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Home , Prussian blue, Thallium

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1 2 3 4 IPCS EVALUATION OF FOR POISONING BY 5 AND 6 7 8 9 PRUSSIAN 10

11 ferric (III) hexacyanoferrate (II) (colloidal soluble , 12 KFe[Fe(CN)6) and ferric (III) hexacyanoferrate (II) (insoluble Prussian Blue, Fe4 13 [Fe(CN)6]3) 14 15 16 17 18 Initial draft (1994):

19 A.N.P. van Heijst (National Poisons Control Centre, Utrecht, The Netherlands), A. von Dijk 20 (Apotheek, Academisch Ziekenhuis, Utrecht, The Netherlands) & J. Ruprecht (Heyl 21 Pharmaceuticals, Berlin, ) 22 23 Update (2009): 24 M McParland and P Dargan (Medical Toxicology Information Services, Guy's & St Thomas' 25 NHS Foundation Trust, London, UK) 26 27 28 29 30 31

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32 33 1. Introduction ...... 3 34 2. Name and of ...... 3 35 3. Physico-chemical Properties...... 4 36 4. Synthesis and Pharmaceutical Formulation ...... 5 37 5. Analytical Methods...... 7 38 6. Shelf-life...... 9 39 7. General Properties...... 9 40 8. In Vitro Studies ...... 11 41 9. Animal Studies...... 12 42 10. Volunteer Studies...... 21 43 11. Clinical Studies – Clinical Trials ...... 23 44 12. Clinical Studies - Case Reports...... 23 45 13. Summary of Evaluation...... 36 46 14. Model Information Sheet...... 40 47 15. References ...... 42 48 16. Author(s) Name, Address...... 51 49 17. Additional Information...... 51 th 50 Results of Cochrane Library Search 4 November 2009 ...... 52 51 Table 3: Summary of evidence of use of Prussian blue in poisoning ...... 1 52 Table 4: Summarized case reports of poisoning...... 6

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53 1. Introduction 54 55 Prussian blue was first prepared in 1704 and used principally as an inorganic 56 (Faustino, 2008). Work in the 1960s investigated the use of Prussian blue as an antidote for 57 thallium poisoning and a decorporation agent for radiocaesium (-134 and caesium- 58 137). From the 1970s Prussian blue was recommended as an antidote in thallium 59 intoxication and it is now routinely used for this purpose. Further studies into its use for 60 increasing the elimination rate of radiocaesium were undertaken following the Chernobyl 61 accident in 1986 but it was only after Goiânia incident in 1987 that it was 62 used in the management of a large scale radiation accident (Faustino, 2008). 63 64 There are two forms of Prussian blue, ferric (III) hexacyanoferrate (II) (insoluble Prussian 65 blue) and potassium ferric (III) hexacyanoferrate (II) (soluble or colloidal Prussian blue). It 66 is essential to be specific about which form of Prussian blue has been used in future 67 publications. The different forms should be identified by citing the correct chemical name 68 and/or chemical formula and/or the physical nature (insoluble or colloidal soluble). 69 70 Prussian blue is given orally and acts via exchange, adsorption and mechanical trapping 71 to bind thallium and caesium within its lattice, interrupting their enterohepatic 72 circulation, enhancing faecal elimination and reducing body burden. 73 74 Insoluble Prussian blue is used to treat patients with known or suspected internal 75 contamination with radioactive caesium, radioactive thallium and non-radioactive thallium, 76 to increase their rates of elimination. Prussian blue can reduce the biological half-life of 77 radiocaesium by about a third. There is one report where Prussian blue has been used for the 78 treatment of non radioactive caesium (Thurgur et al., 2006) and has also 79 been shown to bind to Prussian blue (Hoffman, 2006). 80 81 Prussian blue is well tolerated but can result in constipation. It is essential to treat 82 constipation as it will decrease elimination of thallium and caesium and some authors advise 83 administration of Prussian Blue with laxatives to prevent constipation. 84 85

86 2. Name and Chemical Formula of Antidote 87 88 There are two forms of Prussian blue, ferric (III) hexacyanoferrate (II) (insoluble Prussian 89 blue) and potassium ferric (III) hexacyanoferrate (II) (soluble or colloidal Prussian blue). 90 Synonyms which apply to both compounds include Berlin blue, Milori Blue, Chinese blue, 91 Hamburg blue, blue, Pigment blue 27 and Paris blue. 92 Potassium ferric (III) Ferric (III) hexacyanoferrate (II) hexacyanoferrate (II) Synonyms colloidal soluble Prussian blue, insoluble Prussian blue, IPB, ferric SPB, potassium ferric , (III) ferrocyanide, ferrocyanide, Ferrotsin, Ferrocin, Ferrihexacyanoferrate, ferric cyanoferrate (II) Molecular formula KFe[Fe(CN)6] Fe4[Fe(CN)6]3 Molecular weight 306.9 859.3

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Potassium ferric (III) Ferric (III) hexacyanoferrate (II) hexacyanoferrate (II) CAS Registry No 12240-15-2 14038-43-8 Colour Index No. 77520 77510 RTECS number LJ8200000 Structural diagram (ChemIDplus Lite)

93 94 95 96 97 Prussian blue is a non-stoichiometric compound and the chemical formulae for both forms 98 shown above are therefore idealized. Additionally all precipitates of insoluble Prussian blue 99 contain nonstoichiometric amounts of potassium, and water (Dvořák, 1971; Nielsen 100 et al., 1987; Nielsen et al, 1988b). The water is partially adsorbed at the large surface of this 101 pigment, partially distributed in the cavities of the crystal lattice (zeolithical water) and 102 partially coordinatively bound (Ludi, 1988). Colloidal soluble Prussian blue may + 103 additionally contain nonstoichiometric amounts of cations (H , ) and anions - 104 (Cl , OH) and water (Bozorgzadeh, 1971; Dvořák, 1970; Dvořák, 1971). 105 106 Insoluble Prussian blue is the only commercially available pharmaceutical preparation 107 however in the literature there is confusion over the two available forms of Prussian blue and 108 inconsistencies in the nomenclature of Prussian blue (Hoffman, 2006). For example, some ® 109 authors who used the commercially available Prussian blue Antidotum Thallii-Heyl 110 containing insoluble ferric hexacyanoferrate have incorrectly described the chemical formula 111 as KFe[Fe(CN)6] (Franke et al., 1979) or named the compound as potassium ferric 112 ferrocyanide (Spoerke et al., 1986). In another report both formulae are used KFe[Fe(CN)6] ® 113 and Fe4[Fe(CN)6]3 for Antidotum Thallii-Heyl (Trenkwalder et al., 1984). In other cases ® 114 the drug Antidotum Thallii-Heyl is described as "colloidal soluble Prussian blue" (Jax et 115 al., 1973; Kemper, 1979; Gansser, 1982; Forth, 1986) or potassium ferric (III) 116 hexacyanoferrate (II) is described as insoluble (Lehmann & Favari, 1984). As a result it is 117 often very difficult or impossible to determine which form of Prussian blue was used. 118 119 Where possible, the two forms of Prussian blue are identified here as soluble Prussian blue 120 and insoluble Prussian blue. 121

122 3. Physico-chemical Properties 123

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124 : Not available. 125 126 : Potassium ferric (III) hexacyanoferrate (II) is colloidally soluble in water, ferric 127 (III) hexacyanoferrate (II) is insoluble in water and diluted acids (solubility product -40 59 128 =10 , i.e. practically insoluble) (Ludi, 1988). Fe-labelled measurements resulted 129 in a solubility of 1.1 µmol/L for potassium ferric (III) hexacyanoferrate (II) and 0.7 130 µmol/L for ferric (III) hexacyanoferrate (II) (Dvořák, 1970). 131 132 Optical properties: Not available. 133 134 pH: Not available. 135 136 pKa: Not available. 137 138 Stability in : Prussian blue should be stored in tightly closed containers protected from 139 light. 140 141 Thermal stability: Not available. 142 143 : Not available. 144 145 Specific gravity: 1.8 146 147 Loss of weight on drying: Potassium ferric (III) hexacyanoferrate (II) 6-15% loss of weight 148 on drying at 150°C. Ferric (III) hexacyanoferrate (II) 28-34% loss of weight on 149 drying at 105°C. 150 151 Excipients and pharmaceutical aids: In both pharmaceutical preparations, Antidotum Thallii- ® ® 152 Heyl and Radiogardase -Cs, ferric (III) hexacyanoferrate (II) is provided as 500mg 153 of Prussian blue powder in gelatine capsules with 0-38mg of microcrystalline 154 cellulose. The powder may vary in coarseness and colour shade (Heyl, 2004). 155 156 Pharmaceutical incompatibilities: None known. 157 158 Binding to some therapeutic drugs and essential nutrients is possible. Insoluble Prussian blue 159 may bind electrolytes in the . Hypokalaemia (potassium 2.5-2.9) 160 was reported in 3 of 42 patients (7%) treated with insoluble Prussian blue (Heyl, 161 2004; Thompson & Callen, 2004; Thompson & Church, 2001). There are some 162 anecdotal reports of Prussian blue decreasing the bioavailability of oral tetracycline 163 and the serum levels and, or clinical response to critical orally administered products 164 should be monitored (Heyl, 2004). 165

166 4. Synthesis and Pharmaceutical Formulation 167 168 4.1 Routes of Synthesis 169 Both substances can be synthesized according to the method of Dvořák (1969). The basic 170 chemicals needed are the same for both substances. However, the salt ultimately formed will 171 depend on the ratio of the raw materials and the way in which they are added to each other. 172

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173 FeCl3 + K4[Fe(CN)6] ------> KFe[Fe(CN)6] + 3 KCl 174 175 4 FeCl3 + 3 K4[Fe(CN)6]----- > Fe4[Fe(CN)6]3 + 12 KC1 176 177 Raw materials: 178 Ferric chloride (FeCl3) and (K4[Fe(CN)6]) 179 180 Water suitable for injection or of comparable quality (i.e. water with cation and anion 181 concentrations equal or less than water for injection). 182 183 Synthesis: 184 (a) Basic solutions 185 186 Dissolve appropriate amounts of FeCl3 and K4[Fe(CN)6] in water so that the molar ratio of 187 the two solutions is 2.0. This means that when a 0.2 M ferric chloride solution has been 188 prepared, the molarity of the K4[Fe(CN)6] solution should be 0.1 M. 189 190 (b1) Potassium ferric (III) hexacyanoferrate (II) 191 3+ 4- 192 It is essential that the final stoichiometric ratio Fe / [Fe(CN)6 ] in the reaction 193 mixture equals 0.33. When 1 litre of 0.2 M ferric chloride solution is used, then 4- 194 (1000 x 0.2)/(0.33 x 0.1) 6060 ml of a 0.1 M (Fe(CN)6] solution is necessary. The 4- 195 quantity of [Fe (CN)6] is added drop wise to the ferric chloride solution whilst 196 stirring vigorously. Centrifuge at 120000g after one hour standing. Wash the 197 precipitate three times with water and centrifuge after each washing step. Dry the o 198 precipitate obtained after the last washing step at 105 C until the weight remains 199 constant. 200 201 To remove possible low molecular impurities some authors have dialysed the 202 colloidal soluble Prussian blue exhaustively against water (Müller et al., 1974; 203 Nielsen et al., 1987; Nielsen et al., 1988a; Nielsen et al., 1988b). 204 205 (b2) Ferric (III) hexacyanoferrate (II) 206 3+ 4- 207 It is essential that the final stoichiometric ratio Fe / [Fe(CN)6] in the reaction 208 mixture equals 1.33. When 1 litre of 0.2 M ferric chloride solution is used, then 4- 209 (1000 x 0.2)/(1.33 x 0.1) 1504 ml of a 0.1 M [Fe(CN)6] solution is necessary. The 4- 210 quantity of [Fe(CN)6] is added drop wise to the ferric chloride solution whilst 211 stirring vigorously. Filter standing for one hour. Wash the precipitate and filter 212 three times after each washing step. Dry the precipitate obtained after the last o 213 washing step at 105 C until weight remains constant. The drying process influences 214 the efficacy of the substance (Nigrović et al., 1966). The effectiveness of Radiogar- ® 215 dase -Cs in binding caesium was higher than that of the ferric (III) hexacyanoferrate 216 (II) synthesized by the authors (Nielsen et al., 1987). Among other things this can be 217 attributed to a special drying procedure. 218 219 After synthesis as described, and after one year's storage in tightly closed containers 220 at room temperature, neither substance showed any significant decomposition into 221 ion (Dvořák, 1970). The effectiveness of ferric (III) hexacyanoferrate (II) 222 was not decreased after storage during one year (Nigrović et al., 1966).

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223 224 4.2 Manufacturing processes 225 Not known. 226 227 4.3 Presentation and formulation ® ® 228 Prussian blue is available as Radiogardase -Cs (Radiogardase in the of ® 229 America) and Antidotum Thallii-Heyl distributed by Heyl Chemisch-pharmazeutische 230 Fabrik GmbH, Berlin, Germany. 231 232 Both products are available in bottles of 30 hard gelatine capsules, each containing 0.5 g of 233 insoluble Prussian blue. 234

235 5. Analytical Methods 236 237 5.1 Quality control procedures for the antidote and/or its formulation 238 239 5.1.1 Test for the presence of colloidal Prussian blue: 240 A mixture of 10 g insoluble Prussian blue and 50 ml of distilled water is shaken and filtered 241 using a membrane filter. The filtrate should show no blue colouration (absence of colloidal 242 soluble Prussian blue). 243 244 5.1.2 Test for the presence of insoluble Prussian blue: 245 Shake 100 mg of colloidal soluble Prussian blue with 20 ml of water. An intensely dark blue 246 solution is obtained. Precipitation should not occur before a standing time of two days has 247 passed (test to exclude the insoluble form). 248 249 5.1.3 Test for the presence of hexacyanoferrate (II) and hexacyanoferrate (III): 250 10 g Prussian blue and 50 ml distilled water are shaken and filtered using a membrane filter. 251 The filtrate is then mixed with the salt of ferrous ammonium sulphate 252 ((NH4)2Fe (II)(SO4)2) or ferric ammonium sulphate ((NH4)Fe (III)(SO4)2). The absence of 253 blue colour indicates that K3[Fe (III)(CN)6] is less than 5 ppm and K4[Fe (II)(CN)6] is less 254 than 20 ppm respectively. 255 256 5.1.4 Test for iron content: 257 500 mg Prussian blue in 50 ml water are shaken and filtered. Compared with an iron 258 standard solution the iron content should be not more than 100 ppm. 259 260 5.1.5 Test for heavy metal content: 261 After digestion of Prussian blue with concentrated sulphuric acid the heavy metal content 262 (measured as ) should be less than 50 ppm. content must be less than 5 µg/g. 263 264 5.1.6 Removal of oxalic acid: 265 Prussian blue may contain oxalic acid, originating from the manufacturing process. Shake 1 266 g of Prussian blue with 10 ml sodium acetate solution (Dutch Pharmacopoeia, 1966). 267 Centrifuge and filter the upper layer through a paper filter folded three times. Add to 5 ml 268 filtrate 1 ml of calcium acetate solution (Dutch Pharmacopoeia, 1966). No precipitate of 269 calcium oxalate should occur after 24 hours. Since no oxalic acid is used in the above speci- 270 fied synthesis, quality control for this acid may be omitted provided that the raw materials do 271 not contain it. The commercially available pharmaceuticals Prussian blue in Antidotum

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® ® 272 Thallii-Heyl and Radiogardase -Cs do not contain oxalic acid. 273 274 5.2 Methods for identification of the antidote 275 Detection of ferric (III) and hexacyanoferrate (II) ions: 500 mg of Prussian blue is heated in 276 5 ml of 3M potassium resulting in decomposition to , flocculent ferric 277 hydroxide. After deposition the supernatant fluid appears (K4[Fe(CN)6]). After fil- 278 tration, 2 ml of the fluid is acidified with concentrated hydrochloric acid and mixed with a 279 FeCl3 solution and this will result in blue colouration or formation of a blue precipitate. 280 281 5.3 Methods for analysis of the antidote in biological samples 282 Not applicable. 283 284 N.B. It is often stated that Prussian blue is not absorbed to any significant extent however 285 anecdotal reports suggest prolonged therapy results in a blue discolouration of sweat and 286 tears suggesting some absorption. 287 288 5.4 Analysis of the toxic agents in biological samples 289 290 5.4.1 Analysis of thallium in urine, plasma and erythrocytes 291 Thallium can be measured by spectrophotometry, flame atomic absorption 292 spectrophotometry (AAS) and electrothermal atomic absorption spectrophotometry 293 (ETAAS) (Moffat et al, 2004). 294 295 5.4.1.1 Spectrophotometry 296 A spectrophotometric method was described by de Wolff and Lenstra (1964). It involves the 297 determination of thallium at 550nb using rhodamine 'B' or brilliant and can be used on 298 urine and on diluted with water. The limit of quantification is approximately 50µg/L 299 (Moffat et al, 2004). This method is described in the Thallium Poisons Information 300 Monograph (IPCS, 1990). 301 302 Another spectrophotometric method is described by Flanagan et al (1995). In this method a 303 - colour indicates the presence of thallium at concentrations of 1mg/l or more. A 304 calibration graph of absorbance against thallium concentration in standard samples can be 305 used to measure the thallium concentration more precisely. The limit of sensitivity is 0.1 306 mg/L. 307 308 5.4.1.2 Atomic Absorption Spectrophotometry 309 Flame AAS can be used to measure thallium in plasma, blood or urine by extracting it as a 310 pyrrolidine dithiocarbamate complex in an organic solvent (de Groot et al, 1985). The lowest 311 quantifiable concentration is at least 0.2 mg/l and depends on the atomic absorption 312 spectrophotometer used. Linearity is up to 5 mg/l. Higher concentrations can be measured 313 by diluting the sample under investigation with blank sample. This method is described in 314 the Thallium Poisons Information Monograph (IPCS, 1990). 315 316 ETAAS has been used for monitoring occupational exposure and has a limit of detection of 317 less than 1 µg/L (Moffat et al, 2004). 318 319 320 5.4.2 Analysis of caesium in biological samples 321 Caesium-134 and caesium-137 decay by emitting beta particles and these nuclides are

