C.3.1. Cap tightly and store at room temperature. 
C.7.2 Weigh 20 g product into each of four sterile jars, C.3.1. Cap tightly and store at room temperature. 
C.8 Bacterial inocul um preparation  
C.8.1 Streak each bacteria culture, C.5.1 – C.5.10, onto a nutrient agar slant, C.4.2.3. Incubate for 48 h at 
35 C  2  C. Wash each slant with 5.0 mL sterile saline, loosening the culture from the agar surface. 
Transfer the suspension into a sterile tube. Repeat the wash with second 5.0 mL aliquot of saline. Combine 
washes and mix on Vortex mixer to disperse evenly. 
C.8.2 Adjust each wash with sterile saline to yield a suspension of ca 108 cfu/mL using Mc Farland BaSO 4 
standard No, 2,  C.4.2.8, direct microscopic count, turbidimetry, absorbance, or other method correlated to an 
aerobic plate count (APC), C .13. Perform an APC, C.13, on each suspension to confirm standardization. 
C.9 Fungal inoculum preparation  
C.9.1 Streak  C. albicans,  C.5.9, on 3 slants of Y/M agar, C.4.2.4. Incubate at 25 C  2 C for 48 h. Wash 
each slant sequentially with 5.0 mL aliquot of sterile saline.  Repeat with a second 5.0 mL aliquot of sterile 
saline. Combine washes to produce 10 mL suspension. Mix on Vortex mixer to disperse evenly. 
C.9.2 Adjust the wash with sterile saline to yield a suspension of ca 107 cfu/ml using a Mc Farland Ba SO 4 
standard No. 7, C.4.2.8, direct microscopic count, turbidimetry, absorbance, or other method that has been 
correlated to an APC, C.13. Perform an APC, C.13, on the suspension to confirm standardization. 
C.9.3 Streak  A. Niger , C.5.10, on 5 slants of potato dextrose agar C.3.5. Incubate at 25 C  2 C for 10 
days. Dislodge mould spores by adding 5.0 mL sterile saline containing 0.05 % polysorbate 80 to each tube 
and vigorously rubbing the surface of the agar slant with a sterile swab.  Repeat with a second 5.0 mL aliquot 
in each tube.  Combine the 10 washes to produce 50 mL suspension. Filter into a sterile container through 3- 5 
layers of sterile gauze supported in funnel.  Perform an APC, C.13, using appropriate dilutions. Adjust mould 
suspension to ca 107 per ml using sterile saline.  Use immediately or refrigerate at 2 C - 5 C for up to 1 
month. Verify mould viability by an APC, C.13, before each use. 
C.10  Inoculum pools  
C.10.1 Pool equal parts of the S. aureus  and S . epidermidis   suspensions, C .8.2 in a sterile container to 
make incoculum pool 1: Gram-postive cocci. 
C.10.2 Pool equal parts of the K pneumoniae  , E . coli, and E . gergoviae  suspensions, C .8.2, in a sterile  
container to make inoculum pool  2: Gram- negative fermentors. 
C.10.3 Pool equal parts of the P. aeruginosa , B .cepacia  and A. baumanii suspensions, C .8.2, in a sterile 
container to make Inoculum Pool 3: Gram-negative nonfermentors.  
C.10.4 Pool equal parts of C. Albicans, C.9.2, and A. Niger, C.9.3, suspensions in a sterile container to make 
innoculum pool 4: Fungi. 
C.10.5 Use organism pools immediately or refrigerate them at 2 C - 5 C for more than 72 h. EAS 338: 2013 
© EAC 2013  – All rights reserv ed 13 
 C.11 Inoculation 
C.11.1 Inoculate each of the four 20.0 mL aliquots of sterile saline, C .7.1, with 0.2 mL of its respective 
inoculum pool, C.10.1 - C.10.4. Mix thoroughly.  Use these suspensions to determine inoculums counts (see 
Ka). 
C.11.2 Inoculate each of the four 20 g products suspensions, with 2.0 mL of its respective inoculums, C.10.1 
– C.10.4. Mix thoroughly by shaking, Vortex mixing or stirring, so that each suspension contains 106 bacteria 
