2.2.3 limit test for heavy metals draft revision for the …€¦ · 21/08/2018 · 55 draft...
TRANSCRIPT
Working document QAS/18.769
August 2018
Draft for comments
1 2
2.2.3 LIMIT TEST FOR HEAVY METALS 3
Draft revision for The International Pharmacopoeia 4
(August 2018) 5
DRAFT FOR COMMENTS 6
7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22
© World Health Organization 2018 23 24 All rights reserved. 25 26 This draft is intended for a restricted audience only, i.e. the individuals and organizations having received this draft. The draft 27 may not be reviewed, abstracted, quoted, reproduced, transmitted, distributed, translated or adapted, in part or in whole, in any 28 form or by any means outside these individuals and organizations (including the organizations' concerned staff and member 29 organizations) without the permission of the World Health Organization. The draft should not be displayed on any website. 30 31 Please send any request for permission to: 32 33 Dr Sabine Kopp, Group Lead, Medicines Quality Assurance, Technologies Standards and Norms, Regulation of Medicines and 34 other Health Technologies, Department of Essential Medicines and Health Products, World Health Organization, CH-1211 35 Geneva 27, Switzerland, fax: (41 22) 791 4856, email: [email protected]. 36 37 The designations employed and the presentation of the material in this draft do not imply the expression of any opinion 38 whatsoever on the part of the World Health Organization concerning the legal status of any country, territory, city or area or of its 39 authorities, or concerning the delimitation of its frontiers or boundaries. Dotted lines on maps represent approximate border lines 40 for which there may not yet be full agreement. 41 42 The mention of specific companies or of certain manufacturers’ products does not imply that they are endorsed or recommended 43 by the World Health Organization in preference to others of a similar nature that are not mentioned. Errors and omissions 44 excepted, the names of proprietary products are distinguished by initial capital letters. 45 46 All reasonable precautions have been taken by the World Health Organization to verify the information contained in this draft. 47 However, the printed material is being distributed without warranty of any kind, either expressed or implied. The responsibility 48 for the interpretation and use of the material lies with the reader. In no event shall the World Health Organization be liable for 49 damages arising from its use. 50 51 This draft does not necessarily represent the decisions or the stated policy of the World Health Organization. 52
53
Please send any comments you may have on the attached text to Dr Herbert Schmidt, Technical
Officer, Medicines Quality Assurance, Technologies Standards and Norms ([email protected]), with
a copy to Ms Xenia Finnerty ([email protected]) by 30 September 2018.
Medicines Quality Assurance working documents will only be sent out electronically and will
also be placed on the Medicines website for comment under “Current projects”. If you have
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Working document QAS/18.769
page 2
SCHEDULE FOR THE PROPOSED ADOPTION PROCESS OF DOCUMENT QAS/18.769: 54
Draft revision for The International Pharmacopoeia 55
2.2.3 LIMIT TEST FOR HEAVY METALS 56
57
First draft prepared. November 2017
Discussion at informal consultation on new medicines, quality
control and laboratory standards. 2–4 May 2018
Preparation of a revised document based on feedback received at
the informal consultation June – July 2018
Draft revision sent out for public consultation. August – September 2018
Compilation of the feedback received October 2018
Presentation to WHO Expert Committee on Specifications for
Pharmaceutical Preparations. October 2018
Further follow-up action as required.