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322 detected using gamma spectrometry. 323 324 Specialist advice is essential for dose assessment following a radiation accident as this assists in 325 determining appropriate management and the expected clinical course. Radioactivity 326 measurements of the wound (if applicable), skin or chest (following inhalation), nasal swabs, 327 urine and faeces are also used to assess dose. In many cases the victim is not wearing a 328 dosimeter (and this only measures external exposure not the internal dose). In addition the 329 standard models for calculating intake from routine occupational exposures may not be 330 applicable and individual-specific models may have to be developed and applied for internal 331 dose calculations (Toohey, 2003). 332 333 A quantitative baseline of the internal contamination of radiocaesium should be obtained by 334 appropriate whole body counting and/or by bioassay, or faeces/urine sample whenever 335 possible to obtain the following type of information to establish an elimination curve: 336 • Estimated internalised radiation contamination of caesium, and 337 • Rate of measured elimination of radiation in the faeces (Heyl, 2004). 338 339

340 6. Shelf-life 341 0 342 The pharmaceutical products of Prussian blue should be stored in the dark at 25 C; 0 343 occasional variations of temperature within the range 15 to 30 C are permitted (Heyl, 2004). ® ® 344 For the pharmaceutical preparations Antidotum Thallii-Heyl and Radiogardase -Cs the 345 shelf-life is given as five years. 346

347 7. General Properties 348 349 Orally administered Prussian blue binds thallium or caesium in the gut and increases the 350 concentration gradient, enhancing elimination by gut dialysis. Thallium (Forth & Henning, 351 1979) and caesium (Nigrović, 1965) also undergo enterohepatic circulation and Prussian 352 blue interrupts their reabsorption from the gastrointestinal tract thereby increasing faecal 353 excretion. Prussian blue therefore reduces the biological half-life of caesium and thallium. 354 355 The mechanism of caesium and thallium adsorption by hexacyanoferrates is believed to 356 involve chemical ion exchange whereby nonstoichiometric and stoichiometric cations of the 357 drug are exchanged by thallium or caesium ions. The affinity of Prussian blue increases as 358 the of the cation increases, so Prussian blue preferentially binds caesium (ionic 359 radius 0.169 nm) and thallium (0.147 nm) over potassium (0.133 nm) and sodium (0.116 360 nm) (Forth, 1983; Nielsen et al., 1987). An influence of Prussian blue on potassium and 361 sodium levels is therefore not expected (Nigrović et al., 1966). Rubidium (ionic radius 362 0.148) has also been shown to bind to Prussian blue (Hoffman, 2006). 363 364 Evidence is provided by a number of experimental studies. After binding of thallium to 365 colloidal soluble and insoluble Prussian blue, the content of potassium and hydrogen ions 40 366 was reduced (Dvořák, 1970). K labelled colloidal Prussian blue showed no radioactivity + 367 after mixing with thallium. The pH value was decreased, indicating release of H ions 368 (Dvořák, 1971). In in vitro experiments insoluble Prussian blue bound more caesium than 369 potassium, and hydrogen and iron ions were released.

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370 371 There may be other mechanisms in addition to ion exchange. Nielsen et al. (1987) found that 372 colloidal soluble Prussian blue released more potassium than it adsorbed caesium, 373 suggesting that an additional, physical adsorption on its large surface, possibly interacting 374 with water, may be involved (Ludi, 1983; Richmond, 1968). Yang et al (2008) found that 375 the binding capacity of insoluble Prussian blue for thallium decreased as the water content of 376 Prussian blue was decreased. Forms of Prussian blue with a smaller crystal size, and 377 therefore a larger surface area, have a higher adsorptive capacity and antidotal efficacy for 378 thallium (Kravzov et al., 1993; Yang et al, 2008). 379 380 Non-dialyzed colloidal soluble Prussian blue initially proved to combine with thallium much 381 more effectively than ferric (III) hexacyanoferrate (II) in vitro as well as in vivo (Dvořák, 382 1969; Dvořák, 1970; Kamerbeek et al., 1971). This greater binding was not only the result 383 of the larger surface area of the colloidal soluble preparations (Dvořák, 1970), but also 384 because of the larger amount of extra-stoichiometric potassium in these preparations 385 (Dvořák, 1971). The sodium salt-derived preparation did not meet these requirements 386 (Rauws et al., 1979), and subtle differences in the dimensions of the crystal lattice might also 387 play a role (Keggin & Miles, 1936; Kravzov et al, 1993). 388 389 Colloidal soluble Prussian blue may, however, present problems. The production of a 390 standard colloidal soluble Prussian blue was found to be difficult resulting in a different 391 toxicity of the different preparations (Dvořák et al., 1971). Long-term treatment of rats with 392 non-dialysed colloidal soluble Prussian blue resulted in actual caesium retention in the body 393 in treated rats compared to control animals (Bozorgzadéh & Catsch, 1972; Müller et al., 394 1974). Dresow et al., (1993) investigated the effect of soluble and insoluble Prussian blue in 395 rats, pigs and humans and concluded that for medical use in man, a separation of the low 396 molecular weight compounds from crude commercial preparations is recommended. 397 398 Following the introduction of colloidal potassium ferric (III) hexacyanoferrate (II) as an 399 antidote for thallium poisoning in 1970 (Kamerbeek, 1971; Kamerbeek et al., 1971), the 400 results of treatment with potassium ferric (III) hexacyanoferrate (II) as well as with ferric 401 (III) hexacyanoferrate (II) in patients with severe thallium poisoning published in the 402 literature have been associated with a favourable outcome, particularly when it is used early. 403 Treatment with Prussian blue in patients with thallotoxicosis can be life-saving but it does 404 not improve all the clinical signs, such as neurological signs or alopecia, particularly in late- 405 presenting patients. 406 407 Thompson & Callen (2004) reviewed the English language literature concerning the efficacy 408 of soluble and insoluble Prussian blue and its use as a therapeutic agent in radiocaesium and 409 thallium poisoning. They noted that most of the evidence describing the efficacy of Prussian 410 blue for radiocaesium poisoning is based on the use of the insoluble form, whilst similar 411 evidence for thallium poisoning involves the use of the soluble form. They concluded that 412 while there is sufficient evidence that the insoluble form of Prussian blue is effective in 413 radiocaesium poisoning, there is a lack of analogous data supporting its use in thallium 414 poisoning. The authors acknowledged that further research is needed to determine the 415 significance of any differences between the two forms of Prussian blue and whether their 416 physicochemical differences have any effect on outcomes in human poisoning is currently 417 unknown. However, the commercial products in current use are formulated with the 418 insoluble form of Prussian blue and most data collated from future use is likely to refer to 419 this form only.

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420

421 8. In Vitro Studies 422 423 8.1 In vitro binding of caesium 424 Various in vitro studies have shown that Prussian blue binds caesium (Bozorgzadéh, 1971; 425 Nielsen et al., 1987; Verzijl et al., 1992; Faustino et al., 2008). 426 427 In vitro binding of caesium to insoluble Prussian blue is affected by pH, exposure time, 428 storage temperature (affecting moisture content) and particle size. The lowest caesium 429 binding was at pH 1.0 and 2.0 and the highest at pH 7.5. Dry storage conditions results in 430 loss of moisture from Prussian blue which causes a negative effect on caesium binding 431 capacity. There is batch to batch variation in particle size and variation in binding capacity. 432 At 1, 4 and 24 hours it was determined that caesium binding increases as particle size 433 decreases. The maximum caesium binding capacity of insoluble Prussian blue was 434 approximately 715 mg/g (Faustino et al., 2008). 435 436 In a study comparing the binding capacity of soluble and insoluble Prussian blue, the ® 437 binding of caesium-137 was greater with insoluble Prussian blue (Radiogardase -Cs) and 438 was pH-dependent for both formulations. The maximum binding capacities for insoluble 439 Prussian blue were 87 mg/kg at pH 1.0, 194 mg/g at pH 6.5, and 238 mg/g at pH 7.5. For 440 soluble Prussian blue the maximum binding capacities were 48 at pH 1.0, 73 mg/g at pH 6.5, 441 and 78 mg/g at pH 7.5 (Verzijl et al., 1992). 442 443 Nielsen et al. (1987) investigated caesium binding by various hexacyanoferrate compounds 444 (iron, , cobalt, nickel, , manganese) at pH 1.2 and 6.8. All the compounds bound 445 caesium; potassium copper hexacyanoferrate (II) and potassium zinc hexacyanoferrate (II) 446 were the most efficient at pH 1.2 and 6.8. These compounds had twice the binding capacity 447 of soluble and insoluble Prussian blue. These authors also demonstrated that whey 448 contaminated with caesium-134 and caesium-137 was almost completely decontaminated by 449 dialysing a whey suspension against a buffer solution containing insoluble Prussian blue. 450 The whey had been produced from the milk of South German cows contaminated in the 451 summer of 1986 following the Chernobyl accident. 452 453 8.2 In vitro binding of thallium 454 In vitro studies have demonstrated that thallium binds to both soluble (Dvořák, 1970; 455 Kamerbeek et al., 1971; Krazov et al., 1993) and insoluble Prussian blue (Dvořák, 1970; 456 Yang et al., 2008). The adsorption is rapid: after 10 minutes all thallium was bound (Dvořák 457 et al., 1971). The effectiveness of soluble Prussian blue (as measured by mg of thallium/mg 458 Prussian blue or % adsorbed) was higher than that of insoluble Prussian blue (Dvořák, 1970; 459 Kamerbeek et al., 1971). In vitro, thallium binds more strongly to Prussian blue than to 460 activated charcoal (Kamerbeek et al., 1971). 461 462 As with caesium, the thallium binding capacity of Prussian blue is affected by pH, exposure 463 time, storage temperature (affecting moisture content) and particle size. The adsorption of 464 thallium on Prussian blue depended on the pH-value of the solution and a maximal 465 adsorption could be detected at a neutral or slightly alkaline pH (Dvořák, 1970). In the in 466 vitro study by Yang et al. (2008) the maximum thallium binding capacity of insoluble 467 Prussian blue was approximately 1400 mg/g at pH 7.5 after 24 hours. The lowest binding 468 occurred at pH 1.0 at 1 hour and thereafter, binding increased as pH increased (determined

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469 up to pH 7.5). The binding constant was higher for fully hydrated Prussian blue compared to 470 Prussian blue which had been dried for 24 hours and the smaller the particle size the higher 471 the binding constant. 472 473 An in vitro study examined the adsorption of thallium-201 by insoluble Prussian blue to 474 investigate the use of Prussian blue in reducing the radiation burden in thallium-201 475 myocardial . The maximum adsorption peaked at approximately 30 minutes 476 irrespective of the concentration (1, 10 or 100 mg or Prussian blue in 5 mL of water). It 477 reached a plateau at 30 to 60 minutes with no further increase in adsorption of thallium until 478 4 hours. The rate of absorption was constant for up to 1 hour but slowed down thereafter. 479 Thallium-201 removal increased from 46 to 95% when the pH was increased from 2.0 to 8.0. 480 In the most favourable conditions the maximal adsorption capacity was 5000 MBq of 481 thallium-201/g of Prussian blue and maximal adsorption occurred at 30 minutes (Bhardwaj 482 et al., 2006). 483 484

485 9. Animal Studies 486 487 9.1 Pharmacodynamics 488 489 9.1.1 Caesium 490 A number of animal studies have examined the impact of Prussian blue on the absorption of 491 caesium, its effect on the enhancement of excretion (or reduced retention) of caesium and 492 the impact of these actions on final outcomes. 493 494 9.1.1.1 Prevention of absorption/uptake from gut 495 Administration of a single dose of radiocaesium and concomitant oral dosing of Prussian 496 blue resulted in reduction of caesium uptake from the gastrointestinal tract (Brenot & 497 Rinaldi, 1967; Dresow et al., 1990, 1993; Giese & Hantzch, 1970; Nielsen et al., 1988b; 498 Nigrović, 1963; Nigrović, 1965). In piglets potassium ferric (III) hexacyanoferrate (II) and 499 ferric (III) hexacyanoferrate (II) reduced the caesium-134 uptake by more than 97%. The 500 diminution of the caesium-134 body burden depended on the dose of administered 501 hexacyanoferrate (II). The difference between the colloidal and insoluble Prussian blue 502 compounds in decreasing enteral absorption of caesium-134 was small (Nielsen et al., 503 1988b). 504 505 If Prussian blue was given as late as 60 minutes after caesium-137 administration the enteral 506 caesium-137 absorption was also suppressed (Nigrović, 1963). Autoradiography of rats 507 showed, that the radioactivity was limited to the gastrointestinal-tract (Brenot & Rinaldi, 508 1967). 509 510 In rats administered a composite treatment mixture containing calcium alginate, potassium 511 iodide and insoluble Prussian blue in their diet and exposed to various radionucleotides, the 512 Prussian blue, even as a component of the mixture, decreased the absorption of caesium into 513 the organism and reduced the whole-body retention (Kargacin & Kostial, 1985; Kostial et 514 al., 1980; Kostial et al., 1981; Kostial et al., 1983). 515 516 In a study in pigs the animals (82 kg) were fed twice daily with pellet food and 500 mL of 517 milk together with 200 g of radiocaesium-contaminated whey powder. Prior to each feeding

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518 the pigs were given 0.5g, 1.5g or 2.5 g of soluble Prussian blue, insoluble Prussian blue or 519 ammonium iron hexacyanoferrate (II) (NH4Fe[Fe(CN)6]) in gelatine capsules. The animals 520 were slaughtered after 27 days of feeding. All compounds were found to be effective at 521 reducing caesium-134 and caesium-137 absorption. Administration of increasing quantities 522 of the compounds resulted in a dose-dependent reduction in the radioactivity of the tissues 523 tested. Soluble Prussian blue and ammonium iron hexacyanoferrate were most effective, 524 possibly due to more favourable distribution of colloidal soluble compounds in the 525 gastrointestinal tract (Dresow et al., 1993). Similarly, soluble Prussian blue and ammonium 526 iron hexacyanoferrate were most effective in rats (260 to 280 g) given 0.5 mg of Prussian 527 blue in water 2 minutes before 0.5 mL of water containing a tracer dose of caesium-134, by 528 gastric tube. There was almost complete blockade of radiocaesium absorption as judged by 529 urinary excretion and whole body retention measured 7 days after ingestion. Insoluble 530 Prussian blue was less effective (Dresow et al., 1993). 531 532 9.1.1.2 Enhancement of decorporation (reduced retention) 533 Chronic feeding of rats (Dresow et al., 1993; Stather, 1972) or piglets (Dresow et al., 1993; 534 Giese et al., 1970) with caesium-137 contaminated food and concomitantly administration of 535 Prussian blue resulted in reduced whole body retention. Most of the daily caesium-137 dose 536 was excreted in the faeces. 537 538 After ingestion of radiocaesium, administration of insoluble Prussian blue (Bozorgzadéh & 539 Catsch, 1972; Dresow et al., 1990; Nigrović, 1963; Nigrović, 1965; Nigrović et al., 1966; 540 Stather, 1972;) and colloidal soluble Prussian blue (Bozorgzadeh, 1971; Bozorgzadéh & 541 Catsch, 1972; Dresow et al., 1990; Giese et al., 1970; Müller et al., 1974) decreased the 542 whole-body-retention of the caesium . Also in a mixture with other substances 543 Prussian blue increased the excretion of the caesium-137 (Kostial et al., 1983). In rats the 544 effect on the whole-body retention was age-dependent. In younger animals the whole body 545 retention was lower than in older animals. Probably because of the higher basal metabolic 546 rate more caesium was excreted into the gut in young animals (Bozorgzadeh, 1971; Stather, 547 1972). 548 549 Prussian blue increased the cumulative excretion of incorporated radiocaesium in faeces and 550 urine (Brenot & Rinaldi, 1967; Nigrović, 1965; Nigrović et al., 1966). Whereas in untreated 551 animals most caesium is excreted in the urine, in animals treated with Prussian blue faecal 552 excretion predominates (Nigrović et al., 1966; Brenot & Rinaldi, 1967; Müller, 1969; Giese 553 et al., 1970; Giese & Hantzsch, 1970; Richmond, 1968). The biological half-life was 554 reduced (Madshus et al., 1966; Nigrović et al., 1966; Havliček et al., 1967; Havliček, 1968; 555 Müller et al., 1974; Richmond, 1968; Strömme, 1968). In rats the half-life was reduced by 556 50% (11 days compared to 6 days) (Nigrović et al., 1966; Müller et al., 1974), and in dogs 557 from 11 to 6.5 days (Madshus et al., 1966). 558 559 Colloidal soluble Prussian blue was more efficient in increasing the excretion of 560 radiocaesium than the insoluble form (Müller, 1969). In long-term use however, colloidal 561 soluble Prussian blue decreased the excretion compared with control animals. Insoluble 562 Prussian blue did not show this effect (Bozorgzadéh & Catsch, 1972). After thorough 563 dialysis with water to remove any possible low molecular impurities this effect of the 564 colloidal Prussian blue disappeared (Müller et al., 1974). The efficacy of the dialyzed 565 colloidal soluble Prussian blue, however, was only marginally better than that of insoluble 566 Prussian blue (biological half-lives: control 10.51 days, insoluble Prussian blue 5.6 days, 567 colloidal Prussian blue 4.88 days) (Müller et al., 1974).