or 10 5 fungi per gram, evenly distribute throughout the product. Tightly close inoculated containers and store 
at ambient temperature (20 C - 25 C). 
C.12  Sampling intervals  
C.12.1 Sample each inoculated saline suspension, C.11.1, for APC, C.13 , within 1 h after inoculation to 
obtain inoculum count. 
C.12.2 Test each inoculated product, C .11.2 for APC, C.13, at 7, 14 and 28 days after inoculation to obtain 
product interval count. 
C.13  Aerobic Plate Count (APC) 
C.13.1 Mix suspension thoroughly. Weigh 1.0 g product into screw-capped culture tube containing 9.0 mL 
sterile neutralizing broth for a 1:10 dilution. If necessary to disperse product, add 10 - 20 sterile 3 mm 
diameter glass beads to the tube. Mix on Vortex mixer until homogeneous. 
C.13.2 Aseptically pipette 0.1 mL of the 1: 10 dilution into 9.9 mL tube of neutralizing broth to obtain a 1:1000 
dilution. Vortex mix. Pipette 0.1 mL of the 1:1000 dilution into 9.9 mL neutralizing broth to obtain a 1: 100 000 
dilution. 
The number of dilutions may be decreased if previous counts microbial populations show reduction. 
C.13.3 Using a 2.2 mL pipette, aseptically pipette 1.0 mL and 0.1 mL aliquots from the 1:10 dilution into 
duplicate petri dishes for the 1:10 and 1:100 plates. If necessary, transfer duplicate 1.0 mL and 0.1 mL 
aliquots from the 1: 1000 dilution for plates 1:1000 and 1:10 000, and from the 1:100 000 dilution for plates 
1:100 000 and 1:1000 000. Pour 15 mL - 20 mL sterile Letheen agar C.3.1, (45 C  2 C into each plate. Mix 
by rotating the plates to disperse the suspension thoroughly, and let solidify. 
C.13.4 Invert bacterial plates and incubate at 35 C  2 C.  Examine bacterial plates after 48 h - 72 h.  Count 
in suitable range (30 - 300 colonies). If no countable plates fall in that range, count the plate(s) nearest that 
range showing distinct colonies. Average duplicate plates counts and express results as cfu/g of product. 
C.13.5 Invert and incubate fungal plates at 25 C  2 C.  Read fungal plates at 2 - 3 days and record results. 
Count plates in a suitable range (30 - 300 colonies). If no countable plates fall in that range, count the plate(s) 
nearest that range showing distinct colonies. Re-incubate plates for another 2 - 3 days. Read and record 
additional colonies. Add to previous results to obtain total counts. Average duplicate plate counts and record 
as cfu/g of products.   
C.14  Neutralization check 
Make a 1:10 000 dilution in sterile saline of pools 1, 2 and 3, C.10 .1 - C.10 .3, and a 1:1000 dilution of pool 4, 
C.10.4. Streak each dilution for isolation with a 10 l loop on the plates saved from C.6.5.  If plates are not 
usable due to either desiccation or surface growth, repeat C.6, and streak freshly prepared plates.  Incubate 
as in C .13.4 - C .13.5. EAS 338: 2013 
14 © EAC  2013  – All rights reserved  
 C.15  Data analysis 
C.15.1 Product quality check , C.6 .4, should be found to contain, <100 cfu/g to proceed with the challenge 
test. 
C.15.2 Inoculums counts , C.12.1, should be between 1 to 9.9 x 106 cfu /g product for bacteria and 1 to 9.9 x 
105 cfu/g product for fungi, or the test should be repeated with different dilutions. 
C.15.3 Neutralization check , C.14, should show significant growth of all pools to confirm adequate 
neutralization.  A neutralizing broth other than D/E broth can be used.  If neutralization does not occur, the test 
is invalid.  Refer to references 4-6 in AOAC official method 998.10 for assistance. 
C.15.4 Calculate the percentage reduction: 
count Inoculum100 count interval product count Inoculum% Reduction,
 