58
59
[Note from the Secretariat. Feedback is being sought in relation to the revision of the method of 60
analysis 2.2.3 Limit Test for Heavy Metals. It is proposed to change the provision as follows: 61
62
to add a note to provide users of The International Pharmacopoeia with the option to 63
apply ICH Q3D principles to control elemental impurities; 64
to add a new procedure for the preparation of the test solution: procedure 5, a closed-65
vessel microwave digestion that shall be used as an alternative, in particular, for 66
procedures 3 and 4 employing ignition techniques (in new and revised monographs, 67
procedure 5 shall be preferred to procedures 3 or 4); 68
to replace the reagent hydrogen sulfide R by thioacetamide R; and 69
to align parts of text to the corresponding text included in the European Pharmacopoeia 70
(2.4.8.), thereby keeping and further simplifying the structure of the existing text. 71
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72
The aim of the limit test for heavy metals is to control metal contaminants potentially emanating 73
from reagents, solvents, electrodes, reaction vessels, rubber seals, and so on. The test shall 74
serve as a screening tool indicating the overall quality of the production process, with limits 75
usually set at < 10 ppm or 20 ppm Pb. 76
77
The principle of the test is based on the precipitation of metal sulfides and assumes that all 78
metals behave in a similar manner to a lead standard with which samples are compared. As 79
investigations have shown, the test is effective for detecting Pb, Hg, Pd, Ag, V, Au and Cu which 80
give dark brown or black precipitates. The colour of other precipitates varies from pale yellow 81
to orange. 82
83
It is proposed to complement this document by a review of all tests for heavy metals currently 84
prescribed in The International Pharmacopoeia to evaluate the impact of the new provisions on 85
existing test descriptions. In addition, a document could be developed that provides guidance for 86
collaborating laboratories involved in the development of monographs for The International 87
Pharmacopoeia on how to elaborate and validate this limit test. 88
89
Comments are sought in particular on the Note added to the text providing users with the option 90
to assess and control elemental impurities according to ICH Q3D, for example, to accommodate 91
decisions of responsible national or regional authorities. 92
93
Parts of the text are reproduced from the European Pharmacopoeia with permission and with 94
appropriate editorial modifications. 95
96
Changes from the current monograph are indicated in the text by insert or delete.] 97
98
99
Working document QAS/18.769
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Draft revision for The International Pharmacopoeia 100
2.2.3 LIMIT TEST FOR HEAVY METALS 101
102
Note. The Guideline for Elemental Impurities Q3D, published by the International Council for 103
Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH), presents 104
a process to assess and control elemental impurities in finished pharmaceutical products using 105
the principles of risk assessment. It is within a regulatory authority’s remit to decide whether or 106
not they apply this guideline for the assessment of elemental impurities. If ICH Q3D is 107
implemented, compliance of pharmaceutical substances with the limit test for heavy metals will 108
no longer be required. 109
110
The limit test for heavy metals is provided to demonstrate that the content of metallic impurities 111
that are precipitated as coloured sulfides by thioacetamide does not exceed the heavy metals 112
limits given in the individual monographs in terms of micrograms of lead per gram of the test 113
substance. 114
115
The test consists of three consecutive operations: preparation of the test solutions (precedures 1 116
to 5), development of the coloured precipitate by reaction with thioacetamide, and comparison of 117
the colours thus obtained either by directly comparing the coloration of liquids in suitable 118
comparison tubes (Method A) or by comparing the intensity of coloured residues obtained by 119
filtering the liquid using an appropriate apparatus (Methods B or C). Method A is generally 120
applicable only when the amount of heavy metals in the weight of the test substance used 121