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568 569 The efficacy of therapy with Prussian blue is time-dependent. The best effect resulted in 570 administration of Prussian blue 2 minutes before application of caesium (Dresow et al., 571 1990). Start of treatment immediately after intoxication was also successful (Bozorgzadéh, 572 1971; Bozorgzadéh & Catsch, 1972), but it was still effective after a delayed start of 3.5 573 days (Stather, 1972). 574 575 The reduced whole-body retention of caesium after treatment with Prussian blue can also be 576 seen in individual organs. This reduced caesium content of the different organs has been 577 reported in muscle (Bozorgzadéh, 1971; Bozorgzadéh & Catsch, 1972; Brenot & Rinaldi, 578 1967; Kostial et al., 1983; Müller et al., 1974; Stather, 1972; Wolsieffer et al., 1969), bone 579 (Bozorgzadéh & Catsch, 1972; Müller et al., 1974; Wolsieffer et al. 1969), carcass (Kostial 580 et al., 1983; Wolsieffer et al. 1969), liver (Bozorgzadéh, 1971; Müller et al., 1974; Stather, 581 1972) and kidney (Bozorgzadéh, 1971; Bozorgzadéh & Catsch, 1972; Brenot & Rinaldi, 582 1967; Kostial et al., 1983; Müller et al., 1974; Stather, 1972). Due to a different turnover 583 rate of the organs the rate of diminution of caesium in the different organs varied. In the 584 gastrointestinal-tract the caesium content was increased due to binding on the non-resorbable 585 Prussian blue (Brenot & Rinaldi, 1967). 586 587 After 15 days of dietary acclimatization to insoluble Prussian blue as 1% of their food, rats 588 were given intraperitoneal casesium-137. The animals were killed 30 days later. There was 589 significant reduction in the retention of caesium-137 in the carcass, femur and muscle 590 (Wolsieffer et al., 1969). 591 592 Oral insoluble Prussian blue was given to rats in drinking water (400 mg/kg/day) for 11 days 593 which was started immediately after an intravenous injection with caesium-137. On 594 evaluation at 11 days Prussian blue had increased the faecal excretion of caesium-137 5-fold 595 and this consequently also reduced urine excretion. There was also reduced retention of Cs- 596 137 in all the tissues tested (blood, liver, kidneys, spleen and skeleton) (Le Gall et al., 2006). 597 598 Oral administration of insoluble Prussian blue shortened the retention of caesium-137 in 599 mated, pregnant and lactating rats and the deposition of caesium-137 in the embryos and 600 nursed young animals was reduced (Havliček, 1967; Havliček, 1968). 601 602 The effect of Prussian blue provided in drinking water to rats of various ages, on the 603 excretion of intraperitoneally administered caesium-137 was investigated. In 19-week old 604 rats the body burden of caesium-137 was reduced to 34% of the controls 1 week after 605 injection whilst in 9-week and 4-week old rats, the corresponding values were 28% and 9% 606 respectively. analysis suggested that the rate limiting factor of excretion of caesium- 607 137 during Prussian blue therapy was the turnover rate of caesium-137 in muscle tissue. The 608 turnover rate of caesium-137 in muscle tissues of young animals is faster than that of older 609 animals and this is reflected in an increased efficacy of Prussian blue in removing caesium- 610 137 from the younger animals (Stather, 1972). 611 612 9.1.1.3 Impact on outcomes 613 Richmond & Bunde, (1966) investigated the effect of three different concentrations of 614 Prussian blue on caesium-137 contaminated rats. Each rat (approximately 92 days old with 615 and average bodyweight of 372g) received approximately 0.84 microcurie of caesium-137, 616 the rats were then measured 30 minutes later for total body activity and then returned to their 617 respective cages. Insoluble Prussian blue was incorporated into their drinking water which

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618 they had free access to over the following 60 days. The concentrations of Prussian blue in 619 the water were 0, 0.025, 0.25 and 2.5 g/L, and the estimated daily dose of Prussian blue was 620 0, 0.9, 8.5 and 84 mg/rat, respectively. There was significant reduction in the whole body 621 retention of caesium-137 with continuous ingestion of Prussian blue at 0.25 and 2.5 g/L, but 622 no effect was observed at 0.025 g/L. Retention of caesium was described in 3 exponential 623 terms and the third component had a half-life of 8.77 days in high-dose Prussian blue rats 624 compared to 14 days in control rats. The authors concluded that the rate of caesium-137 625 secretion would depend partially on the turnover rate in body tissues, particularly muscle, 626 which accounts for a large proportion of the total body caesium-137 activity. 627 628 The effect of Prussian blue was studied in sheep fed wheat and grass contaminated with 629 caesium after the Chernobyl accident. The lactating sheep (average 35 kg) were given 0.5 630 kg of wheat (average radiocaesium content 1684 ± 17 Bq/kg) and 200 g (9840 ± 442 Bq/kg) 631 daily. The average milk yield was 110 g per day. When the radiocaesium content of the 632 milk had reached a state of equilibrium half the animals were given colloidal Prussian blue, 633 5 g in 5 L of drinking water for 23 days. The colloidal Prussian blue settled in the container 634 and the drinking water had to be stirred repeatedly during the day and it took about 2 days 635 before the sheep became used to the water. Treatment with Prussian blue reduced the 636 radiocaesium content of the milk by approximately 85% and the effect was seen within a few 637 days of the start of treatment (Ioannides et al., 1991). 638 639 The role of Prussian blue in removing caesium-137 internal contamination from rats was 640 studied to evaluate the possible side effects caused by chronic consumption on various 641 biomarkers for exposure. Rats of two age groups (growing rats (2-months old) and adult rats 642 (4month old)), were observed over a 60 day having had either caesium-137 alone, 643 Prussian blue alone, or a combination of both, administered at different time intervals (e.g. 644 both given simultaneously or with a time lapse between). The authors reported that in both 645 growing and adult rats Prussian blue administration, particularly when given before or 646 immediately after Caesium-137 intake, could eliminate the effects of caesium-137 647 irradiation on red blood cell count and haemoglobin content, and on serum levels of: total 648 proteins (in adult rats only), globulins (in adult rats only), creatinine, urea, urea nitrogen, 649 ALT activity, T3, and T4. When given one or seven days post caesium-137 irradiation, 650 Prussian blue eliminated the effects of caesium-137 treatment on serum cholesterol, serum 651 calcium and serum bilirubin, in both growing and adult rats (Fekry et al., 2003). 652 653 The efficacy of insoluble and colloidal soluble Prussian blue in removing a single dose of 654 internally deposited caesium-134 was compared in rats (6 rats in each experimental ). 655 Prussian blue was administered twice daily and treatment started immediately after injection 656 of caesium-134. For the first few days the colloidal form initially showed greater efficacy in 657 removing the caesium but with continuous administration it brought about a complete 658 blockage of caesium-134 excretion from the liver, spleen and skeleton. This was attributed 4- 659 to in vivo disintegration of the colloidal compound yielding free [Fe(CN)6] which when 660 absorbed from the gut reacts with endogenous metal ions. The authors highlight the practical 661 clinical consequences of this finding in that the insoluble compound should be the antidote 662 of choice for caesium-134 toxicity; the slight transient superiority of the colloidal form is 663 over-shadowed by its untoward long-term effect (Bozorgzadéh & Catsch, 1972). 664 665 9.1.2 Rubidium 666 There is limited information on the decorporation effect of Prussian blue in rubidium 667 exposure. Rats were fed insoluble Prussian blue in their food (as 5%) from 2 days before an

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668 intraperitoneal injection of rubidium-86. Prussian blue reduced the half-life of rubidium-86 669 from 10.3 days to 1.7 days, reducing the whole body retention to 9% that of controls after 7 670 days (Stather, 1972). 671 672 9.1.3 Thallium 673 674 9.1.3.1 Prevention of absorption/uptake from gut 675 Concomitant oral administration of thallium and of Prussian blue in rats resulted in a lower 676 uptake of the metal and lower concentrations found in organs (Dvořák, 1969; Heydlauf, 677 1969; Rauws, 1974). Soluble Prussian blue was more effective than the insoluble Prussian 678 blue (Dvořák, 1969). 679 680 In a study by Heydlauf (1969) aqueous solutions of thallium-204 sulphate were administered 681 to rats by gastric tube. Aqueous suspensions of ferric cyanoferrate (II) (insoluble Prussian 682 blue) in doses of 0.5-50 g were then administered by gastric tube 1 to 60 minutes later. The 683 maximal protective effect, i.e. approximately ten times lower absorption of thallium-204, 684 was observed when Prussian blue was given immediately, although an effect was still seen 685 when Prussian blue was administered at 60 minutes. As expected, there was a marked 686 dependence of antidotal efficacy on the dose of Prussian blue administered. 687 688 9.1.3.2 Enhancement of decorporation (reduced retention) 689 It was also shown that insoluble Prussian blue was able to remove thallium across the 690 gastrointestinal wall. Carrier-free thallium-204 was injected intravenously and the animals 691 fed with Prussian blue pellets at will (Heydlauf, 1969). The thallium-204 content of 692 Prussian blue treated animals was drastically reduced even when treatment was initiated on 693 the fourth day. This was due to markedly enhanced faecal excretion whereas urinary 694 elimination did not reach the control level. 695 696 Insoluble Prussian blue (Heydlauf, 1969) as well as colloidal soluble Prussian blue (Dvořák, 697 1969; Günther, 1971) reduced the retention of thallium in the body. The excretion in the 698 faeces was increased and decreased in the urine when compared to the control animals 699 (Heydlauf, 1969; Lehmann & Favari, 1985; Leloux et al. 1990; Manninen et al., 1976; 700 Rauws, 1974; van der Stock & de Schepper, 1978). The cumulative excretion in faeces and 701 urine was increased (Heydlauf, 1969; Lehmann & Favari, 1985; Rauws, 1974). The bio- 702 logical half-life of thallium in the body was reduced. In dogs the biological half-life was 703 decreased from 6.5 days (measured in control animals) to 2.5 days (animals treated with 704 Prussian blue) (van der Stock & de Schepper, 1978), in rats it was decreased from 4 days to 705 2 days (Rauws, 1974). 706 707 A study in rats demonstrated that soluble Prussian blue increased excretion of thallium and 708 reduced the LD50 but only if started within 24 hours of exposure. Thereafter thallium- 709 induced pathological changes were irreversible (Günther, 1971). 710 711 The reduced retention and the increased excretion of thallium by Prussian blue results in a 712 decrease of the thallium content in liver (Dvořák, 1969; Günther, 1971; Heydlauf, 1969; 713 Kravzov et al., 1993; Manninen et al., 1976; Rìos & Monroy-Noyola, 1992; Sabbioni et al., 714 1982), kidney (Dvořák, 1969; Günther, 1971; Heydlauf, 1969; Kravzov et al., 1993; 715 Manninen et al., 1976; Rauws, 1974; Rìos & Monroy-Noyola, 1992; Sabbioni et al., 1982), 716 skeleton (Heydlauf, 1969) blood (Rìos et al., 1991), heart (Kravzov et al., 1993; Rìos & 717 Monroy-Noyola, 1992) and muscles (Dvořák, 1969; Günther, 1971; Heydlauf, 1969;

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718 Manninen et al, 1976; Rauws, 1974). 719 720 9.1.3.3 Impact on outcomes 721 Kamerbeek et al. (Kamerbeek, 1971; Kamerbeek et al., 1971) showed the influence of 722 Prussian blue on the concentration of thallium in the brain. Thirty-five rats, divided into 723 seven groups of five animals, were given 0.075 mM/kg thallous nitrate in 5% glucose 724 solution by intraperitoneal injection. After 24 hours, one group was sacrificed. Three of the 725 remaining groups were subsequently treated with 50 mg Prussian blue suspended in saline, 726 twice daily by gavage. The other groups served as controls. At 48, 72 and 120 hours after 727 administration of thallium, one control and one treated group were killed. The thallium 728 concentration was determined in the brain and in a muscle specimen (quadriceps). After 729 four days of Prussian blue therapy the concentration of thallium in the brain of the treated 730 groups was less than half that of the control group. The muscle thallium concentration in the 731 treated group was almost one-fourth of that of the control group. A dose-dependent 732 relationship was observed. Also in other experiments reduced thallium levels in the brain 733 were measured (Kravzov et al., 1993; Leloux et al., 1990; Manninen et al., 1976; Rauws. 734 1974; Rìos et al., 1991; Rìos & Monroy-Noyola, 1992; Sabbioni et al., 1982). 735 736 Trying to establish an optimal dosage scheme for use of Prussian blue in human 737 thallotoxicosis, Kamerbeek (1971) gave five groups of five rats 0.1 mm/kg thallous nitrate 738 intraperitoneally. After 24 hours four groups were treated by gavage once daily with 10, 50, 739 250 and 1000 mg/kg Prussian blue, suspended in 15% mannitol. After four days of 740 treatment, the animals were sacrificed and the thallium concentrations were determined in 741 the brain and in a muscle specimen. A daily dose of 250 mg/kg Prussian blue appeared to be 742 as effective as 1000 mg/kg/day with respect to thallium-concentration in muscle specimen 743 but with respect to thallium in the brain the highest dosage was more efficacious. 744 745 Kamerbeek (1971) further investigated the protection afforded by Prussian blue against 746 thallium toxicity. Two groups of rats were given 0.25 mg/kg thallous nitrate intraperi- 747 toneally. Four hours later one group received Prussian blue 100 mg/kg in a 15% mannitol 748 solution by gavage. The other group received mannitol only. This regimen was repeated for 749 10 days. In the control group, 10 of 20 animals died, while in the treated group only two 750 deaths occurred. 751 752 It was further shown that enhanced thallium-204 excretion as a result of Prussian blue 753 therapy was accompanied by reduced thallium toxicity (Kravzov et al., 1993; Rìos et al., 754 1991; Rìos & Monroy-Noyola 1992). Treatment with potassium ferric hexacyanoferrate (II) 755 (colloidal soluble Prussian blue) increased the LD50 by a factor 2.3 (Günther, 1971). After 756 application of 30 mg thallium/kg the survival in the control group was 0% and in the 757 Prussian blue group 50% (Heydlauf, 1969). 758 759 After intraperitoneal injection of thallium (32 mg/kg) on day 1, a group of 16 rats was given 760 soluble Prussian blue (50 mg/kg orally twice daily), D- (intraperitoneal 761 injection 25 mg/kg) or a combination of the two from days 2 to 5. The mortality in the 762 different treatment groups by day 6 was: control group 87.5%, Prussian blue group 56.25%, 763 D-penicillamine group 100% and Prussian blue + D-penicillamine group 25%. Only the 764 combination of antidotes produced a significant difference in mortality compared to controls. 765 Prussian blue alone protected against thallium-induced neurotoxicity (as measured by the 766 number of altered Purkinje cells) but the effect was greater with combined Prussian blue + 767 D-penicillamine. D-penicillamine alone did not protect against thallium-induced changes in

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768 Purkinje cells (Barroso-Moguel et al, 1994). 769 770 9.2 Pharmacokinetics 771 772 9.2.1 Oral 773 There is limited information on the pharmacokinetics of Prussian blue as these compounds 774 are very poorly absorbed from the gastrointestinal tract. Most studies have been performed 775 with iron-59 or carbon-14 labelled Prussian blue examining intestinal absorption and 776 bioavailability of iron and cyanide (see section 9.4 for studies on assessment of cyanide 777 toxicity). 778 779 The release of iron from potassium ferric (III) hexacyanoferrate (II) and ferric (III) 780 hexacyanoferrate (II) was examined in piglets. The compounds were labelled with iron-59 781 in the ferric or ferrous position. When labelled in the ferric position only 1.47% of the iron 782 was absorbed from potassium ferric (III) hexacyanoferrate (II) and 1.34% from ferric (III) 783 hexacyanoferrate (II), as determined by the whole-body retention 14 days after oral dosing. 784 Only 0.2% and 0.15%, respectively, of the iron was absorbed from the ferrous position. 785 Most of the dose was excreted in the faeces; 0.1 to1% of the iron-59 was in the urine but it 786 could not be determine how much of may have been due to faecal contamination (Nielsen et 787 al., 1988a). This study suggests that iron is not significantly absorbed from Prussian blue. 788 59 14 789 Administration of labelled ( Fe, C) Prussian blue to rats resulted whole-body retention of 790 0.03% of the dose (only in the gastrointestinal tract) and in traces of radioactivity in the 791 urine (0.15%). The amount in blood and skeleton was below the detection limit. After 792 administration of iron-59 labelled potassium ferric (III) hexacyanoferrate (II) 59 793 (K Fe[Fe(CN)6]) to rats traces of radioactivity were found in the skeleton (0.11% of the 794 administered dose) and in blood (0.046%). Again, with radio-labelled iron in the ferric and 795 ferrous positions, the differences in distribution showed that Prussian blue is not absorbed, + 3+ 4- 796 but the different ions K , Fe and [Fe(CN)6] are metabolized instead. No evidence was 4- 797 obtained for decomposition of [Fe(CN)6] (Dvořák et al., 1971). Histopathological 798 examination of organs showed no deposits of Prussian blue after oral administration of 799 insoluble and colloidal soluble Prussian blue (Giese & Hantzsch, 1970). 800 801 9.2.2 Parenteral 802 After intraperitoneal administration of radio-labelled colloidal soluble Prussian blue the 803 substance is eliminated by the reticuloendothelial system. On the first day 40.5% of the 804 radioactivity was excreted in the urine, the content in the faeces was very small. On the 805 second day 42% was found in the faeces with only traces in the urine. After 4 days the body 806 retention was 4.5%, mostly in the liver (Müller, 1969). 807 59 59 808 Intravenous administration of KFe[ Fe(CN)6] and K Fe[Fe(CN)6] resulted in entirely 809 different metabolic behaviour in rats between the two forms. With potassium ferric (III) 59 810 hexacyanoferrate (II) labelled in the ferrous position (KFe[ Fe(CN)6]) more than 50% of the 811 radioactivity was excreted in the urine, by contrast when labelled in the ferric position 59 812 (K Fe[Fe(CN)6]) only 0.06%. The faecal excretion was low for both. The distribution of 59 813 the radioactivity into the organs after administration of KFe[ Fe(CN)6] differed from that of 59 59 814 K Fe[Fe(CN)6]. Whereas the radioactivity of KFe[ Fe(CN)6] persisted in the liver for 8 59 815 days, the activity of K Fe[Fe(CN)6] varied from the liver to the blood (Dvořák et al., 1971). 816 817 9.3 Toxicology