 
C.15.5 The test product is considered adequately preserved if  
a) bacteria show at least 99.9 % (3 log) reduction within one week following challenge and remain at or 
below that level thereafter, and  
b) fungi show at least a 90 % (1 log) reduction within one week following challenge, and remain at or below 
that level thereafter. This criteria applies to freshly prepared formulations. 
 EAS 338: 2013 
© EAC 2013  – All rights reserv ed 15 
 Annex D 
(normative) 
 
Determination of pH 
D.1 Apparatus 
pH meter , equipped with a glass electrode 
D.2 Procedure 
D.2.1 For oil- in-water emulsion creams  
Weigh 5 g ± 0.01 g of the cream in a 100 mL beaker. Add 45 mL of water and disperse the cream in it. 
Determine the pH of the suspension at 25 C using the pH meter. 
D.2.2 For water- in-oil emulsion creams  
Weigh 10 g of the cream to the nearest 0.1 g. Add 90 mL of rectified spirit previously adjusted to pH 6.5 to 7.0. 
Warm, if necessary, to 45 C and stir thoroughly for 15 min. Filter the alcoholic layer through a filter paper and 
measure the pH of the filtrate at 25 C using pH meter. 
Determine the form of cream by placing some of it on spot tile and sprinkling with a mixed indicator consisting 
of an intimate mixture of oil soluble dye of one colour, e.g. oil orange, and a water-soluble dye of a different 
colour e.g. methylene blue. After a few minutes the predominant colour indicates whether the continuous 
phase is oil or water. In case of doubt matter is confirmed by checking whether the product is capable of 
conducting electricity: if so the cream is deemed to be water continuous. EAS 338: 2013 
16 © EAC  2013  – All rights reserved  
 Annex E 
(normative)  
 
Determination of free alkali content 
E.1 Outline of the method 
This method consists of dissolving the sample in alcohol, and titrating against standard acid. 
E.2 Reagents 
E.2.1 Phenolphthalein indicator solution , dissolve 1 g of phenolphthalein in 100 mL of 95 % (v/v) rectified 
spirit. 
E.2.2 Ethyl alcohol, freshly boiled, and neutral to phenolphthalein, 95 % (v/v) 
E.2.3 Standard hydrochloric acid, 0.1 N 
E.3 Procedure 
Dissolve 2 g of hair relaxer in 100 mL of ethyl alcohol by warming, if necessary. Cool and add a few drops of 
phenolphthalein indicator. Titrate with standard hydrochloric acid. 
E.4 Calculation 
The free alkali content shall be calculated as follows: 
Free alkali, % by mass=
.C
MVN  
where 
C is a Constant 
C = 2.4 for LiOH 
C = 4 for NaOH, KOH and LiOH 
C = 5.6 for KOH 
C = 3.7 for Ca(OH) 2 
C = 3.5 for NH 4OH 
V is the volume, in millilitres, of standard hydrochloric acid; 
N is the normality of standard hydrochloric acid; and 
M is the mass, in grams, of the sample. EAS 338: 2013 
© EAC 2013  – All rights reserv ed 17 
 Where mixtures of sodium, lithium and potassium hydroxide are present in the product, the free alkali content 
shall not exceed 2.5 % by mass when calculated as sodium hydroxide. EAS 338: 2013 
18 © EAC  2013  – All rights reserved  
 Annex F 
(normative) 
 
Determination of total alkali content 
F.1 Outline of the method 
The method consists of refluxing the sample in alcohol, and titrating with standard acid. 
F.2 Reagents 
F.2.1 Ethanol , 95 % (v/v) solution, free from carbon dioxide. Reflux this solution for 5 min. Cool to room 
temperature and neutralize with the ethanolic potassium hydroxide solution per 200 mL of ethanol. 
F.2.2 Hydrochloric acid , 1 N standard solution 
F.2.3 Ethanolic potassium hydroxide , 0.1 N ethanolic standard solution 
F.2.4 Phenolphthalein solution , of 1 g per 100 mL in 95 % (v/v) ethanol 
F.3 Apparatus 
F.3.1 Conical flask capacity 250 mL , with a conical ground-glass joint at the bottom 
F.3.2 Reflux condenser , water cooled, with a conical ground-glass joint at the bottom 
F.3.3 Analytical balance 
F.4 Procedure 
Weigh accurately 2 g of the sample into the flask. Add 100 mL of the ethanol solution to the test portion. Fit 
the flask to the reflux condenser, and gently heat until the sample is completely dissolved. 
Add 3.0 mL of the hydrochloric acid solution and boil gently for 10 min. If after boiling, the pink colour re-
appears, add a further quantity of hydrochloric acid. Titrate, at 70 C, with the ethanolic potassium hydroxide 
solution in the presence of phenolphthalein indicator. Carry out two determinations on the same sample. 
F.5 Calculation 
The total alkali content shall be calculated as follows: 
Total alkali content = 
C
MNVNVx11 00  
where 
C is a Constant EAS 338: 2013 
© EAC 2013  – All rights reserv ed 19 
 C = 2.4 for LiOH 
C = 4 for NaOH,  
C = 5.6 for KOH 
C = 3.7 for Ca(OH) 2 
C = 3.5 for NH 4OH 
V0 is the volume, in millilitres, of hydrochloric acid used; 
V1  is the volume in millilitres of potassium hydroxide used in titration; 
N0 is the exact normality of hydrochloric acid; and 
N1  is the exact normality of the potassium hydroxide solution used. 
M is the mass, in grams, of the sample. 
Where mixtures of sodium, lithium and potassium hydroxide are present in the product, the total alkali content 
shall not exceed 2.5 % by mass, calculated as sodium hydroxide. 
 