exceeds 5 μg; Methods B or C can also be used for amounts of 2 to 5 μg of heavy metals. 122
123
PREPARATION OF THE TEST SOLUTIONS 124
125
For the standard solution, unless otherwise specified, dilute lead PbTS containing 10 µg of lead 126
per mL solvent to obtain a solution containing 1 µg of lead per mL or 2 µg of lead per mL, 127
depending on the limit prescribed in the monograph. Use the solvent used to prepare the sample 128
solution. 129
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130
Procedure 1. For the sample solution, unless otherwise specified in the monograph, weigh the 131
quantity of the substance to be examined and dissolve it in 25 mL of water R. For the reference 132
solution, add 2 mL of the sample solution to 10 mL of the standard solution. For the blank 133
solution, add 2 mL of the sample solution to 10 mL of water R. 134
135
Procedure 2. For the sample solution , unless otherwise specified in the monograph, weigh the 136
quantity of the substance to be examined and dissolve it in about 25 mL of the organic solvent 137
specified in the monograph, containing a minimum percentage of water R (for example, dioxan 138
R containing 15% of water R or acetone R containing 15% of water R). For the reference 139
solution, add 2 mL of the sample solution to 10 mL of the standard solution. For the blank 140
solution, add 2 mL of the sample solution to 10 mL of the solvent used to prepare the sample 141
solution. 142
143
Procedure 3. For the sample solution, place the prescribed quantity (not more than 2 g) of the 144
substance to be examined in a silica crucible with 4 mL of a 250 g/L solution of magnesium 145
sulfate R in sulfuric acid (~98 g/L) TS. Mix using a fine glass rod. Heat cautiously. If the 146
mixture is liquid, evaporate gently to dryness on a water bath. Progressively heat to ignition and 147
continue heating until an almost white or, at most, greyish residue is obtained. Carry out the 148
ignition at a temperature not exceeding 800 °C. Allow to cool. Moisten the residue with a few 149
drops of sulfuric acid (~98 g/L) TS. Evaporate, ignite again and allow to cool. The total period 150
of ignition must not exceed two hours. Take up the residue in two quantities, each of 5 mL, of 151
hydrochloric acid (~70 g/L) TS. Add 0.1 mL of diluted phenolphthalein/ethanol TS, then 152
ammonia (~35 g/L) TS, until a pink colour is obtained. Cool, add anhydrous acetic acid R until 153
the solution is decolorized and add 0.5 mL in excess. Filter if necessary and wash the filter. 154
Dilute with water R to 20 mL. 155
156
For the reference solution, follow the procedure described for the sample solution, using the 157
prescribed volume of dilute lead PbTS containing 10 µg of lead per mL instead of the substance 158
to be examined. To 10 mL of the solution obtained, add 2 mL of the sample solution. 159
160
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For the monitor solution, follow the procedure described for the sample solution, adding to the 161
substance to be examined the volume of dilute lead PbTS prescribed for the preparation of the 162
reference solution. To 10 mL of the solution obtained add 2 mL of the sample solution. 163
164
For the blank solution, add 2 mL of the sample solution to 10 mL of water R. 165
166
Procedure 4. For the sample solution, unless otherwise specified in the monograph, mix 167
thoroughly in a silica crucible the prescribed quantity of the substance to be examined with 0.5 g 168
of magnesium oxide R1. Ignite to a dull redness until a homogeneous white or greyish-white 169
mass is obtained. If after 30 minutes of ignition the mixture remains coloured, allow to cool, mix 170
using a fine glass rod and repeat the ignition. If necessary, repeat the operation. Heat at 800 °C 171
for about one hour. Take up the residue in two quantities, each of 5 mL, of a mixture of equal 172
volumes of hydrochloric acid (~250 g/L) TS and water R. Add 0.1 mL of diluted 173
phenolphthalein/ethanol TS and then ammonia (~35 g/L) TS until a pink colour is obtained. 174
Cool, add anhydrous acetic acid R until the solution is decolorised, then add 0.5 mL in excess. 175
Filter if necessary and wash the filter. Dilute with water R to 20 mL. 176
177
For the reference solution, follow the procedure described for the sample solution, using the 178