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818 819 9.3.1 Acute toxicity 820 Ferric (III) hexacyanoferrate (II) 821 Oral: 822 According to a Soviet study, 8g/kg body weight was not lethal to laboratory animals 823 (presumed to be rats or mice) and produced no clinical signs of toxicity (BIBRA, 1997) 824 825 Intraperitoneal administration: 826 LD50 rat: 1.13 mg/g body weight (Brenot & Rinaldi, 1967). 827 LD50 rat: 2.1g/kg body weight (BIBRA, 1997). 828 LD50 mouse: 2 g/kg body weight (BIBRA, 1997). 829 830 Rats or mice given lethal doses suffered inertia, breathlessness and sluggishness with excess 831 blood in the liver, spleen and kidney (BIBRA, 1997). 832 833 834 Potassium ferric (III) hexacyanoferrate (II) 835 In studies by Dvořák et al. (1971) the lethality of intravenous injection of 1 mg of colloidal 836 soluble Prussian blue in rats varied from 0% to 100%, despite the same manufacturing 837 processes. Some animals became unwell within 15 minutes and developed respiratory 838 distress. A blue colouration was noted in the lungs at post-mortem examination. This 839 variation in toxicity was thought to be due to differences in the degree of dispersion of the 840 Prussian blue in the solution of each batch. 841 842 843 Pigment blue 27 844 Oral LD50 rat: >5g/kg body weight (BIBRA, 1997). 845 846 847 848 9.3.2 Chronic toxicity 849 In rats colloidal soluble Prussian blue given as 2% of drinking water for 12 weeks resulted in 850 no significant body weight changes or histopathological changes in the organs, including the 851 gut (Dvořák et al., 1971). Similarly, sheep (average weight 35 kg) given colloidal soluble 852 Prussian blue, 5 g in 5 L of drinking water daily for 23 days had no change in body weight 853 (Ioannides et al., 1991). 854 855 There were no significant differences in average fluid intake in rats given insoluble Prussian 856 blue in drinking water (0.025, 0.25 or 2.5 g/L) for 60 days. The estimated daily dose of 857 Prussian blue was 0.9, 8.5 and 84 mg/rat, respectively (equating to 2.4, 23, 226 mg/kg) 858 (Richmond & Bunde, 1966). 859 860 Oral insoluble Prussian blue caused no adverse effects and no impairment of growth in 861 young rats when given as 1% of the diet for 120 days (Nigrović et al., 1966) or as 1% of 862 their food for 60 days in rats (Wolsieffer et al., 1969). Also in rats, food consumption and 863 body weight were unchanged during 9 days of treatment with a mixture of sodium alginate 864 (daily consumption 2 g), insoluble Prussian blue (250 mg) and sodium perchlorate (100 mg) 865 (Kostial et al., 1980) or during 4 weeks treatment with a mixture of calcium alginate 866 (average 4.8 g/day), insoluble Prussian blue (average 0.8 g/day) and 867 (0.0048 g/day) (Kostial et al., 1981).

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868 869 There were no adverse effects in dogs (7 to 8 kg) given oral insoluble Prussian blue (3 or 6 870 doses of 0.5 g daily) for 11 days. The doses equate to approximately 200 and 400 mg/kg, 871 respectively (Madshus et al., 1966). At autopsy no pathological changes were observed 872 (Nigrović et al., 1966). 873 874 9.3.3 Reproductive toxicology and teratogenicity 875 There is no information available on the reproductive toxicity of Prussian blue. 876 877 In pregnant rats intoxicated with oral thallium soluble Prussian blue started 8 hours later 878 increased the survival rate, reduced the thallium content of the placenta by 5-fold and in the 879 foetuses reduced the thallium content of the brain and liver (Sabbioni et al., 1982). 880 881 9.3.4 Genotoxicity 882 No information available. 883 884 9.4 Assessment of possible cyanide toxicity 885 Prussian blue contains cyanide ions bound to iron. At extremely low pH values in the 886 presence of oxidizing agents Prussian blue decomposes and, under these circumstances, 887 cyanide can be released. Since oral administration of Prussian blue is indicated in the 888 treatment of thallium poisoning and caesium incorporation, various studies have examined 889 the possibility of cyanide release from these hexacyanoferrate compounds. 890 891 When gastric juice (pH 2) and soluble Prussian blue were incubated for 4 hours no cyanide 892 was detected. Similarly no cyanide was detected when the study was conducted with 0.1 N 893 hydrochloric acid at room temperature. Cyanide was only detected when this last 894 experiment was repeated at 100 °C (Kamerbeek, 1971). Other in vitro studies have also 895 shown that the release of cyanide is negligible (Dvořák, 1970). 896 897 Verzijl et al. (1993) studied in vitro cyanide release of four Prussian blue salts, potassium 898 ferric (III) hexacyanoferrate (II), ferric (III) hexacyanoferrate (II) and ammonium ferric (III) 899 hexacyanoferrate (II), both unpurified and purified compounds (that is, with and without 900 33% ammonium chloride as a manufacturing impurity). These salts were added to water, 901 artificial gastric (pH 1.2) or intestinal (pH 6.8) juices and the content flasks were allowed to 0 902 stand for 5 hours, protected from light, at 37 C. Cyanide was detected in all tests and the 903 quantity released ranged from 22 to 535 µg/g of Prussian blue in water, 64 to 418 µg/g in 904 artificial gastric juice and 15 to 58 µg/g in artificial intestinal juice. For all salts tested the 905 release of cyanide was greatest in artificial gastric juice than the other test media. The 906 unpurified ammonium ferric (III) hexacyanoferrate (II) released the most cyanide and ferric 907 (III) hexacyanoferrate (II) (insoluble Prussian blue) the least in all test media. 908 909 In an in vitro study the release of cyanide from insoluble Prussian blue was measured over a 0 910 pH range of 1.0 to 12 following incubation for 1 to 48 hours in a shaking water bath at 37 C 911 (Yang et al., 2007). Five batches of active pharmaceutical ingredients were tested and three 912 batches of drug product. The release of cyanide was both pH-dependent and incubation-time 913 dependent. The greatest release occurred at pH 1.0 with a gradual decline as the pH 914 increased to 7.0. At this pH the lowest quantity of cyanide was released and as the pH 915 increased again the cyanide concentration also increased. Increasing the incubation time at 916 different pH also increased the amount of cyanide released. The highest cyanide 917 concentration occurred when Prussian blue was incubated at pH 1.0 for 48 hours. The

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918 authors concluded that, based on a dose of 17.5 g of Prussian blue per day, a total of 1.5-1.6 919 mg of cyanide would be released, which was well below the minimum toxic dose of cyanide 920 of 14.4 mg. 921 922 The release of cyanide from potassium ferric (III) hexacyanoferrate (II) and ferric (III) 923 hexacyanoferrate (II) was examined in piglets. The compounds were labelled with carbon- 924 14 in the cyanide group. No carbon-14 dioxide was detected in expired air after ferric (III) 925 hexacyanoferrate (II), indicating that the quantity of cyanide released is very small or nil 926 (Nielsen et al., 1988a). 927 928

929 10. Volunteer Studies 930 931 10.1 Pharmacokinetics 932 There is very little published pharmacokinetic data on Prussian blue in humans. 933 934 10.1.1 Release of iron and cyanide from Prussian blue 935 Three volunteers (all male, 36 years, 81 kg; 38 years, 81 kg; 45 years, 70 kg) were given 936 radio-labelled soluble Prussian blue (500 mg) to determine the release of iron and cyanide in 937 humans in vivo. The compound was labelled with iron-59 in the ferric or ferrous position 938 and carbon-14 in the cyanide group. Only 0.22% of iron (II) and <0.04% of iron (III) was 939 absorbed. Only 2 mg of non-complex bound carbon-14 labelled cyanide was absorbed. This 940 is a factor of 20 to 100 below the lethal dose of 0.5 to 3.5 mg cyanide/kg in humans (Nielsen 941 et al., 1990a). 942 943 10.2 Caesium 944 945 10.2.1 Studies on absorption 946 In a series of volunteer studies on the effect of Prussian blue on the pharmacokinetics of 947 caesium the studies involved self-dosing by the study authors. These authors (to include 8 948 observations, 5 volunteers undertook the study once and one author repeated the experiment 949 twice) showed that 3 g daily of Prussian blue given before caesium-137 did not reduce 950 caesium absorption. The increase in caesium-137 excretion was small following 0.5 g of 951 Prussian blue three times daily (Madshus & Strömme, 1968). 952 953 In 6 volunteers a preliminary study showed that insoluble Prussian blue (4 x 0.5 g or 10 x 954 0.2 g daily for 2 to 3 weeks) did not fully caesium uptake from contaminated food 955 (Volf et al., 1987). 956 957 Two male volunteers (age 36 and 38 years, both 81 kg) ingested three single test meals 958 consisting of 170 g of milk labelled with a tracer dose of caesium-134 along with bread, 959 margarine and cheese 10 minutes after ingestion of 1 g of Prussian blue in gelatine capsules. 960 Both forms of Prussian blue were equally effective in reducing radiocaesium absorption. 961 The absorption of radiocaesium from the meal judged by urinary excretion of caesium-134 962 and whole body retention 14 days after administration was reduced from 100.9% (control 963 without Prussian blue) to 5.6% by soluble Prussian blue and to 6.4% by insoluble Prussian 964 blue (Dresow et al., 1993). In a similar study in two male adult volunteers, the ingestion of 965 Prussian blue ten minutes before eating a test meal containing caesium-134 labelled milk 966 (along with bread, margarine and cheese) reduced the caesium absorption more than the

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967 simultaneous administration of Prussian blue along with the labelled test meal. 968 Administration of Prussian blue prior to the meal reduced absorption of the radiocaesium to 969 3-10% of the ingested dose whereas simultaneous ingestion of Prussian blue and the test 970 meal only reduced absorption to 38-63%. The 100% control was the absorption rate of the 971 radiocaesium test meal alone without Prussian blue (Nielsen et al., 1991). 972 973 10.2.2 Studies on decorporation/excretion 974 In a series of volunteer studies on the effect of Prussian blue on the pharmacokinetics of 975 caesium the studies involved self-dosing by the study authors. Studying the decorporation 976 of caesium involved ingestion of Prussian blue (3 g daily as 2 or 3 doses for several weeks) 977 in 2 adult males given 180 days after ingestion of caesium-137 and it was found to reduce 978 the biological half-life of caesium from the pre-treatment values of 110 and 115 days to 40 979 days. The only adverse effect was mild constipation (Madshus et al., 1966). 980 981 In five cases when Prussian blue (1g three times daily) was given several months after 982 caesium ingestion the biological half-life of caesium was reduced on average from 94 to 31 983 days, that is, to one third of its original half-life (Madshus & Strömme, 1968; Strömme, 984 1968). 985 986 A 37-year-old male was given oral caesium-137 for 24 days followed by 2 g of insoluble 987 Prussian blue (as 10 x 200 mg daily over a 9 hours period) from day 12 to day 17 then after a 988 rest period of 2 days was given Prussian blue for another 5 days. The biological half-life of 989 the caesium was reduced from 140 days to approximately 50 days. There was no 990 constipation and no change in whole-body potassium values (Richmond, 1968). 991 992 Fifteen Chinese exchange students in Bulgaria were exposed to caesium-134 and 137 993 released following the accident at the Chernobyl Nuclear Power Station in April 1986, and 994 following their return to China in June they were assessed for contamination. In three 995 volunteers the biological half-life of caesium ranged from 42 to 71 days. Insoluble Prussian 996 blue (1 g three times daily for 6 days repeated for 3 courses with a 6 day rest period in 997 between) was given from day 114 to 145 after exposure. This reduced the half-life of 998 caesium and enhanced elimination (Tang et al., 1988). 999 1000 10.3 Rubidium exposure 1001 There are no volunteer studies on the effect of Prussian blue in rubidium exposure. 1002 1003 10.4 Thallium poisoning 1004 There are no volunteer studies on the effect of Prussian blue in thallium exposure. 1005 1006 Bhardwaj et al. (2006) studied the effect of insoluble Prussian blue on whole body 1007 radioactivity in two patients following thallium-201 myocardial scintigraphy. Each patient 1008 had two sessions of scintigraphy, one with and one without Prussian blue (100 mg 3 times 1009 daily after meals for 3 days), so each patient acted as their own control. In the first patient 1010 whole body radioactivity was reduced by 18 and 30% after 24 and 48 hours, respectively, of 1011 oral Prussian blue therapy. The second patient developed constipation and did not pass any 1012 stools after oral Prussian blue for 48 hours. The whole body radiation counts were similar to 1013 those when Prussian blue was not given but there was a concentration of radioactivity in the 1014 colon suggesting that the radioactivity was unavailable for resorption. 1015 1016

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1017 11. Clinical Studies – Clinical Trials 1018 1019 There are no controlled clinical trials on the use of Prussian blue in human thallium poisoning or 1020 radiocaesium decorporation. 1021 1022

1023 12. Clinical Studies - Case Reports 1024 1025 12.1 Decorporation of radiocaesium 1026 1027 12.1.1 Goiânia incident, Brazil, 1987 1028 At the end of 1985 a private radiotherapy institute moved premises and left a caesium-137 1029 teletherapy unit behind. The building was partly demolished and in 1987 two men removed 1030 the source assembly head from the machine thinking it may have scrap value but without 1031 being aware of what it was. They took this home and tried to dismantle it during which they 1032 ruptured the source capsule. This contained which is highly soluble and 1033 easily dispersed. After the rupture the source capsule was sold for scrap to a junkyard 1034 dealer. He observed that the material glowed blue in the dark and over the next 5 days this 1035 was a source of interest to family and friends. During this time several individuals started to 1036 become unwell with gastrointestinal signs and eventually the source capsule was suspected 1037 and it was taken to the public health department. This triggered a major response to the 1038 incident. In total 112,000 people were assessed and 249 were found to contaminated either 1039 internally or externally with caesium-137. Some had very high exposure due to eating with 1040 contaminated hands or rubbing the glowing material over their body. Twenty patients 1041 developed suppression. Eight developed acute radiation syndrome (Brandão- 1042 Mello et al., 1991) and four of these victims died within 4 weeks of their admission to ® 1043 hospital (IAEA, 1988). Prussian blue (Radiogardase -Cs) treatment was given to 46 1044 patients (aged 4 to 46 years) for up to 150 days. The adults were initially given 3 g daily and 1045 the 13 children were given 1 to 1.5 g daily. These doses were later increased to 10 g and 3 g 1046 daily, respectively, when it was established that larger doses resulted in higher radioactivity 1047 of faecal samples. In four cases 20 g of Prussian blue was given over 24 hours. Of 46 1048 patients, 10 developed mild to moderate constipation and this was managed with a high fibre 1049 diet or laxatives (Farina & Brandão-Mello, 1991). Prussian blue treatment significantly 1050 increased the rate of faecal caesium excretion and reduced whole body retention of caesium 1051 (IAEA, 1988). The physiological faeces to urine excretion ratio of caesium was 1:4 and this 1052 was changed to 4:1 with Prussian blue treatment (Farina & Brandão-Mello, 1991). In 15 1053 adult patients who received Prussian blue the body burden of caesium-137 was reduced by 1054 51% to 84% with an average of 71% within the first 2 months after exposure. This dose 1055 reduction was independent of the Prussian blue dose in the range of 3 to 10 g/day (Melo et 1056 al., 1994). 1057 1058 In vivo data from patients internally contaminated with caesium-137 in the Goiânia accident 1059 was analysed to compare the half-life of caesium-137 with and without Prussian blue 1060 treatment. Additionally the possible influences of various body parameters (age, height, 1061 weight and radioactivity) on the half-lives were evaluated. Subjects were monitored using a 1062 whole-body counter and the findings are from data collected for the period of one year post 1063 the accident. Patients under treatment had previously followed different Prussian blue dosing 1064 patterns but during the monitoring period received 3 g/day, 6 g/day or 10 g/day. Caesium- 1065 137 elimination from the body followed first order kinetics with or without Prussian blue

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1066 therapy. Without Prussian blue treatment the half lives of caesium-137 in the 10 adult 1067 females studied varied widely (range 39 to 104 days; mean: 65.5 days) with less variation 1068 seen in the 8 adult males (66 to 106 days; mean: 83 days). Overall Prussian blue reduced the 1069 half-life of caesium-137 by about 32%. The actual calculations showed that for those 1070 subjects receiving 3 g/day of Prussian blue the mean reduction was 28%, it was 31% in those 1071 receiving 6 g/day and was 32% in subjects receiving 10 g/day. The strongest parameter 1072 influencing the half-life in both males and females was body weight. Height and age were 1073 correlated to the half-lives through their correlation to the weight parameter but were not 1074 additional variables. Investigating the estimated caesium-137 body burden at the initial time 1075 of elimination found an inverse relationship between initial activity and half-life: the larger 1076 the initial body burden, the faster the nucleotide removal from the body. The influence of 1077 this parameter was much weaker than that of body weight (Lipsztein et al., 1991). 1078 1079 12.1.2 Chernobyl incident, 1986 1080 Measurements were made on 15 Chinese subjects internally contaminated with th 1081 radionucleotides released from the Chernobyl accident on 26 April 1986. The students had th 1082 been visiting Sofia, Bulgaria from 19 April until 23 May 1986 and monitoring was done on 1083 their return to Beijing. Internal contamination with radioactive caesium (caesium-134 and 1084 caesium-137) was measured in all 15 students. The measured activity in the body for the 15 1085 volunteers ranged from 68-840 Bq for cesium-137 and 110-630 Bq for caesium-134. The 1086 estimated intakes were calculated and ranges were 95-1200 Bq (Caesium-137) and 170-930 1087 Bq (Caesium-134). The biological half-life was calculated for three volunteers along with 1088 the effect of Prussian blue on their rate of elimination of radiocaesium. Prussian blue was 1089 given at doses of 1g three times a day for a 6 day course, 3 courses in total were give with a 1090 6 day time interval between each course. The biological half-life of the radiocaesium ranged 1091 from 43-71 days. Prussian blue was given to the three volunteers in the period of 114-141 1092 days after contamination and the body retention of radiocaesium declined more rapidly 1093 following Prussian blue administration than in those of controls (Tang et al., 1988). 1094 1095 12.1.3 Other case reports 1096 Five persons, two adults (aged 34 and 38 years, weight 56 and 55 kg) and three children 1097 (aged 4 to 11 years; weight 13.5 to 34 kg) accidentally received caesium-137 chloride for 1098 approximately 20 days (no details given). The patients were evaluated as soon as the 1099 accident was discovered and started on Prussian blue. The adults received 3 g daily from 1100 days 35 to day 128 or 143. The children received 1, 1.5 or 2 g daily from days 35 to 86, 76 1101 or 99. The half-life of the caesium was very variable and was 124, 54, 61, 36 and 36 days 1102 without treatment. With Prussian blue treatment the half-life of caesium was 38, 39, 25, 17 1103 and 16 days, respectively (Ma et al., 1985). 1104 1105 12.1.4 Non-radioactive caesium 1106 A 65 year old woman presented to a hospital accident and emergency department with a one 1107 day history of recurrent fainting. The patient claimed to have essential hypertension and was 1108 having treatment for this from her family doctor. Six months prior to her presentation she 1109 had been diagnosed with rectal cancer and liver metastasis and she had experienced frequent 1110 episodes of watery diarrhoea in the past 4 weeks. On admission her blood pressure was 0 1111 138/55 mmHg, pulse 52/min regular, temperature 36.8 C, respiratory rate 18/min and blood 1112 glucose 10.7 mmol/L. She developed and episode of Torsades de points (TDP) polymorphic 1113 ventricular tachycardia with transient loss of consciousness during initial assessment. The 1114 arrhythmia spontaneously converted back to normal sinus rhythm in about 10 seconds. Her 1115 electrocardiogram (ECG) showed QT prolongation with a corrected QT interval of 620 ms