 EAS 338: 2013 
20 © EAC  2013  – All rights reserved  
 Annex G 
(normative) 
 
Determination of thioglycollic acid 
G.1 Reagents 
G.1.1 Concentrated hydrochloric acid 
G.1.2 Standard iodine solution , 0.1 N 
G.2 Procedure 
Accurately weigh about 5 g of the sample in a 250 mL conical flask. Add about 75 mL of water and 15 mL of 
concentrated hydrochloric acid and heat on a water bath for 10 min. Cool and titrate with iodine solution using 
starch solution as the indicator. 
G.3 Calculation 
Calculate on the basis that each ml of 0.1 N iodine solution is equivalent to 0.00921 g of thioglycollic acid. 
% of thioglycollic acid, m/m = 
MV 100x 0.00921x  
where 
M is the mass, in gram s, of sample taken for test ; and  
V is the volume, in millilitres, of 0.1 N iodine solution used 
 EAS 338: 2013 
© EAC 2013  – All rights reserv ed 21 
 Annex H 
(normative) 
 
Test for lead using atomic absorption spectrophotometer (AAS) 
H.1 Scope 
This method describes the determination of lead in various cosmetic products. No interference occurs from 
the high concentrations of bismuth which do interfere in the dithzone calorimetric procedure. 
H.2 Reagents 
H.2.1 Lead nitrate,  Pb(NO 3)2 
H.2.2 Dimethylacetamide  DMA  
H.2.3 Nitric acid HNO 3, 3N. Prepare by diluting 195 mL of concentrated HNO 3 (15.4 N) to one litre with 
deionized water. 
H.2.4 Ethanol  C 2H5OH  
H.2.5 Bismuth oxychloride  BiOCl 
H.2.6 Hydrochloric acid  HCl, 6 N. Prepare by diluting 516 mL of concentrated HCl (11.6 N) to one litre with 
deionized water. 
H.2.7 Hydrochloric acid  HCl, 2 N. Prepare by diluting 172 mL of concentrated HCl (11.6 N) to one litre with 
deionized water. 
H.2.8 Hydrochloric acid  HCl, 0.5 N. Prepare by diluting 25 mL of 2 N HCl to 100 mL with deionized water. 
H.3 Standard solutions 
H.3.1 Lead standard solution , 100 μg/mL. Dissolve 0.1598 g of Pb(NO 3)2 in 10 mL of dilute HNO 3 and 
dilute to 1000 mL with deionized water . 
H.3.2 Lead standard solution , 1000 μg/mL in DMA. Dissolve 0.1598 g of Pb(NO 3)2 in DMA. Dilute to 100 
mL.  
H.4 Sample preparation 
H.4.1 Aerosols 
For hair, deodorant spray or similar aerosols, weigh accurately about 5 g of sample and dissolve in 50 mL of 
ethanol. For shaving cream spray or similar aerosols weigh accurately about 1 g of sample and dissolve in 50 
mL of ethanol. EAS 338: 2013 
22 © EAC  2013  – All rights reserved  
 H.4.2 BiOCl or cosmetics containing BIOCl 
Dissolve 1 g of sample (5 g if the lead level is expected to be less than 10 μg/g) in 15 mL of 6 N HCI and dilute 
to 100 mL with 0.5 N HCI. If the sample is coated with an organic material, it is necessary to ignite the sample 
to 500 C to ash before analysis. 
H.5 Instrument conditions 
H.5.1 Standard atomic absorption conditions for lead 
H.5.1.1 Recommended flame air-acetylene, oxidizing (lean, blue)  
H.5.1.2  Data obtained with a standard nebulizer and flow spoiler. Operation with a High sensitivity 
nebulizer or impact bead will typically provide a 2-3x sensitivity improvement.  
H.5.1.3 Characteristic concentration with a N 2O-C 2H2 flame at 283.3 nm: 2.7 mg/L.  
Table H.1 — HCl data EDL sensitivity values for lead 
Wavelength , 
nm Slit  
Nm Relative noise  Characteristic 
concentration , 
mg/L  Characteristic 
concentration 
check ,  
mg/L  Linear range , 
mg/L  
283.3  0.7 0.43 0.45 20.0 20.0 
217.0  0.7 1.0 0.19 9.0 20.0 
205.3  0.7 1.4 5.4 250.0  - 
202.2  0.7 1.8 7.1 350.0  - 
261.4  0.7 0.35 11.0 500.0  - 
368.3  0.7 0.40 27.0 1200.0  - 
364.0  0.7 0.33 67.0 3000.0  - 
 