prescribed volume of dilute lead PbTS containing 10 µg of lead per mL instead of the substance 179
to be examined and drying in an oven at 100 °C to105 °C. To 10 mL of the solution obtained, 180
add 2 mL of the sample solution. 181
182
For the monitor solution, follow the procedure described for the sample solution, adding to the 183
substance to be examined the volume of dilute lead PbTS prescribed for the preparation of the 184
reference solution and drying in an oven at 100 °C to105 °C. To 10 mL of the solution obtained 185
add 2 mL of the sample solution. 186
187
For the blank solution, add 2 mL of the sample solution to 10 mL of water R. 188
189
Procedure 5. For the sample solution, unless otherwise specified in the monograph, place the 190
prescribed amount of the substance to be examined (not more than 0.5 g) in a suitable, clean 191
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beaker. Add successively 2.7 mL of cadmium-free and lead-free sulfuric acid (~1760 g/L) TS, 192
3.3 mL of cadmium-free and lead-free nitric acid (~1000 g/L) TS, and 2.0 mL of hydrogen 193
peroxide (~330 g/L) TS using a magnetic stirrer. Allow the substance to react with the reagent 194
before adding the next one. Transfer the mixture to a dry high-pressure digestion vessels 195
(fluoropolymer or quartz glass). 196
197
For the reference solution, follow the procedure described for the sample solution, using the 198
prescribed volume of dilute lead PbTS containing 10 µg of lead per mL instead of the substance 199
to be examined. 200
201
For the monitor solution, follow the procedure described for the sample solution, adding to the 202
substance to be examined the volume of dilute lead PbTS prescribed for the preparation of the 203
reference solution. 204
205
For the blank solution, prepare the solution as described for the sample solution, omitting the 206
substance to be examined. 207
208
CAUTION. When using high-pressure digestion vessels, the safety precautions and operating 209
instructions given by the manufacturer must be followed. The digestion cycles have to be 210
elaborated depending on the type of microwave oven to be used (for example, energy-controlled 211
microwave ovens, temperature-controlled microwave ovens or high-pressure ovens). The cycle 212
must conform to the manufacturer′s instructions. The digestion cycle is suitable if a clear 213
solution is obtained. 214
215
Close the vessels and place them in a laboratory microwave oven. Digest using a sequence of 216
two separate suitable programmes. Design the programmes in several steps in order to control 217
the reaction, monitoring pressure, temperature or energy depending on the type of microwave 218
oven available. After the first programme, allow the digestion vessels to cool before opening. 219
220
Add to each vessel 2.0 mL of hydrogen peroxide (~330 g/L) TS and digest using the second 221
programme. After the second programme, allow the digestion vessels to cool before opening. If 222
Working document QAS/18.769
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necessary to obtain a clear solution, repeat the addition of hydrogen peroxide (~330 g/L) TS and 223
the second digestion programme. 224
225
Cool, dilute cautiously with water R and rinse into a flask, ensuring that the total volume does 226
not exceed 25 mL. 227
228
Colour development and measurement 229
230
For Procedures 1 to 4 231
232
Method A. Use matched flat-bottomed comparison tubes of transparent glass with a uniform 233
internal diameter of 16 mm for the comparison of the colours. “Matched tubes" means tubes that 234
are matched as closely as possible in internal diameter and in all other respects. 235
236
Transfer 12 ml of each of the test solutions prepared as described under Preparations of the test 237
solutions to comparison tubes, add 2 mL of acetate buffer, pH 3.5, TS and mix. Add 1.2 mL of 238
freshly prepared thioacetamide reagent TS, mix and allow to stand for two minutes. 239
240
Compare the colours of the solutions by viewing down the vertical axis of the tube in diffused 241
light against a white or, if necessary, a black background, or by another suitable method. The 242
test is not valid unless the colour of the reference solution is more intense than the colour of the 243