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1116 calculated by Bazett’s formula. Serum electrolytes showed mild hypokalaemia (2.8mmol/L) 1117 whilst serum magnesium and serum calcium were normal. She was treated with intravenous 1118 magnesium sulphate and vigorous potassium replacement however there was no 1119 improvement in the QT prolongation after these therapies. At this stage it was discovered 1120 that in the previous 6 weeks the patient had been taking anticancer naturopathic drugs 1121 obtained from an alternative medical centre in Hong Kong. A detailed drug history was 1122 obtained. Her medications included methyldopa, Dologesic™ (dextropropoxyphene 32.5mg 1123 and paracetamol 325mg) and Lomotil™ (diphenoxylate 2.5mg, 0.025mg) all 1124 prescribed by her family physician. In addition, a bottle of herbal powder (1 teaspoon taken 1125 daily), 3 oral medications including “Gigamax” (labelled as multivitamins), Slow K™ (slow 1126 release potassium supplement) and “multi-C” were prescribed by the naturopathic 1127 practitioner. On the basis of the clinical findings and along with previous case reports of 1128 caesium chloride use in anticancer therapy, the diagnosis of caesium poisoning was highly 1129 suspected. Whole blood and serum were assayed and the serum caesium concentration was 1130 elevated markedly at 288µmol/L (normal range 0.0045-0.0105µmol/L). Whole blood 1131 arsenic concentration was normal. One of the naturopathic medicines (“multi-C”) was found 1132 to contain 89% caesium chloride by weight. No undeclared contents or toxins were found 1133 on analysis of the herbal powder “Gigamax” and Slow K™ tablets. The patient was 1134 hospitalized for 2 weeks with intensive cardiac assessment and monitoring. The use of 1135 naturopathic medicines was stopped after hospitalization. Oral Prussian blue 3g 3 times 1136 daily was started on day 7 after hospital admission and continued for 4 weeks (day 7 to day 1137 34). Hypokalaemia was noticed during Prussian blue therapy and an oral potassium 1138 supplement was given to keep the serum potassium concentration at around 4mmol/L. Serial 1139 serum and urine caesium concentrations were measured. The calculated serum half-life of 1140 caesium was shortened from 61.7 days to 29.4 days with Prussian blue therapy. The 1141 corrected QT interval of her ECG returned to normal baseline on day 27 (Chan et al. 2009). 1142 1143 Thurgur et al. (2006) report a case where Prussian blue was used in the treatment of non- 1144 radioactive caesium. The patient was a 58 year old female with chronic caesium toxicity 1145 from the use of caesium chloride as an alternative cancer therapy. High levels of caesium are 1146 arrhythmogenic and this patient showed recurrent syncope, polymorphic ventricular 1147 tachycardia, hypokalaemia, and a QT prolongation of 690 ms. Along with conventional 1148 measures Prussian blue was used to treat her caesium toxicity. The Prussian blue treatment 1149 decreased the half-life of caesium from 86.6 days to 7.9 days, with associated normalization 1150 of QT interval and cardiac rhythm. 1151 1152 1153 12.2 Rubidium 1154 There are no reports on the use of Prussian blue in rubidium exposure in humans. 1155 1156 12.3 Thallium poisoning 1157 Reference values for thallium (Walker, 1998): 1158 Blood <5 nmol/L (<1 µg/L) 1159 Urine <5 nmol/L (<1 µg/L) 1160 1161 A 45 year old man with no significant medical history was hospitalized with peripheral 1162 neuropathy and paraesthesia of the extremities of all 4 limbs for 2 days. Clinical 1163 examination revealed an erythematous popular rash on the face and folliculitis on the lower 1164 limbs associated with hyperaesthesia of the feet and hands. An electromyogram showed 1165 severe polyneuropathy. The patient had a cardiac arrest 9 days after admission followed by

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1166 post-anoxic coma. The patient worked in a technical factory and handled thallium, 1167 bromide, caesium and iodide. The urinary thallium concentration measured twenty days 1168 after his cardiac arrest was 5118µg/g of creatinine. Despite the late diagnosis treatment with 1169 insoluble Prussian blue (Radiogardase™) was started 2 weeks later (42 days from his 1170 original admission to hospital) and continued for 24 days at a dose of 6g three times daily. 1171 Urinary thallium concentrations decreased from 1333µg/g of creatinine to 166µg/g of 1172 creatinine. After evaluation of the efficacy of the treatment a second course of Prussian blue 1173 therapy was started at 77 days post admission and continued for 1 month when the patient 1174 died. The urinary thallium concentration decreased from 86µg/g of creatinine to 3µg/g of 1175 creatinine (Villa et al., 2009). 1176 1177 Ten members of two families (family A and family B) sought treatment at a health care 1178 facility in Baghdad, Iraq. All patients were experiencing , abdominal pain and 1179 dysphagia. Over the next 4 days, 5 of the patients developed neurological signs and 1180 symptoms of varying severity (pain, abnormal sensations and weakness-particularly of the 1181 lower limbs). Four days after admission biological samples and a sample of a cake that all 1182 10 patients had consumed were submitted for toxicology testing. Thallium was detected in 1183 both the biological samples and the cake. On the eighth day after admission one of the 1184 patients, a child aged 11 years, died and two days later the 9 surviving patients were 1185 evacuated to Jordan for Prussian blue therapy which was not available in Iraq. A second 1186 patient, a 2 year old child died soon after arrival in Jordan, prior to receiving therapy. 1187 Prussian blue therapy was begun in the 8 surviving patients, now 11 days after they had 1188 eaten the cake. Two of these 8 patients were already comatose with severe cerebral oedema 1189 and subsequently died. Over the next thirty days, all 6 long-term survivors developed hair 1190 loss and 5 of the 6 survivors developed muscle weakness and spasticity of the lower limbs, 1191 with differing severity. An epidemiological investigation was started and it was discovered 1192 that the fathers of the two families were both board members of an Iraqi sporting club and 1193 had attended a routine board meeting on the day before hospital attendance in the club’s 1194 conference room. The cake, prepared by a local bakery and pre-divided into 10 pieces, had 1195 been delivered to the board meeting as a gift from a former board member. However the 1196 cake arrived late, after most board members had already left the meeting so no cake was 1197 eaten then. The two members that remained (the fathers of the two families) divided the 1198 cake and took the halves home to their families and it was eaten by both families at home 1199 that evening. Family A comprised seven members (parents and five children) and family B 1200 comprised five members (parents, two children and an uncle). Ten cases of abdominal pain, 1201 vomiting and dysphagia were identified among family members who consumed any amount 1202 of the cake. No other board members or their families were ill and no similar illnesses were 1203 reported at the health facility in Baghdad or at any nearby health facilities. Food exposure 1204 histories were collected in Jordan through interviews with family members. Ten people who 1205 ate portions of the cake became unwell; neither of the two persons who did not eat cake 1206 became unwell, however one of the two had tasted some of the cake icing and although 1207 asymptomatic, his blood and urine samples tested positive for thallium. A more rapid onset 1208 of illness occurred in adults and in persons who ate the most cake. Fatality was not 1209 significantly associated with sex, age, the amount of cake eaten, or the time to illness onset. 1210 Quantitative thallium levels were determined from blood and urine samples of nine patients 1211 taken on day 16 after eating the cake. Thallium was detected in all nine patients; the median 1212 blood thallium level was 289µg/L (range 53-1,408 µg/L; reference range expected <2µg/L), 1213 and the median calculated 24 hour urine excretion of thallium was 3063 µg/L (range 542- 1214 12,556 µg/L; reference range expected <5µg/L). Blood thallium levels correlated weakly 1215 with the amount of cake reported to have been eaten (CDC., 2008)

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1216 1217 A previously healthy forty-year old male was admitted to hospital complaining of 1218 progressive weakness of his limbs, repeated vomiting and recurrent episodes of confusion. 1219 He had initially presented to the hospital six weeks previous to this admission complaining 1220 of thirst, nausea, vomiting, dizziness, parathaesias and arthralgias (predominantly of the 1221 lower limbs). The paraesthesias were not described as ascending or notably painful. A 1222 supine blood pressure of 180/90mmHg and a mild fever were the only physical findings 1223 noted. Neurological examination showed the cranial nerves to be intact and the results of 1224 muscle strength tendon reflex and cerebellar function tests were normal. There was no 1225 evidence of impaired superficial or position senses. The results of a complete blood cell 1226 count and blood chemistry tests were normal. He was discharged and symptomatic 1227 treatment (non-steroidal anti-inflammatory drugs) was prescribed. Two weeks before the 1228 present admission he returned complaining of general weakness, anxiety, myalgias 1229 (particularly of the legs), delayed growth of facial hair after shaving and thirst. Physical 1230 examination and routine laboratory tests were again normal. His symptoms were diagnosed 1231 as “non-specific”, partly attributed to stress. On his present final presentation he was alert 1232 and complained of double vision. Physical examination revealed hyperhidrosis, tachycardia 1233 (100beats/min) and supine blood pressure of 140/100 mmHg. Alopecia of the scalp was 1234 noted but eyebrows, eye lashes and body hair were intact. Neurological assessment 1235 disclosed horizontal and upbeat nystagmus, severe weakness of the lower extremities (more 1236 prominent proximally), bilateral absence of Achilles tendon reflexes, and lower limb ataxia. 1237 He did not complain of extreme pain and no objective signs of sensory changes were 1238 detected. He was found to have raised blood alanine aminotransferase and raised aspartate 1239 aminotransferase. There was no evidence of proteinuria. Lumbar tap revealed an elevated 1240 protein. Electroencephalogram showed persistent generalized slowing; the electromyogram 1241 displayed bilateral, severe, lower limb motor axonal neuropathy. Rapid deterioration of his 1242 neurological state was observed over the next few days, including flaccid paraparesis, lower 1243 limb areflexia, severe sensory impairment, mild distal arm and neck weakness, as well as 1244 occasional urinary and faecal incontinence. Visual disturbance progressed from impaired 1245 colour vision and decreased acuity to optic disc atrophy. Cognitive disturbances and 1246 hoarseness were also noted. Sural nerve showed early acute axonal degeneration 1247 with no evidence of vasculitis. At this stage he was started on intravenous immunoglobulin 1248 as a variant of Guillain-Barré syndrome (Milllar-Fisher type) could not be excluded, and 1249 alternative causes were explored. Several days later thallium poisoning was diagnosed as 1250 heavy metal urinalysis showed renal thallium excretion of 7mg/24 hours. Prussian blue was 1251 administered (250mg/kg/day, dissolved in 15% mannitol) daily through a nasogastric tube, 1252 along with forced diuresis. At this stage Mees’ lines appeared on his nail beds. No 1253 improvement in his general state was noted within the next two weeks. He became drowsy, 1254 required respiratory assistance and subsequently developed aspiration pneumonia. The 1255 suspicion of cardiotoxicity was raised by elevations of alanine aminotransferase, aspartate 1256 aminotransferase and creatine phosphokinase (MB fraction) levels however this was not 1257 substantiated by electrocardiographic follow-up and echocardiography. There was swelling 1258 and pain in his right knee, however, some clear fluid drained was inconsistent with any 1259 particular diagnosis. Rheumatoid factor was mildly elevated, without any other supporting 1260 evidence of concurrent arthritic disease. His condition stabilized over the next weeks and 1261 there was a gradual decrease in thallium urinary output (table below). He was transferred to 1262 a rehabilitation hospital after forty two days. A follow-up examination 3 months later the 1263 alopecia and Mees’ lines had completely resolved. There were no cognitive disturbances, his 1264 proximal strength was restored and his colour vision was normal. Decreased visual acuity 1265 and bilateral drop-foot were still evident however and the patient had only a vague

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1266 recollection of his period in hospital. 1267 Daily excretion of thallium in urine Hospitalization day Daily excretion (mg/day) 9 7 17 9 18 7.6 19 3.2 21 5.6 24 2.9 25 2.9 28 2.5 38 0 1268 (Atsmon et al., 2000) 1269 1270 A 67 year old woman was admitted to a county hospital because of acute pain in the chest, 1271 abdomen and lower limbs. The pain in the lower limbs was described as burning and severe. 1272 There was no vomiting or diarrhoea. The patient was discharged after 3 days with the cause 1273 of her illness undetermined. The patient then presented one week later to a private clinic 1274 because of persistent symptoms. Physical examination showed mild tenderness over the 1275 abdomen and electrocardiography showed non-specific T-wave inversion in the anterior 1276 waves. Routine urinalysis results were normal and laboratory tests showed normal blood 1277 cell counts, blood glucose level and kidney and liver function. The patient was treated 1278 symptomatically with analgesics. She was soon readmitted to hospital because of fainting 1279 spells and persistent symptoms. The patient suspected she was being poisoned. Paranoid 1280 psychosis and trichotillomania were diagnosed and she was admitted under restraint to the 1281 psychiatric department for observation. After being discharged home at week 5, the woman 1282 then re-presented at the private clinic. According to the patient the pain had become less 1283 severe. Physical examination showed diffuse alopecia which had started 2 weeks after the 1284 onset of the initial symptoms. Thallium poisoning was suspected. Urinalysis showed a 1285 thallium level of 8.56µmol/L (normal level, 0.003µmol/L; a level of 0.98µmol/L is toxic). 1286 The case was reported to the police and the prime suspect was the patient’s 73 year old 1287 cohabitant. A dose of activated charcoal was given in the emergency department of a nearby 1288 district hospital and the patient was discharged home with a 2-week supply of succimer (2, 1289 3-) (no reason for this was given). At follow-up at week 6 however, 1290 it was found that the pain in her chest, abdomen and lower limbs had subsided, but 1291 symptoms of peripheral neuropathy had emerged - namely bilateral numbness and loss of 1292 exteroceptive and proprioceptive sensations in the toes. She had mild weakness in the 1293 proximal muscles of the lower limbs, as indicated by the patient’s difficulty in rising from 1294 the squatting position. She also complained of right-sided headache and tachycardia. She 1295 was immediately admitted to a University Medical Centre for treatment with oral Prussian 1296 blue (potassium ferric hexacyanoferrate) 4g every 8 hours. The blood thallium level at the 1297 time of hospitalization was 0.15µmol/L. No other were present. The patient 1298 tolerated Prussian blue very well, but hypoaesthesia developed over the medial aspect of her 1299 left calf on the second day of treatment. She was discharged 1 week later, when the urine 1300 level of thallium was 0.14µmol/L. In week 9 the patient experienced neurological 1301 deterioration, impairment of short-term memory, double incontinence, tremor ataxia and 1302 falls. Physical examination showed hypoaesthesia of the right trigeminal nerve, general 1303 weakness of the extremities, cerebellar ataxia, tremor and dyskinesia. Plantar reflexes were 1304 normal and the urine thallium level was 0.33µmol/L. By week fourteen the right facial