H.5.2 Standard flame emission conditions for lead 
Stock standard solution, lead, 1000 mg/L. Dissolve 1.598 g of lead nitrate, Pb(NO 3)2 in 1 %(v/v) HNO 3 and 
dilute to 1 litre with 1 % (v/v) HNO 3 
Table H.2 — Flame emission conditions for lead 
Wavelength (nm)  Slit (nm)  Flame  
405.8  0.2 Nitrous oxi de- acetylene  
 
H.5.3 Light sources 
Both Electrodeless Discharge Lamps (EDLS) and Hollow Cathode Lamps are available for lead. EDLs provide 
greater light output and longer life than Hollow Cathode Lamps. For lead, both EDLs and Hollow Cathode 
Lamps provide approximately the same sensitivity and detection limit. With multi-element lamps containing 
copper, the Cu 216.5 nm resonance line may interfere with lead determinations at the lead 217.0 nm line. The 
lead 283.3 nm line should be used instead.  EAS 338: 2013 
© EAC 2013  – All rights reserv ed 23 
 H.5.4 Interferences 
Large excesses of other elements (for example, 10 000 mg/L Fe) May interfere with the lead signal.  
H.6 Analysis 
Determine the concentration of lead in the sample solutions of aerosols and similar products using the 
standard conditions for lead and standards prepared in ethanol. The standard solutions shall be the following 
concentrations: 0 ppm, 10 ppm, 20 ppm, 30 ppm, 40 ppm and 50 ppm. Determine the concentration of lead in 
the BiOCI solutions using the standard conditions for lead and the method of additions. 
H.7 Calculations 
The total amount of lead shall be calculated as follows: 
µg/g Pb =
sampleofgPbμg/mL x 50 
H.8 Interferences 
No interference was found from 1000 μg/mL of Na, Mg, K, or Ca, or from 500 μg/mL of Mn, Co or Ni. A 
background absorption interference was noted with 1000 μg/mL of Al, Fe or Bi, which can be eliminated by 
correcting the lead absorption at 283 nm by any absorption observed at 280 nm, or by using the Deuterium 
Background Corrector. 
 EAS 338: 2013 
24 © EAC  2013  – All rights reserved  
 Annex I 
(normative) 
 
Test for arsenic using atomic absorption spectrophotometer (AAS) 
I.1 Scope 
This method describes the determination of arsenic in various cosmetic products.  
I.2 Reagents 
The reagents used should be of analytical reagent grade. Water shall be distilled or di-ionised.  
I.3 Instrument conditions 
I.3.1 Standard atomic absorption conditions for arsenic 
I.3.1.1  Recommended flame; air-acetylene, reducing (rich, slightly yellow) 
I.3.1.2  Data obtained with a standard nebulizer and flow spoiler. Operation with a High Sensitivity 
nebulizer or impact bead will typically provide a 2-3X sensitivity improvement. 
I.3.1.3  Characteristic concentration with a N 2O-C 2H2 flame at 193.7 M: 1.4mg/L 
I.3.1.4  Table contains EDL data.  HCL sensitivity values are more than 25 % poorer. 
Table I.1 — HCl data EDL sensitivity values for arsenic 
Wavelength ,  
nm Slit,  
Nm Relative noise  Characteristic 
concentration ,  
mg/L  Characteristic 
concentration 
check ,  
mg/L  Linear range , 
mg/L  
193.7  0.7 1.0 1.0 45.0 100.0  
189.0  0.7 1.8 0.78 40.0 180.0  
197.2  0.7 0.95 2.0 90.0 250.0  
 