blank solution. If the use of a monitor solution is prescribed, the colour of the monitor solution 244
is at least as intense as the colour of the reference solution. 245
246
The sample complies with the requirements of the test when the colour of the test solution is not 247
darker than the reference solution. 248
249
Method B. Transfer 12 ml of each of the test solutions prepared as described under Preparations 250
of the test solutions to a beaker, add 2 mL of acetate buffer, pH 3.5, TS and mix. Add 1.2 mL of 251
freshly prepared thioacetamide reagent TS, mix and allow to stand for two minutes. 252
253
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Filter the solutions through a suitable membrane filter (nominal pore size 0.45 µm). Carry out 254
the filtration slowly and uniformly, applying moderate and constant pressure to the piston. 255
256
Compare the intensity of the coloration of the residues obtained with the different test solutions 257
on the membrane filters. The test is not valid unless the coloured residue obtained with the 258
reference solution is more intense than the coloured residue obtained with the blank solution. If 259
the use of a monitor solution is prescribed, the coloured residue obtained with the monitor 260
solution is at least as intense as the coloured residue obtained with the reference solution. 261
262
The sample complies with the requirements of the test when the coloured residue obtained from 263
the test solution is not more intense than the coloured residue from the lead standard. 264
265
For Procedure 5: 266
267
Method C. Using short-range pH indicator paper, adjust the test solutions to pH 3.0-4.0 with 268
ammonia (~260 g/L) TS. (Ammonia (~100 g/L) TS may be used as the specified range is 269
approached). To avoid heating of the solutions, use an ice bath and a magnetic stirrer. Dilute to 270
40 mL with water R and mix. Add 2 mL of acetate buffer, pH 3.5, TS and mix. Add to 1.2 mL 271
of thioacetamide reagent R. Mix immediately. Dilute to 50 mL with water R, mix and allow to 272
stand for two minutes. Filter the solutions through a suitable membrane filter (nominal pore size 273
0.45 µm). Carry out the filtration slowly and uniformly, applying moderate and constant 274
pressure to the piston. 275
276
Compare the spots on the filters obtained with the different solutions. The test is not valid unless 277
the coloured residue obtained with the reference solution is more intense than the coloured 278
residue obtained with the blank solution. The coloured residue obtained with the monitor 279
solution is at least as intense as the coloured residue obtained with the reference solution. 280
281
The sample complies with the requirements of the test when the coloured residue obtained with 282
the sample solution is not more intense than the coloured residue obtained with the reference 283
solution. 284
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285
2.2.3 Limit test for heavy metals 286
287
The limit test for heavy metals is provided to demonstrate that the content of metallic impurities 288
that are coloured by hydrogen sulfide does not exceed the heavy metals limit given in the 289
individual monograph in terms of micrograms of lead per gram of the test substance. 290
291
The test consists of two consecutive operations: preparation of the test solution, and the colour 292
development by reaction with hydrogen sulfide, followed by comparison of the colour obtained 293
with that produced with standard lead solution. 294
295
The preparation of the test solution is carried out, as specified in the monograph, according to 296
procedures 1 to 4 described below. A blank is prepared in a similar manner. 297
298
The reaction with hydrogen sulfide is carried out by mixing the test solution with freshly 299
prepared hydrogen sulfide TS. The comparison of the colour thus obtained is carried out either 300
by directly comparing the coloration of the liquid in suitable comparison tubes (Method A) or by 301
comparing the intensity of coloration of spots obtained by filtering the liquid using an 302
appropriate apparatus (Method B). 303
304
Method A is generally applicable only when the amount of heavy metals in the weight of test 305
substance used exceeds 5 μg; for amounts of 2-5 μg of heavy metals Method B should be used. 306