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1305 numbness fully recovered, by week twenty there was recovery of sphincter control and 1306 regrowth of hair. Urine thallium was undetectable 2 weeks later. The weakness in the lower 1307 limbs, unsteady gait, falls and bruises persisted until 11 months after the initial presentation, 1308 when her condition improved noticeably although residual weakness continued (Pau., 2000) 1309 1310 A 24-year old female was admitted to hospital with a 4-month history of illness. Eight days 1311 after admission a diagnosis of thallium poisoning was made based on rapid diffuse alopecia, 1312 gastro-intestinal disturbance and a worsening neurological state combined with laboratory 1313 results. Whole blood thallium was measured and the first level was 300 µg/L (normal 1314 <10µg/L, toxic >100 µg/L); the corresponding 24-hour urinary thallium value was 4300 1315 µg/L (normal < 10 µg/L, toxic > 200 µg/L), consistent with severe intoxication. Colloidal 1316 soluble Prussian blue (KFe[Fe(CN)6]) was given orally at a dose of 250mg/kg/day in 2-4 1317 divided doses for 14 days. The patient also received mannitol as a cathartic, cisapride for th 1318 her persistent constipation (started on the 5 day of treatment) and forced diuresis, which 1319 was achieved with furosemide, glucose, potassium and . The patient’s 1320 clinical course after treatment was uncomplicated with recovery of her vital signs within a 1321 week. Four months after treatment thallium was undetectable in a 24-hour assay. However 1322 after 6 months she still suffered from a lack of concentration and insomnia and never fully 1323 returned to her previous functional level. The source of thallium was never established 1324 Days before/after treatment 24 hour-urinary thallium Whole blood thallium level level µg/L µg/L -8 4300 0 4200 300 4 1450 < 10 10 330 14 468 18 440 31 260 1325 (Vrij et al., 1995). 1326 1327 1328 A thirty nine year old male with a history of heavy consumption became ill one 1329 week after returning from holiday in Spain. His symptoms started acutely with generalized 1330 pain and tingling all over his head and body and he was admitted to hospital where he was 1331 greatly distressed and complaining of shooting pains in his legs and back and leg weakness. 1332 It was thought initially that his illness was an alcoholic syndrome. Because of his continued 1333 deterioration despite multivitamin treatment he was transferred initially to the regional 1334 neurology unit and then further to an intensive care unit, 20 days after first becoming ill. On 1335 initial neurological examination he had respiratory distress, evidence of scalp hair loss, gaze 1336 evoked nystagmus in all directions, bilateral lower motor neuron, facial and bulbar 1337 weakness; his arms were minimally weak with normal reflexes but his legs showed a flaccid 1338 paralysis with absent reflexes. Initial biochemical investigations were within normal ranges 1339 (electrolytes, urea, creatinine and liver function tests). The electrophoresis pattern of serum 1340 proteins was normal and a non-specific auto-immune profile did not detect any auto- 1341 antibodies. Screening tests for abnormal urinary porphyrins gave negative results. A 1342 complete blood count was within the reference range, except for a slightly increased mean 1343 corpuscular volume. Cerebrospinal fluid was acellular with a protein count of 1.5g/L. 1344 Nerve conduction studies showed absent sensory and motor responses for the legs but 1345 normal values for the arms. On the basis of the clinical picture at this stage, in particular

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1346 with respect to the hair loss, serum and urine were assayed for thallium and it was found to 1347 be present at toxic levels (value not stated). The patient deteriorated further and developed 1348 visual failure, complete external ophthalmoplegia, and total arreflexic paralysis of all limb 1349 and neck muscles. He was given two treatments of plasma exchange and treatment with 1350 potassium ferrihexacyanate (colloidal soluble Prussian blue), 5g every 6 hours by 1351 nasogastric tube was started (now thirty five days into the illness) and continued for 2 1352 months. At the same time intravenous potassium supplements (100-400mmol/day) were 1353 given. Excretion concentrations of thallium in serum and urine are tabulated below. The 1354 patient made a slow recovery which was complicated by septicaemia, recurrent 1355 supraventricular tachycardias and psychosis. He required 96 days of assisted ventilation and 1356 a total of 224 days in hospital. Some 500 days after the initial insult he still had a significant 1357 visual handicap, no fine finger function and could only walk a few steps with assistance. 1358 The source of the thallium poisoning was never discovered. 1359 1360 Excretion of thallium in serum Hospitalization day Excretion (nmol/L) 28 914.4 46 54.8 48 25.4 49 19.0 55 8.8 56 6.9 57 6.9 58 7.5 59 6.3 60 7.4 1361 1362 Excretion of thallium in whole blood Hospitalization day Excretion (nmol/L) 42 156.5 43 190.7 44 141.8 45 92.2 50 39.1 51 36.7 52 14.7 56 7.8 58 7.8 59 5.4 1363 Day zero = date of admission to hospital 1364 (Chandler et al., 1990) 1365 1366 Villanueva et al (1990) describe 5 cases of thallium poisoning in Spain. Four were members 1367 of a single family and the source of thallium was never ascertained. The fifth case was a 1368 female adult with a history of depression who intentionally ingested a thallium sulphate 1369 . The family members (2 adults and two children aged 10 years and 3 years) 1370 presented initially with varying symptoms and the two children required admission to

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1371 intensive care. Total hair loss of the 10-year old 2 weeks after admission led to the 1372 diagnosis of thallium poisoning. In all of the 5 cases urinary thallium was measured. The 1373 10 year old child’s first measured thallium level was 18.4 mg/L, seven days later it was 5.8 1374 mg/L, at 27 days later it was 0.14 and by approximately 4 months it was 0.004 mg/L. The 1375 child had been treated with Prussian blue at a dose of 250mg/kg by duodenal tube 1376 administration every 6 hours until the urinary excretion of thallium was 0.5 mg/day. Her 1377 symptoms resolved in 20 days. The female adult who intentionally ingested the thallium had 1378 an initial urinary level of 2.4mg/L which decreased to 0.9mg/L in 4 days and to 0.1 mg/L in 1379 5 weeks. She had also been given Prussian blue at a dose of 250mg/kg/day (duration of 1380 treatment not stated) and 6 weeks later recovery was complete. The case reports also outline 1381 the thallium concentrations of the other family members but it is unclear whether or not they 1382 received Prussian blue therapy. 1383 1384 A 20-year-old chemistry student presented with a 3 day history of malaise and polyuria with 1385 paraesthesia of the fingers and lips. He had been using thallium and blood and urine 1386 analyses showed very high concentrations (5750 µg/L and 60000 µg/L, respectively), 1387 although he denied ingestion. Tests for thallium in hair samples were negative which 1388 excluded chronic ingestion however, despite exhaustive enquiries by the police and college 1389 authorities, the mode of thallium administration remained undetermined. Shortly after 1390 admission he became drowsy and had a convulsion. He developed progressive weakness 1391 over the next 12 hours, with severe pain in the calf muscles. He had peripheral and sensory 1392 impairment, dysarthria, dysphagia, paralytic ileus, tachycardia and ECG changes. He was 1393 started on soluble Prussian blue (5 g in 50 ml of 15% mannitol 4 times daily) and forced 1394 diuresis. On day 6 dialysis and haemofiltration were started and diethyldithiocarbamate was 1395 given. The diethyldithiocarbamate produced a short-lived increase in thallium excretion 1396 (from 17 to 142mg/24 h in urine, and 63 to 81mg/6 h dialysis) but also led to a rise in serum 1397 thallium concentration (from 800 to 1350 µg/L and worsening neurological signs (including 1398 respiratory failure) and it was stopped. He required ventilation for 4 weeks. He developed 1399 almost complete alopecia before hair regrowth started. By 13 weeks he could swallow fluids 1400 and by 20 weeks he had normal upper limb function. But after 12 months he was still in a 1401 wheelchair owing to nerve damage to the lower limbs. Prussian blue and laxatives were the 1402 most effective means of enhancing thallium elimination, even though paralytic ileus caused 1403 long periods of constipation. High concentrations of thallium were present in the faeces up 1404 to day 18, and it was estimated that at least 2000 mg of thallium was excreted via this route 1405 in the first 20 days (see table). This is twice the quantity excreted by all the other methods 1406 in the same period. Forced diuresis was estimated to have eliminated 820 mg of thallium in 1407 46 days with 225 mg eliminated via haemodialysis in 25 days. 1408 No. of days from Concentrations Excretion admission Serum Urine Urine Faeces Dialysate Urine (µg/L) (µg/L) (mg) (mg) (mg) Filtrate (mg) 1 5750 60000 129 2-5 2390 35900 398 6-10 680 7750 230 550 146 3.5 11-15 225 1520 42 155 64 1.0 16-20 35 640 12 1280 5.0 <0.1 21-25 15 280 2.6 0.5 5.4 <0.1

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26-30 12 315 2.7 2.5 4.5 <0.1 31-40 11 110 2.9 41-50 <7 64 1.0 51-70 <7 24 0.9 Total quantities excreted (mg): 820 1990 225 4.5 1409 1410 The serum and urine concentrations presented in the table above are daily averages for the 1411 periods stated. The quantities excreted are totals for each period. All values are given as 1412 elemental thallium, measured by atomic absorption spectrometry (Wainwright et al., 1988). 1413 1414 1415 A group of 14 people ate dinner together and the following morning all complained of 1416 abdominal pain, vomiting and diarrhoea and were taken to hospital. Five of the 14 patients 1417 died in hospital within the next four days. The remaining nine patients (age range 16-70 1418 years) were seen over the following 4-7 days. One patient was pregnant, one had pre- 1419 existing Parkinson’s disease and a third had elephantiasis of the legs. All of them 1420 complained of varying degrees of pain in the feet and legs and 6 were unable to walk. Some 1421 had headaches, constipation, abdominal pains, chest discomfort, loss of appetite, and loss of 1422 sleep. On examination they all had varying degrees of peripheral neuritis with 1423 hyperaesthesia, hyperalgesia, mental confusion, tachycardia, muscle and abdominal 1424 tenderness. A diagnosis of heavy metal poisoning was made based on the sudden onset of 1425 peripheral neuritis in a group of people who had eaten the same food and also because of 1426 similar case presentations in the previous year that had proven to be thallium poisoning. 1427 Blood and urine samples were sent for heavy metal analysis. Thallium was detected 1428 quantitatively in all the samples. The food they ate was suspected of being contaminated 1429 with a thallium-containing rodenticide but this could not be confirmed. Prussian blue was 1430 started between the 4th and 7th day at a dose of 2g three times a day orally, together with 1431 magnesium sulphate solution (30ml three times a day) to avoid constipation. For the first 1432 few days some of the patients required parenteral pethidine (100mg every six hours) for 1433 severe lower limb pain later changed to oral pentazocine (50mg three times a day). They 1434 were also given oral vitamin B complex and amiloride/hydrochlorothiazide daily. Over the 1435 next 5 days the leg pains rapidly subsided and analgesics were stopped. Seven of the 1436 patients started walking freely without much pain. In the 3rd week all of them gradually 1437 started losing scalp hair and there was almost complete alopecia after 4 weeks. Six patients 1438 developed Mee's lines on the finger nails along with hyperpigmentation over the knuckles. 1439 Four patients were discharged from hospital after the 4th week and the remaining patients 1440 after 6 weeks. Prussian blue was continued in all patients at the same dose for a total of 6 1441 weeks and no side effects were noted. On discharge 5 patients had fine tremor of the upper 1442 limbs with slight incoordination of movement. After sixteen weeks regrowth of scalp hair 1443 was complete. The pregnant woman had a premature delivery in the sixth month of 1444 pregnancy at another hospital but no details were available. By sixteen weeks all patients 1445 had returned to active life. 1446 Serum and urine thallium concentrations after two weeks of Prussian blue therapy Patient number Patient age (years) Serum Thallium Urine Thallium (µgm/ml) (µgm/ml) 1 62 16 2200 2 37 4 48 3 29 4 250 4 70 4 250

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5 60 6 660 6 16 11 1100 7 29 5 37 8 30 6 87 9 26 4 23 1447 (Pai., 1987) 1448 1449 A 21-month old girl arrived at a hospital in the UK from Qatar for investigation of ataxia of 1450 five days duration. On admission she was semi-conscious and irritable but extensive 1451 investigations including routine toxicology screen on blood and urine, could only achieve a th 1452 diagnosis of encephalopathy. On the 5 day of hospitalization hair loss was noted and a 1453 suggestion of thallium poisoning was made by a nurse in charge of the child (the subject of 1454 the crime thriller she was reading at the time, Agatha Christie’s A Pale horse). The child 1455 was treated with oral Prussian blue and potassium chloride (doses not stated) and after 3 1456 weeks of treatment there was marked clinical improvement and no thallium was detected in 1457 her urine. The clinical improvement continued and after four months the child was alert, 1458 ataxic but able to walk with help. The source of thallium was never determined but was 1459 thought to be the cockroach bait that was used in the home (Robb-Smith, 1987). 1460 1461 A 32 year-old woman was admitted to hospital 6 hours after allegedly ingesting 100mg of 1462 thallium sulphate. On admission the patient had malaise, nausea, vomiting and a burning 1463 retrosternal pain. After gastric lavage Prussian blue was administered 250mg/kg in divided 1464 doses) and combined haemoperfusion-haemodialysis (HP-HD) was started 7 hours after the 1465 alleged ingestion. HP-HD was instigated because the dose of thallium and whether there 1466 were possible co-ingestants were unknown. The patient was treated for 4 hours by HP-HD. 1467 Blood samples were taken before and after the charcoal column and after the artificial 1468 kidney at 1 hour intervals. Blood flow was 200ml/min. After HP-HD treatment 7 more 1469 blood samples were taken, initially at 1-hour intervals later at 2-hour intervals. During HP- 1470 HD treatment the patient’s blood pressure remained constant however there were frequent 1471 supraventricular extrasystoles. The patient improved and after 4 hours of HP-HD she felt 1472 well. Subsequent to her treatment no neurological or dermatological symptoms were noted. 1473 The combination of HP-HD in this patient obtained an overall clearance of 150ml/min in 1474 blood compared to 47ml/min (mean value) using HD only (De Backer et al., 1982). 1475 1476 A 28-year-old female presented 4 days after ingestion of nearly 1 g of thallium sulphate with 1477 lower abdominal pain, nausea, and hyperaesthesia of the limbs. Thallium was detected in 1478 the urine (3 mg/L) and gastric aspirate (10.8 mg/L). She was started on intravenous fluids 1479 and soluble Prussian blue (5 g four times daily via duodenal tube with 50 mL of 15% 1480 mannitol). Over the next two days the abdominal and lower limb pain persisted, with 1481 drowsiness and vomiting. On the third day she became hypotensive with bilateral ptosis. 1482 Alopecia was also noted. Neurological signs and gastrointestinal symptoms began to 1483 improve 4 days later, but hair loss continued. She had severe constipation for the first 6 days 1484 despite laxative administration. By 11 days her symptoms improved; Prussian blue was 1485 discontinued after 20 days, by which time hair regrowth had started. Thallium 1486 concentrations fell dramatically over the first 2 days of admission, with a slower decline th 1487 thereafter (see table). No faecal samples could be obtained until the 10 day after poisoning 1488 (hospital day 6) owing to the severe constipation. At this time 1.6 mg of thallium was 1489 detected in the 24 hour faeces and 1.93 mg in the urine. In 28 days of hospitalization 1490 approximately 5 mg of thallium was eliminated via the intestinal tract and 35 mg in the 1491 urine. Approximately 55 mg was eliminated in the saliva between days 9 and 26 after

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1492 poisoning. In total the quantity eliminated was thought to be <5% of the dose ingested. She 1493 was discharged asymptomatic at 28 days. 1494 Excretion of thallium (mg/day) in urine, faeces and saliva at various times during hospital course No. days after Urine (mg/day) Faeces (mg/day) Saliva (mg/day) poisoning 4 5.68* Not determined Not determined 5-6 4.54 Not determined Not determined 7-9 2.26 Not determined 10.15 10-12 1.93 0.84 4.69 13-16 1.08 Not determined 2.03 17-22 0.27 0.41 0.38 23-26 0.25 0.02 0.18 1495 *Value calculated on 14 hour urine (Richelmi et al., 1980). 1496 1497 Comment: In the above case study the patient’s response to Prussian blue was slow to 1498 manifest and this may possibly have been due to her constipation. A very small amount of 1499 thallium was excreted in the faeces, and in fact, a very small amount was excreted overall so 1500 possibly the patient may have improved anyway regardless of therapy. 1501 1502 The efficacies of different therapies were evaluated in 18 cases of thallium poisoning treated 1503 between 1971 and 1979 at the University Hospital of Utrecht, the Netherlands. Patients 1504 were treated with gastric lavage if ingestion had occurred within the preceding 48 hours and 1505 then given 10 g soluble Prussian blue with 100 ml 15% mannitol via a duodenal tube twice 1506 daily. Eight patients were also treated with forced diuresis. Furosemide was given only if 1507 necessary to prevent fluid overload. Sixteen patients survived and two patients with cardio- 1508 vascular insufficiency died. The cases are summarized in the table below. 1509 Patient Sex Age Amount Time Thallium Treated Treated of between concentra with with thallium thallium tion in forced haemope ingested ingestion urine on diuresis rfusion (mg) - & admissio from admissio n (mg/L) patient n (days) history A F 21 500 28 2.1 - - B1) F 22 2400 2 47.4 - - C F 21 350 1 8.8 - - D F 55 1000 4 20.2 - - E M 23 1000 2 71.1 - - F M 28 480 ½ 1.0 - - G F 19 unknown 2 2.0 - - H2) F 24 1000 1 84.0 - - I F 32 1000 2 hours 40.0 - - J M 45 unknown ? 3.0 - - K M 50 750 4 24.6 + - L M 26 875 1 54.0 + -