I.3.2 Stock standard solution 
Arsenic , 1000 mg/L. Dissolve 1.320 g of Arsenious oxide As 2O3, in 25 mL of 20 % (w/v) KOH solution. 
Neutralize with 20 % (v/v) H 2SO 4 to a phenolphthalein endpoint. Dilute to one litre with 1 % (v/v) H 2SO 4. The 
standard solutions shall be of the following concentrations: 0 ppm, 2 ppm, 4 ppm, 6 ppm, 8 ppm and 10 ppm.  
I.3.3 Flames 
The air-acetylene flame absorbs or scatters more than 60 % of the light source radiation at the 193.7 nm 
arsenic line. Flame absorption is reduced with the use of the nitrous oxide-acetylene flame, although EAS 338: 2013 
© EAC 2013  – All rights reserv ed 25 
 sensitivity is also reduced. Use of background correction is recommended, as it will correct for flame 
absorption and thus improve the signal to noise ratio.  
I.3.4 Light sources 
Both HCL and EDL sources are available for arsenic. EDLS, which are more intense, provide better 
performance and longer life. 
I.3.5 Interferences 
Sample with high total salt content (greater than 1 %) can produce non-specific absorption at the 193.7 mn 
arsenic line, even when the metal is absent. It is therefore advisable to set background correction. 
I.4 Sample preparation 
As in H.4.  
I.5 Analysis 
As in H.6.  
I.6 Calculation 
As in H.7 EAS 338: 2013 
26 © EAC  2013  – All rights reserved  
 Annex J 
(normative) 
 
Test for mercury using the atomic absorption spectrophotometer (AAS) 
J.1 Scope 
This method describes the determination of mercury in various cosmetic products.  
J.2 Reagents 
The reagents used should be analytical reagent grade.  Water shall be distilled or de-ionised. 
J.3 Instrument conditions 
J.3.1 Standard atomic absorption conditions for mercury 
J.3.1.1  Recommended flame: air-acetylene, oxidizing (lean, blue) 
J.3.1.2  Data obtained with a standard nebulizer and flow spoiler. Operation with a High Sensitivity 
nebulizer or impact bead will typically provide a 2-3X sensitivity improvement. 
J.3.1.3  Characteristic Concentration with a N 2O-C 2H2 flame at 253.7 nm: 12 mg/L 
J.3.1.4  Table contains EDL data.  HCL sensitivity values more than 25 % poorer  
Table J.1 — HCl data EDL sensitivity values for mercury 
Waveleng th,  
nm Slit,  
nm Relative noise  Characteristic 
concentration ,  
mg/L  Characteristic 
concentration 
check ,  
mg/L  Linear range , 
mg/L  
253.7  0.7 1.0 4.2 200.0  300.0  
 
J.3.2 Standard flame emission conditions for mercury 
Table J.2 — Flame emission conditions for mercury 
Wavelength (nm)  Slit (nm)  Flame  
253.7  0.2 Nitrous oxide - acetylene  
J.3.3 Stock standard solution 
Mercury , 1000 mg/L — Dissolve 1.080 g of mercury (II) oxide, HgO, in a minimum volume of (1 + 1) HCI. 
Dilute to one litre with deionized water. The standard solutions shall be of the following concentrations: 0 ppm, 
2 ppm, 4 ppm, 6 ppm, 8 ppm and 10 ppm. EAS 338: 2013 
© EAC 2013  – All rights reserv ed 27 
 J.3.4 Light sources 
Both Electrodeless Discharge Lamps (EDLS) and Hollow Cathode Lamps are available for Mercury. However, 
the light output of mercury Hollow Cathode Lamp is significantly poorer than with EDLs, and the sensitivity and 
detection limit achieved also are much poorer. In addition, the life of Hollow Cathode Lamps is much shorter. 
J.3.5 Interferences 
Large concentrations of cobalt will absorb at the mercury 253.7 nm resonance line. A 1000 mg/L cobalt  
solution produces approximately 10 % absorption. Ascorbic acid, stannous chloride, or other reducing agents 
may reduce the mercury present to Hg (I) or elemental mercury. These give higher sensitivities than Hg (II), 
and their presence can generate erroneously high results.  
J.4 Sample preparation 
As in H.4.  
J.5 Analysis 
As in H.6.  
J.6 Calculations 
As in H.7. 
 
 
 
 EAS 338: 2013    
 
© EAC 2013 – All rights reserved