307
The standard lead solution used in the test; dilute lead PbTS contains 10 μg of lead in 1 mL. 308
When 0.1 mL of this solution is employed to prepare the standard for comparison with a solution 309
of 1 g of the substance being tested, the standard solution thus prepared contains 1 μg of Pb and 310
represents the equivalent of 1 μg of lead per g of the substance tested. 311
312
Apparatus 313
314
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For determination of heavy metals by Method A carry out the test in matched flat-bottomed 315
comparison tubes of transparent glass of about 70 mL capacity and about 23 mm internal 316
diameter bearing a 40-mL and a 50-mL mark. Nessler cylinders complying with the above 317
dimensions are suitable. The expression "matched tubes" means tubes that are matched as closely 318
as possible in internal diameter and in all other respects. For mixing the solution use a stirring 319
rod preferably having a loop at the lower end. 320
321
For determination of heavy metals by Method B use a 50-mL syringe made of suitable material 322
(usually plastic) with a detachable plunger and a male Luer conical joint of 9 mm internal 323
diameter at the lower end (Millipore syringe XX 11 050 05 is suitable) to which an adaptor for 324
filtration is attached. 325
326
The adaptor is made of suitable material (a filtration adapter Millipore SX00 013 00 in 327
polypropylene is suitable) and has a female joint for connecting it with the syringe. It is devised 328
so as to be separable into two parts to permit the exchange of filters, the lower part containing a 329
support for membrane filters 13 mm in diameter. A suitable prefilter (Millipore prefilter AP 2001 330
300 is suitable) and a membrane filter made of mixed cellulose esters, 13 mm in diameter, with a 331
pore opening of 3 μm (a Millipore filter SSWP 013 00 is suitable) are used for the filtration. 332
333
Recommended procedure 334
335
Preparation of test solution 336
337
Procedure 1. Weigh the quantity of substance specified in the monograph, dissolve it in 25 mL of 338
water, adjust the pH of the solution to 3-4 with acetic acid (~60 g/l) PbTS, or with ammonia 339
(~100 g/l) PbTS, as necessary, then dilute to 40 mL with water and mix. 340
341
Procedure 2. Weigh the quantity of substance specified in the monograph, dissolve it in about 30 342
mL of solvent specified (ethanol (~750 g/l) TS, methanol R, acetone R, or dioxan R may be 343
used), add 0.5 mL of acetic acid (~300 g/l) TS, and dilute to 40 mL with the solvent. 344
345
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Procedure 3. Place the quantity of the substance specified in the monograph in a suitable crucible, 346
preferably made of silica, and carefully ignite at a low temperature until the contents are 347
thoroughly charred. The crucible may be loosely covered with a lid during the charring. Add to 348
the contents of the crucible 2 mL of nitric acid (~1000 g/l) TS and 5 drops of sulfuric acid 349
(~1760 g/l TS), and cautiously heat until white fumes are evolved, and then ignite, preferably in 350
a muffle furnace, at 500°C until all the carbon is burned off. Cool, add 2 mL of hydrochloric acid 351
(~250 g/l) TS, and slowly evaporate in a water-bath to dryness. Moisten the residue with 1 drop 352
of hydrochloric acid (~250 g/l) TS, add 10 mL of hot water, and digest for 2 minutes. Add, drop 353
by drop, ammonia (~100 g/l) PbTS, until the pH of the solution is between 8 and 8.5, then add, 354
drop by drop, acetic acid (~60 g/l) PbTS, to adjust the pH to between 3 and 4. Filter if necessary, 355
wash the crucible and the filter with about 10 mL of water, dilute with water to 40 mL, and mix. 356
357
Procedure 4. Place the quantity of substance specified in the monograph in a suitable crucible, 358
preferably made of silica, mix it well with about 0.5 g of magnesium oxide R and incinerate until 359
a homogeneous white mass is obtained. If after 15 minutes of incineration the residue is still 360
coloured, let the crucible cool, mix the contents well with a glass rod and resume heating. Next, 361
dissolve the residue in hydrochloric acid (~70 g/l) TS, add, drop by drop, a solution of ammonia 362
(~100 g/l) PbTS, until the pH of the solution is between 8 and 8.5, then add, also drop by drop, 363