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M M 21 1500 14 79.8 + - N F 25 1500 1 80.0 + + O3) M 44 unknown ? 8.0 + - P3) M 19 unknown ? 10.0 + - Q F 58 unknown 2 2.2 + - R F 19 3000 1 50.0 + + 1510 1) Patient died after 4 days. 1511 2) The patient survived this suicide attempt. Four months later she died within 6 hours 1512 after ingestion of an unknown amount of thallium 1513 3) These patients were admitted with hair loss of the head 1514 1515 With soluble Prussian blue therapy, the mean half-life of thallium was 3.0 ± 0.7 days. When 1516 administration of Prussian blue was combined with forced diuresis, the mean half-life of 1517 thallium was 2.0 ± 0.3 days (van Kesteren et al., 1980). 1518 1519 An early report on the use of Prussian blue therapy in thallium poisoning reviewed 11 1520 patients hospitalized between 1971 and 1973. There were three men (aged 30 to 44 years), 1521 six women (aged 24 to 64 years) and two children (both female, aged 2 and 6 years). Three 1522 patients received Prussian blue by duodenal tube (2 x 10 g every two days, 2 x 10 g or 20 g 1523 daily). The other nine patients received oral Prussian blue (4 x 5 g daily in the adults, 4 x 1 1524 g daily in the 6-year-old and 4 x 1.7 g daily in the 2-year-old). Four patients treated within 1525 24 hours of ingestion of thallium did not develop any signs of toxicosis. Improvement also 1526 occurred in most of the other patients even though treatment was started late (between 9 and 1527 151 days after exposure). Thallium elimination was primarily via the faeces. Although no 1528 adverse effects were attributed to the Prussian blue therapy, one patient developed 1529 constipation and had constantly high blood thallium concentrations during the first few 1530 weeks of treatment (Stevens et al., 1974). 1531 1532 1533 A 26 year old female student, epileptic and treated with phenobarbital and phenytoin, 1534 ingested approximately 700mg of thallium sulphate when she was in a depressed mood. She 1535 ingested half of the amount during the evening and the remainder the following morning. 1536 She was admitted to hospital twelve hours later and was at this stage asymptomatic. On the 1537 third day hyperaesthesia of the legs and feet developed and she had abdominal discomfort. 1538 The next day (day 4) she developed a prickly, burning sensation in the feet and pain in the 1539 legs and shoulders. No other neurological symptoms could be demonstrated. The pain 1540 subsided during the subsequent days and then a slight polyneuropathy was found. Hair loss 1541 began on the tenth day and progressed but was not extreme. After two weeks only traces of 1542 neurological damage could be demonstrated and this disappeared during the following 1543 weeks. No Mees’ lines were seen on the nails. On initial admission to hospital, no thallium 1544 was evident after gastric lavage but it was shown to be present pharmacologically. She was 1545 treated with Prussian blue 250mg/kg body weight/day (given in 4 daily doses of 3.75g) 1546 along with 15% mannitol through a duodenal tube. Potassium chloride and activated 1547 charcoal were also given on the first two days. A multivitamin preparation was given 1548 intramuscularly 3 times a week and a fluid intake of 3-4 litres/day was prescribed. The 1549 decrease in urinary thallium concentration during the patient’s hospitalization is shown in th 1550 the table below. Prussian blue was stopped on the 13 day when the amount of thallium in 1551 the urine was below 0.5 mg/24hours. 1552 1553

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Daily excretion of thallium in urine Hospitalization day Daily excretion (mg/24 hr) 2 13 6 5.1 8 1.4 13 0.5 1554 (Van der Merwe., 1972-Case study 1) 1555 1556 A twenty two year-old alcoholic ingested approximately 700 mg of thallous sulphate in a 1557 suicide attempt. He was admitted to hospital 6 hours later with no complaints. Gastric 1558 lavage showed only a trace of thallium. He was treated with Prussian blue 250mg/kg body 1559 weight in 15% mannitol via duodenal tube in 4 doses a day. Fluid intake of 3-4 litres a day 1560 was prescribed. After 4 days slight paraesthesia and sensory impairment of the toes could be 1561 demonstrated, however this improved and disappeared during the next few days. Hair loss 1562 started after 2 weeks but was mild. Thallium excretion in the urine is shown in the table th 1563 below. Prussian blue treatment was stopped after the 10 day results were known. 1564 Daily excretion of thallium in urine Hospitalization day Daily excretion (mg/24 hr) 2 3.3 3 3.4 5 1.8 10 0.2 1565 (Van der Meerwe., 1972-Case study 2) 1566 1567

1568 13. Summary of Evaluation 1569 1570 13.1 Indications 1571 Prussian blue is indicated in the treatment of patients with known or suspected internal 1572 contamination with radioactive caesium, radioactive thallium and non-radioactive thallium, 1573 to increase their rates of elimination. Treatment should be started as soon as possible after 1574 exposure. 1575 1576 13.1.1 Thallium 1577 The recommendation for the use of Prussian blue in thallium poisoning is based on evidence 1578 derived from animal studies and limited clinical data from human poisoning in the form of 1579 case reports and case series. 1580 1581 In animals experimentally poisoned with thallium, Prussian blue has been shown 1582 significantly to reduce absorption of thallium, increase its elimination and reduce its 1583 concentration in the brain. Moreover, Prussian blue significantly lowered mortality in 1584 animals poisoned by this metal. 1585 1586 Uncontrolled case reports / series have generally been associated with a favourable outcome, 1587 although there are some cases where patients were slow to respond to treatment and some 1588 patients have been left with long-term sequelae. In many of these cases other treatments 1589 were given concurrently (e.g. activated charcoal) and it is therefore difficult to clearly

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1590 identify the benefits of Prussian blue alone. 1591 1592 Soluble Prussian blue has been shown to more than halve the elimination half-life, when 1593 compared to historical controls, in a large series of thallium poisoned patients. Although 1594 such extensive clinical documentation is not available for insoluble Prussian blue, the overall 1595 evaluation is that insoluble Prussian blue is also effective. 1596 1597 13.1.2 Caesium 1598 The recommendation to use Prussian blue in patients having ingested radioisotopes of 1599 caesium is based on its ability significantly to decrease absorption of, and increase the faecal 1600 excretion of, caesium in animals exposed to caesium-137. The body half-life was halved in 1601 the species tested and reduced content of caesium-134 in various organs was also 1602 documented. In healthy volunteers, insoluble Prussian blue more than halved the body half 1603 life of this radioisotope of caesium. The limited case studies available are consistent with 1604 the effect in healthy volunteers. There appears to be no data on the use of soluble Prussian 1605 blue in this situation. 1606 1607 13.2 Advised routes and dose 1608 Prussian blue should only be given orally. The manufacturers of the pharmaceutical 1609 preparation recommend the following doses (Heyl data sheets Radiogardase and Antidotum 1610 Thallii-Heyl, 2004): 1611 1612 • Adults: In early-presenting cases when thallium or caesium may still remain in the gut an 1613 initial dose of 3g is suggested. In late-presenting cases when thallium or caesium have 1614 already been mostly absorbed, 3 to 20 g per day in divided doses should be given. 1615 1616 The individual dose should be based on the severity of exposure and clinical features. 1617 1618 • Children 2 to 12 years: 1 g orally 3 times a day. 1619 1620 The efficacy and dosing for the paediatric population of insoluble Prussian blue has been 1621 extrapolated from adult data and supported by paediatric patients who were internally 1622 contaminated with cesium-137 and treated with Prussian blue in the Goiânia incident. 1623 Paediatric patients aged 2-4 years are expected to have biliary and gastrointestinal function 1624 that is comparable with a 4-year old (Heyl, 2004). 1625 1626 • Children less than 2 years 1627 The dosing regimen for children less than 2 years old has not been established although it 1628 has been used in this age group (Robb-Smith, 1987). Variations exist in the developmental 1629 maturity of the gastrointestinal and biliary systems of neonates and infants and the dose- 1630 related effects of Prussian blue on an immature gastrointestinal tract are unknown (Heyl, 1631 2004). 1632 1633 Administration 1634 The capsules of Prussian blue can be swallowed whole with liquid or if the patient is unable 1635 to swallow large numbers of capsules they can be opened and dispersed in bland food or 1636 fluid. A suspension of Prussian blue can also be administered via a stomach tube following 1637 gastric lavage. 1638 1639 In many patients with thallium poisoning mannitol (100 ml of 15% solution) has also been

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1640 given with each dose in an effort to prevent constipation. This may not work and other 1641 measure to prevent or treat constipation may need to be undertaken. 1642 1643 End point of therapy 1644 The clinical end-point of Prussian blue therapy is generally considered to be when urinary 1645 thallium levels fall below 0.5mg/day (however this is clearly only a guide as most of the 1646 elimination will be faecal particularly in patients receiving Prussian blue therapy). 1647 1648 1649 13.3 Other consequential or supportive therapy 1650 1651 13.3.1 Caesium 1652 Specialist advice should be sought for the management of radiation accidents. This may require 1653 a multidisciplinary approach with radiation protection and dosimetry professionals, and medical 1654 and nursing staff trained and experienced in managing victims of radiation exposure 1655 (Breitenstein, 2003). 1656 1657 It is essential to prevent further incorporation of any radioactive material. Additional measures 1658 include the administration of laxatives to enhance gastrointestinal transit, antacids for 1659 that become or insoluble in the gastrointestinal tract (and therefore less 1660 absorbable), nasal and/or lung lavage and decontamination of skin and wounds (Gerber & 1661 Thomas, 1992). 1662 1663 In preliminary studies based on animal data, co-administration of Prussian blue with other radio- 1664 eliminators does not affect the efficacy of Prussian blue (Heyl, 2004; Kostail et al., 1983). 1665 1666 13.3.2 Thallium poisoning 1667 The treatment of thallium poisoning is primarily concerned with the prevention of absorption 1668 from the intestinal tract and enhanced elimination from the body. 1669 1670 Gastric lavage should be considered in patients who present early. As patients generally 1671 present to healthcare facilities many hours after exposure it is unlikely that lavage would be 1672 beneficial however gastric decontamination has been has been undertaken as late as 48 hours 1673 after thallium ingestion in some previous cases (van Kesteren et al, 1980). Many patients 1674 will also have vomited spontaneously by the time they attend hospital so as with gastric 1675 lavage, emesis should be considered but may not be of any benefit. Enhanced elimination is 1676 often required in severe cases and haemodialysis (Barckow & Jenss, 1976; Pederson et al., 1677 1978), forced diuresis (Nogué et al., 1982; Heath et al., 1983; de Groot & van Heijst, 1988; 1678 Malbrain et al., 1997) and charcoal haemoperfusion (de Groot et al., 1985; van Kesteren et 1679 al., 1980) have all been used in thallium-poisoned patients. 1680 1681 13.4 Controversial issues and areas of use where there is insufficient information to 1682 make recommendations 1683 1684 13.4.1 Optimal form of Prussian blue 1685 Uncertainty exists in the historical literature regarding which of the two forms of Prussian 1686 blue (soluble or insoluble) is most effective as an antidote for radiocaesium and thallium. 1687 Human case study data is limited and this combined with the lack of analogous animal data 1688 on Prussian blue use (for both thallium and radiocaesium), means that whether the 1689 physicochemical differences between soluble and insoluble Prussian blue have any effect on

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1690 outcomes in human poisoning is currently unknown. A relatively recent literature review by 1691 Thompson & Callen, (2004) highlights these controversies although they do conclude that 1692 that there is sufficient evidence to state that insoluble Prussian blue is effective in 1693 radiocaesium toxicity but that data are inconclusive for thallium. 1694 1695 From a pragmatic point of view, however, preference should be given to insoluble Prussian 1696 blue as this is the only commercially available pharmaceutical preparation. 1697 1698 13.4.2 Optimal dosage regimen 1699 The optimal dose of Prussian blue has not been established in clinical studies. The doses 1700 recommended by the manufacturer are empirical, reflecting the doses that have been used to 1701 treat cases of poisoning with thallium and radioactive caesium. In the case reports of 1702 poisoning with radioactive caesium the doses of Prussian blue used were smaller than those 1703 used for cases of thallium poisoning, typically 3g compared with 20g. Moreover, in one case 1704 series adults given doses of 3g, 6g and 10g per day showed very similar reductions in the 1705 half-life of caesium-137 (Lipsztein et al 1991). Since, however, the half-life of caesium-137 1706 was also found to be influenced by patient body weight and body burden of caesium-137 1707 interpretation of the impact of dosing is difficult. 1708 1709 In the case reports of poisoning with radioactive caesium, treatment was with insoluble 1710 Prussian blue, whereas for thallium poisoning treatment was often with the soluble form. 1711 Whether the form of Prussian blue has an impact on the effective dose for thallium poisoning 1712 is unknown. 1713 1714 Food may increase the effectiveness of insoluble Prussian blue by stimulating bile secretion 1715 and increasing enterohepatic circulation. The increase in enterohepatic circulation may 1716 increase the amount of caesium and thallium in the gastrointestinal lumen and hence 1717 increase the amounts available for binding with Prussian blue (Heyl, 2004). 1718 1719 13.5 Proposals for further studies 1720 In their review Thompson & Callen (2004) concluded that further research is needed to 1721 determine the significance of any differences between the two forms of Prussian blue and 1722 whether their physicochemical differences have any effect on outcomes in human poisoning. 1723 1724 Studies of the effect of Prussian blue in exposures to other radioisotopes may be warranted, 1725 e.g. in the case of rubidium-86. Although animal experiments suggest that Prussian blue may 1726 reduce the whole body retention of rubidium-86, the use of Prussian blue in this situation 1727 should be considered controversial. 1728 1729 13.6 Adverse effects 1730 Severe adverse effects have not been reported with Prussian blue (Hoffman, 2003). Mild to 1731 moderate constipation may occur which can be managed with a high fibre diet or bulk 1732 laxatives. In 46 patients involved in the Goiânia incident treated with Prussian blue, 10 1733 developed constipation (Farina & Brandão-Mello, 1991). It is essential to monitor for and 1734 treat constipation because elimination of thallium and caesium are dependent on the transit 1735 and elimination of Prussian blue from the gut. Faeces will be coloured blue and blue sweat 1736 and tears have been reported with prolonged administration, however this effect appears to 1737 be benign and transient (Hoffman, 2003). If capsules are opened and mixed with food or 1738 fluid, the teeth and mouth may be discoloured blue (Heyl, 2004). 1739

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1740 Hypokalaemia is a potential risk as Prussian blue can bind potassium, however no significant 1741 variations in serum potassium concentrations have been reported, even when large doses 1742 have been given (IAEA, 1988). In 46 patients involved in the Goiânia incident treated with 1743 Prussian blue there were only 3 cases of hypokalaemia (2.5 to 2.9 mmol/L) without clinical 1744 complications. Oral and intravenous potassium supplements resulted in prompt correction of 1745 hypokalaemia (Farina & Brandão-Mello, 1991). 1746 1747 Cyanide toxicity has not been reported from oral dosing with Prussian blue. 1748 1749 1750 13.7 Restrictions for use 1751 None known. Prussian blue has been used in the treatment of thallium poisoning in 1752 pregnancy (Hoffman, 2000). 1753 1754

1755 14. Model Information Sheet 1756 1757 14.1 Uses 1758 Prussian blue (soluble or insoluble) is indicated 1759 • for antidotal therapy in acute or chronic thallium poisoning, 1760 • in the case of incorporation of radioisotopes of caesium (caesium-134, caesium- 1761 137), 1762 1763 14.2 Dosage and route ® ® 1764 Prussian blue is available as Radiogardase -Cs (Radiogardase in the United States of ® 1765 America) and Antidotum Thallii-Heyl distributed by Heyl Chemisch-pharmazeutische 1766 Fabrik GmbH, Berlin, Germany. Both products are available in bottles of 30 hard gelatine 1767 capsules, each containing 0.5 g of insoluble Prussian blue. 1768 1769 Prussian blue should only be given orally. 1770 1771 Adults and adolescents see comments above 1772 The recommended dose of Prussian blue is 3g orally three times/day 1773 1774 When the dose of radioactivity is substantially decreased the dose may be reduced to 1 or 2g 1775 three times/day to improve gastrointestinal tolerance 1776 1777 Paediatric dose (2-12 years) 1778 The recommended dose of Prussian blue is 1g orally 3 times/day 1779 1780 Neonates and infants 1781 Dose has not been established 1782 1783 In patients who cannot tolerate swallowing large numbers of capsules, the capsules may be 1784 mixed with bland food or liquids (this may result in blue discolouration of mouth and teeth). 1785 1786 Radiocaesium contamination: Treatment should be initiated as soon as possible after 1787 contamination is expected. Treatment should continue for a minimum of 30 days and then 1788 the patient should be reassessed for the amount of whole body radioactivity. The duration of

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1789 treatment after exposure will be dictated by the level of contamination. 1790 1791 Thallium contamination: Ideally treatment should be initiated as soon as possible after 1792 exposure however Prussian blue is still thought to be effective after a delay in starting 1793 treatment and should not be withheld. 1794 1795 14.3 Precautions/contraindications 1796 As the antidotal effect of Prussian blue is due to the binding of thallium or caesium in the 1797 gut, it is only effective if the motility of the intestines is intact. In patients in coma or 1798 needing sedation, with reduced intestinal motility, medications to increase intestinal motility 1799 should be considered, although they are not proven effective in this setting. 1800 1801 Prussian blue decreases the duration of radiation exposure but does not treat the 1802 complications of radiation exposure. Supportive treatment for radiation toxicity symptoms 1803 should be given concomitantly with Prussian blue treatment. 1804 1805 In radiological emergencies the type of elemental exposure may not be known, Prussian blue 1806 may not bind to all the radioactive components and these elements may not undergo 1807 enterohepatic circulation which is necessary for Prussian blue binding and elimination. Thus 1808 patients contaminated with unknown or multiple radioactive elements may require additional 1809 treatments. 1810 1811 In severe thallium poisoning additional means for enhancing elimination should be 1812 considered; both charcoal haemoperfusion and haemodialysis have been used, although there 1813 are limited data to suggest that they have an impact on outcome. . 1814 1815 14.4 Pharmaceutical incompatibilities and drug interactions 1816 Prussian blue may bind oral drugs. When given together with oral tetracycline it is 1817 anecdotally reported to decrease the bioavailability of tetracycline. 1818 1819 Prussian blue may bind electrolytes in the gastrointestinal tract and asymptomatic 1820 hypokalaemia has occasionally been reported with its use. 1821 1822 14.5 Adverse effects 1823 Prussian blue is well tolerated; death or serious adverse effects have not been reported with 1824 Prussian blue. Mild to moderate constipation may occur which can be managed with a high 1825 fibre diet or bulk laxatives. It is essential to treat constipation as it will decrease elimination 1826 of thallium and caesium. Faeces will be coloured blue and blue sweat and tears have been 1827 reported with prolonged administration. 1828 1829 14.6 Use in pregnancy and lactation 1830 No contraindications. Prussian blue has been used in pregnant women and since it is not 1831 absorbed from the gastro-intestinal tract, effects on the fetus are not expected. The risk of 1832 toxicity from untreated radioactive caesium or thallium exposure is expected to be greater 1833 than the risk of reproductive toxicity from Prussian blue. 1834 1835 Prussian blue is unlikely to be excreted in breast milk as it is not significantly absorbed from 1836 the gastrointestinal tract, however, women with thallium toxicosis or exposure to 1837 radiocaesium should not breast feed due the risks to the baby from these elements. 1838

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1839 14.7 Storage 0 1840 The pharmaceutical products of Prussian blue should be stored in the dark at 25 C; 0 1841 occasional variations of temperature within the range 15 to 30 C are permitted (Heyl, 2004). 1842 1843