acetic acid (~60 g/l) PbTS, to adjust the pH to 3-4, filter, dilute with water to 40 mL, and mix. 364
365
Colour development and measurement 366
367
Method A 368
369
To 40 mL of the liquid contained in the comparison tube add 10 mL of freshly prepared 370
hydrogen sulfide TS, mix and allow to stand for 5 minutes. 371
372
In another comparison tube place a volume of solution of dilute lead PbTS, containing the lead 373
equivalent of the heavy metals limit specified in the monograph, dilute with water, adjust the pH 374
with ammonia (~100 g/l) PbTS and acetic acid (~60 g/l) PbTS to 3-4; dilute with water or the 375
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solvent used to 40 mL, mix, add 10 mL of freshly prepared hydrogen sulfide TS, mix and allow 376
to stand for 5 minutes. 377
378
Compare the colours by viewing down the vertical axis of the tube in diffused light against a 379
white background, or by another suitable method. The colour of the test solution is not darker 380
than that of the lead standard. 381
382
Method B 383
384
Take the filtration syringe, arrange the prefilter and the membrane filter as indicated in the 385
diagram for prefiltration, remove the plunger from the syringe, place the test solution inside the 386
syringe, replace the plunger, and filter the test solution slowly by exerting a regular pressure on 387
the plunger. Collect the filtrate in a beaker or a test-tube. Open the adapter and check whether the 388
membrane filter is free from impurities. If not, replace it and repeat the operation in the same 389
manner. Then rearrange the prefilter and membrane filter as indicated in Fig. 4. Adjust the pH of 390
the filtrate with ammonia (~100 g/l) PbTS and acetic acid (~60 g/l) PbTS to 3-4, add 10 mL of 391
freshly prepared hydrogen sulfide TS, all the reagents previously filtered through a membrane 392
filter, mix, allow to stand for 5 minutes, take out the plunger, place the solution inside the 393
syringe, and filter it through the membrane filter by exerting slowly a regular and moderate 394
pressure on the plunger. Open the adaptor and take out the membrane filter. 395
396
FIG. 4. LIMIT TEST FOR HEAVY METALS: METHOD B 397
398
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399
400
Take a volume of solution of dilute lead PbTS containing the lead equivalent to the heavy metals 401
limit specified in the monograph, dilute with water, adjust the pH with ammonia (~100 g/l) PbTS 402
and acetic acid (~60 g/l) PbTS to 3-4, dilute with water to 40 mL, mix, and proceed as described 403
above. 404
405
Compare the intensity of the coloration of spots obtained on the membrane filters. The colour 406
obtained from the test solution is not more intense than that from the lead standard. 407
408
REAGENTS TO BE ADDED OR REVISED 409
410
Acetate buffer, pH 3.5, TS 411
Procedure. Dissolve 25.0 g of ammonium acetate R in 25 mL of water R and add 38.0 mL of 412
hydrochloric acid (~250 g/l) TS. Adjust the pH, if necessary, with hydrochloric acid (~70 g/L) 413
TS or ammonia (~100 g/L) TS. Dilute with water R to 100.0 mL. 414
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Magnesium oxide R1 415
Complies with the requirements prescribed for magnesium oxide R with the following 416
modifications: 417
Arsenic: maximum 2 ppm. 418
Heavy metals (2.2.3): maximum 10 ppm. 419
Iron: maximum 50 ppm. 420
421
Phenolphthalein/ethanol TS, diluted 422
Procedure. Dissolve 0.1 g of phenolphthalein R in sufficient ethanol (~750 g/L) TS to produce 423
100 mL. 424
Sensitivity test. To 0.1 mL of the phenolphthalein solution add 100 mL of carbon dioxide-free 425
water R. The solution is colourless. Not more than 0.2 mL of 0.02 M sodium hydroxide is 426
required to change the colour to pink. 427
Colour change: pH 8.2 (colourless) to pH 10.0 (red). 428
429
Thioacetamide R 430
C2H5NS = 75.13 (62-55-5). 431
General reagent grade of commerce. 432
White crystals or crystalline powder; melting point, about 113 °C. 433
434
Thioacetamide reagent TS 435
Add 1 mL of a mixture of 15 mL of 1m sodium hydroxide, 5 mL of water and 20 mL of glycerol 436
(85%) to 0.2 mL of thioacetamide solution, heat in a water bath for 20 seconds, cool and use 437
immediately. 438
439
Thioacetamide solution TS 440
A 4% w/v solution of thioacetamide R. 441
442
*** 443