1844 15. References 1845 1846 Atsmon J, Taliansky E, Landau M, Neufeld M (2000) Thallium poisoning in Israel. Am J 1847 Med Sci, 320(5):327-330 1848 1849 Barckow J & Jenss H (1976) [Thallium intoxication treated by haemodialysis, forced 1850 diuresis and antidote] (in German). Med Klin, 71(35): 1377-1382. 1851 1852 Barroso-Moguel R, Villeda-Hernández J, Méndez-Armenta M, Rìos C & Monroy-Noyola A 1853 (1994) Combined D-penicillamine and Prussian blue as antidotal treatment against 1854 thallotoxicosis in rats: evaluation of cerebellar lesions. Toxicology, 89:15-24. 1855 1856 Bhardwaj N, Bhatnagar A, Pathak DP & Singh AK (2006) Dynamic, equilibrium and human 201 1857 studies of absorption of TI by Prussian blue. Health Phys, 90(3): 250-257 1858 1859 BIBRA (1997) Toxicity Profile: Prussian Blue. 1860 1861 Bozorgzadéh A (1971) [Decorporation of radiocaesium by various hexacyanoferrates (II)] 1862 (German). Strahlentherapie, 142(6): 734-738. 1863 1864 Bozorgzadéh A & Catsch A (1972) Evaluation of the effectiveness of colloidal and insoluble 1865 ferrihexacyanoferrates (II) in removing internally deposited radiocaesium. Arch Intern 1866 Pharmacodyn Ther, 197(1): 175-188. 1867 1868 Brandão-Mello CE, Oliveira AR, Valverde NJ, Farina R & Cordeiro JM (1991) Clinical and 1869 hematological aspects of 137Cs: The Goiana radiation accident. Health Phys, 60(1): 31-39. 1870 1871 Breitenstein BD (2003). The medical management of unintentional intakes. Radiat 1872 Prot Dosimetry, 105: 495-497. 1873 1874 Brenot A & Rinaldi R (1967) Toxicite et efficacite comparees de quatre ferrocyanures dans 1875 la decontamination du caesium radioactif 134. Pathol Biol (Paris), 15(1): 55-59. 1876 1877 CDC (2008) Thallium poisoning from eating contaminated cake-Iraq 2008. Morb Mortal 1878 Wkly Rep 57(37):1015-1018 1879 1880 Chan CK, Chan ML, Tse ML, Chan IHS, Cheung RCK, Lam CWK, Lau FL (2009) Life- 1881 threatening Torsades de Pointes resulting from “natural“ cancer treatment. Clin Toxicology 1882 47 (6): 592-594 1883 1884 Chandler HA, Archbold GPR, Gibson JM, O’Callaghan P, Marks JN, Pethybridge RJ 1885 (1990). Excretion of a Toxic Dose of Thallium. Clinical Chemistry, 36(8): 1506-1509 1886 1887 De Backer W, Zachee P, Verpooten GA, Majelyne W, Vanheule A & De-Broe ME (1982)

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1888 Thallium intoxication treated with combined hemoperfusion-. J Toxicol Clin 1889 Toxicol, 19(3): 259-264. 1890 1891 de Groot G (1982) Haemoperfusion in clinical toxicology. PhD Thesis, State University of 1892 Utrecht, The Netherlands. 1893 1894 de Groot G (1985) Thallium concentrations in body fluids and tissues in a fatal case of 1895 thallium poisoning . Veterinary and Human Toxicology, 27:1115-1119. 1896 1897 de Groot G & van Heijst ANP (1988) Toxicokinetic aspects of thallium poisoning. Methods 1898 of treatment by toxin elimination. Sci Total Environ, 71(3): 411-418. 1899 1900 de Groot G, van Heijst ANP, van Kesteren RG & Maes RAA (1985) An evaluation of the 1901 efficacy of charcoal hemoperfusion in the treatment of three cases of acute thallium poiso- 1902 ning. Arch Toxicol, 57: 61-66. 1903 1904 De Wolff JNM, Lenstra JB (1964) The determination of thallium in urine. Pharmaceutisch 1905 Weekblad 99: 377-382. 1906 1907 Díaz V, Tapia R, Brinck G, Gutiérrez M, Hurtado C, del Peso G. [Thallium 1908 poisoning: Prussian blue treatment in 4 cases]. (Spanish) Rev Med Chil. 1990 1909 118(2):183-5. 1910 1911 Dresow B, Nielsen P, Fischer R, Pfau AA & Heinrich HH (1993) In vivo binding of 1912 radiocesium by two forms of prussian blue and by ammonium iron hexacyanoferrate (II). J 1913 Toxicol Clin Toxicol, 31(4): 563-569. 1914 1915 Dresow B, Nielsen P & Heinrich HC (1990) Efficacy of different hexacyanoferrates (II) in 1916 inhibiting the intestinal absorption of radiocaesium. Z Naturforsch, 45(6C): 676-680. 1917 1918 Dutch Pharmacopoeia (1966) 6th Edition. The Hague, Staatsuitgeverij. 1919 1920 Dvořák P (1971) [Binding by hexacyanoferrates (II) of thallium (I)] (German). Z 1921 Naturforsch, 26(4B): 277-281. 1922 1923 Dvořák P (1970) [Hexacyanoferrates (II) as thallium antidotes. Preparation and properties] 1924 (German). Arzneimittelforschung, 20(12): 1886-1888. 1925 1926 Dvořák P (1969) [Colloidal hexacyanoferrates (II) as antidotes in thallium poisoning] 1927 (German). Z Gesamte Exp Med, 151(1): 89-92. 1928 1929 Dvořák P, Günther M, Zorn U & Catsch A (1971) [Metabolic behaviour of colloidal 1930 ferrihexacyanoferrate (II)] (German). Naunyn Schmiedebergs Arch Pharmakol, 269(1): 48- 1931 56 1932 1933 Farina R & Brandão-Mello CE (1991) Medical aspects of 137 Cs decorporation: The Goiana 1934 radiological accident. Health Phys, 60(1): 63-66. 1935 1936 Faustino PJ, Yongsheng Y, Progar JJ, Brownell CR, Sadrieh N, May JC, Leutzinger E, Place 1937 DA, Duffy EP, Houn F, Loewke SA, Mecozzi VJ, Ellison CD, Khan MA, Hussain AS &

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1938 Lyon RC (2008) Quantitative determination of caesium binding to ferric hexacyanoferrate: 1939 Prussian blue. J Pharm Biomed Anal, 47(1): 114-125. 1940 1941 Fekry AE, El-Bieh NM, Elwan KM, Mangood SA (2003) The role of Prussian Blue in 1942 reducing the physiological effects of Cs-137 internal contamination. J Radioanalyt Nuclear 1943 Chem, 257 (1):75-82 1944 1945 Flanagan RJ, Braithwaite RA, Brown SS, Widdop B, de Wolff FA. Basic Analytical 1946 Toxicology, pp222-3. Geneva, World Health Organization, 1995. 1947 1948 Forth W (1986) [How useful is administration of colloidal Berlin blue for the 1949 decontamination of radio-caesium?] (German). Klin Wochenschr, 64(17): 810-812. 1950 1951 Forth W (1983) [Thallium poisoning] (German). Münch Med Wochenschr, 125(3): 45-50. 1952 1953 Forth W & Henning CH (1979) [Thallium intoxication and its therapy] (German). Dtsch 1954 Ärzteblatt, 76(43): 2803-2808. 1955 1956 Franke A, Rodiek S & Neu I (1979) [Thallium poisoning] (German). Notfallmedizin, 5(3): 1957 141-151. 1958 1959 Fred HL, Accad MF.(1997) Abdominal pain, leg weakness, and alopecia in a teenage boy. 1960 Hosp Pract (Minneap). Apr 15;32(4):69-70. 1961 1962 Gansser R (1982) [Acute thallium poisoning after a stay in the Near East] (German). Med 1963 Klin, 77(26): 60-67. 1964 1965 Gerber GB & Thomas RS (1992) Guidebook for the treatment of accidental internal 1966 radionuclide contamination of workers. Radiat Perot Dosimetry, 41: 3-49. 1967 1968 Ghezzi R, Bozza Marrubini M. (1979) Prussian blue in the treatment of thallium 1969 intoxication. Vet Hum Toxicol. 21 (Suppl):64-6. 1970 1971 Giese W & Hantzsch D (1970) [Comparative studies on caesium-137 elimination by various 1972 hexacyanoferrate complexes in the rat] (German). Zbl Vet Med, 185-190. 1973 1974 Giese W, Schanzel H & Hill H (1970) [Biological decontamination of highly radioactive 1975 milk given to pigs. 1. The behaviour of Cs-137 after daily dosage and under specific feeding 1976 conditions] (German). Zbl Vet Med, 11: 191-197. 1977 1978 Günther M (1971) [Influence of colloidal ferrihexacyanoferrate (II) on distribution and 1979 toxicity of thallium] (German). Arch Toxikol, 28(1): 39-45. 1980 1981 Havliček F (1968) Metabolism of radiocesium during gestation and lactation as influenced 1982 by ferric cyanoferrate (II). Int J Appl Radiat Isot, 19: 487-488. 1983 1984 Havliček F (1967) [The effect of ferric cyanoferrate (II) on the action of radiocaesium in 1985 pregnant and lactating rats] (German). Studia Biophysica Berlin, 2(3): 239-246. 1986 1987 Havliček F, Kleisner I, Dvořák P & Pospisil J (1967) [Effects of cyanoferrates on the

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2038 1357-1358. 2039 2040 Kostial K, Kargaćin B, Rabar I, Blanusa M, Maljkovic T, Matkovic V, Ciganovic M, 2041 Simonovic I & Bunarevic A (1981) Simultaneous reduction of radioactive strontium, 2042 caesium and iodine retention by single treatment in rats. Sci Total Environ, 22(1): 1-10. 2043 2044 Kostial K, Kargaćin B & Šimonović I (1983) Efficiency of a composite treatment for mixed 2045 fission products in rats. J Appl Toxicol, 3(6): 291-296. 2046 2047 Kostial K, Vnučec M, Tominac Č & Šimonović I (1980) A method for a simultaneous 2048 decrease of strontium, caesium and iodine retention after oral exposure in rats. Int J Radiat 2049 Biol, 37(3): 347-350. 2050 2051 Kravzov J, Rìos C, Altagracia M, Monroy-Noyola A & Lopez F (1993) Relationship 2052 between physiochemical properties of prussian blue and its efficacy as antidote against 2053 thallium poisoning. J Appl Toxicol, 13: 213-216. 2054 2055 Le Gall B, Taran F, Renault D, Wilk JC & Ansoborlo E (2006) Comparison of Prussian blue 137 2056 and apple-pectin efficacy on Cs decorporation in rats. Biochimie, 88(11): 183718-41. 2057 2058 Lehmann PA & Favari L (1985) Acute thallium intoxication: kinetic study of the 2059 relative efficacy of several antidotal treatments in rats. Arch Toxicol, 57(1): 56-60 2060 2061 Lehmann PA & Favari L (1984) Parameters for the adsorption of thallium ions by activated 2062 charcoal and Prussian blue. J Toxicol Clin Toxicol, 22(4): 331-339. 2063 2064 Leloux MS, Nguyen PL & Claude JR (1990) Experimental studies on thallium toxicity in 2065 rats. II. The influence of several antidotal treatments on the tissue distribution and 2066 elimination of thallium after subacute intoxication. J Toxicol Clin Exp, 10(3): 147-156. 2067 2068 Lipsztein JL, Bertelli L, Melo DR, Azeredo AMGF, Juliao L & Santos MS (1991) 2069 Studies of Cs retention in the human body related to body parameters and Prussian blue 2070 administration. Health Phys, 60(1): 57-61. 2071 2072 Ludi A. (1988) Berliner Blau. Chemie in unserer Zeit; 4:123-127 2073 2074 Ludi A (1983) A comment on "Isotopic exchange in Prussian blue". J Chem Educat, 60: 528. 2075 2076 Ma R, Jin Y, Wang S & Zhou Y (1985) Study of 137-Cs metabolism in humans. In: 2077 Assessment of radioactive contamination in man 1984: proceedings of an International 2078 Symposium on the Assessment of Radioactive Contamination in Man. International Atomic 2079 Energy Agency in cooperation with the World Health Organization. Paris, 19-23 November 2080 1984. Vienna, International Atomic Energy Agency, pp 499-506. 2081 137 2082 Madshus K & Strömme A (1968) Increased excretion of Cs in humans by Prussian Blue. 2083 Z Naturforsch, 23(3B): 391-392. 2084 2085 Madshus K, Strömme A, Bohne F & Nigrović V (1966) Diminution of radiocaesium body- 2086 burden in dogs and human beings by Prussian blue. Int J Rad Biol, 10(5): 519-520. 2087

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2088 Malbrain MLNG, Lambrecht GLY, Zandijk E, Demedts PA, Neels HM, Lambert W, de 2089 Leenheer AP, Lins RL & Daelmans R (1997) Treatment of severe thallium intoxication. J 2090 Toxicol Clin Toxicol, 35 (1):97-100. 2091 2092 Manninen V, Mälkönen M & Skulskii IA (1976) Elimination of thallium in rats as 2093 influenced by Prussian blue and sodium chloride. Acta Pharmacol Toxicol, 39: 256-261. 2094 137 2095 Melo DR, Lipsztein JL, de Olivera CAN & Bertelli L (1994) Cs internal contamination 2096 involving a Brazilian accident, and the efficacy of Prussian blue treatment. Health Phys, 2097 66(3): 245-252. 2098 2099 Meggs WJ, Cahill-Morasco R, Shih RD, Goldfrank LR, Hoffman RS (1997) Effects of 2100 Prussian blue and N- on thallium toxicity in mice. Clin Toxicol, 35(2):163-166 2101 2102 Moffat AC, Osselton MD, Widdop B, Galichet LY (eds) Clarke's Analysis of Drugs and rd 2103 Poisons 3 ed, London, Pharmaceutical Press, 2004 2104 2105 Moore D, House I, Dixon A.(1993) Thallium poisoning. Diagnosis may be elusive but 2106 alopecia is the clue. British Medical Journal. 306(6891):1527-9. Erratum in: BMJ 1993 Jun 2107 12;306(6892);1603. 2108 2109 Müller WH (1969) [Cs137 decorporation using colloidal soluble Berlin blue in rats] 2110 (German). Strahlentherapie, 137(6): 705-707. 2111 2112 Müller WH, Ducousso R, Causse A & Walter C (1974) Long-term treatment of cesium-137 2113 contamination with colloidal and a comparison with insoluble Prussian blue in rats. 2114 Strahlentherapie, 147(3): 319-322. 2115 2116 Niehues R, Horstkotte D, Klein RM, Kühl U, Kutkuhn B, Hort W, Iffland R, 2117 Strauer BE.(1995) [Repeated ingestion with suicidal intent of potentially lethal amounts of 2118 thallium](German) Deutsch Med Wochenschr 24;120(12):403-8 2119 137 2120 Nielsen P, Dresow B & Heinrich HC (1987) In vitro study of Cs sorption by 2121 hexacyanoferrates (II). Z Naturforsch, 42B 1451-1460. 2122 2123 Nielsen P, Dresow B, Fischer R, Gabbe EE, Heinrich HC & Pfau AA (1988a) Intestinal 59 2124 absorption of iron from Fe-labelled hexacyanoferrates (II) in piglets. 2125 Arzneimittelforschung, 38(10): 1469-1471. 2126 2127 Nielsen P, Dresow B, Fischer R Heinrich HC (1991) Inhibition of intestinal absorption of 2128 radiocaesium in humans by hexacyanoferrates (II). Nucl Med Biol, 18(7): 821-826. 2129 2130 Nielsen P, Dresow B, Fischer R & Heinrich HC (1990a) Bioavailability of iron and cyanide 2131 from oral potassium ferric hexacyanoferrate (II) in humans. Arch Toxicol, 64(5): 420-422. 2132 2133 Nielsen P, Fischer R, Heinrich HC & Pfau AA (1988b) Prevention of enteral radiocesium 2134 absorption by hexacyanoferrates (II) in piglets. Experientia, 44(6): 502-504. 2135 2136 Nigrović V (1965) Retention of radiocaesium by the rat as influenced by prussian blue and 2137 other compounds. Phys Med Biol, 10(1): 81-91.

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2138 2139 Nigrović V (1963) Enhancement of the excretion of radiocaesium in rats by ferric 2140 cyanoferrate (II). Int J Rad Biol, 7(3): 307-309. 2141 2142 Nigrović V, Bohne F & Madshus K (1966) [Decorporation of radionuclide: research on 2143 radiocaesium] (German). Strahlentherapie, 130(3): 413-419. 2144 2145 Nogué S, Mas A, Parés A, Nadal P, Bertrán A, Millá J, Carrera M, To J, Pazos MR & 2146 Corbella J (1982) Acute thallium poisoning: An evaluation of different forms of treatment. J 2147 Toxicol Clin Toxicol, 19(10): 1015-1021. 2148 2149 Pai V (1987) Acute thallium poisoning: Prussian blue therapy in 9 cases. West Ind Med J, 2150 36: 256-258 2151 2152 Pau PWI (2000) Management of thallium poisoning. HKMJ, 6(3): 316-319 2153 2154 Pederson RS, Olesen AS, Freund LG, Solgaard P & Larsen E (1978) Thallium intoxication 2155 treated with long-term hemodialysis, forced diuresis and prussian blue. Acta Med Scand, 2156 204(5): 4 2157 2158 Pelclová D, Urban P, Ridzon P, Senholdová Z, Lukás E, Diblík P, Lacina L. (2009)Two- 2159 year follow-up of two patients after severe thallium intoxication. Human & Experimental 2160 Toxicology ;28(5):263-72. 2161 2162 Rangel-Guerra R, Martínez HR, Villarreal HJ.(1990) [Thallium poisoning. Experience with 2163 50 patients] (Spanish). Gac Med Mex.126(6):487-94; 2164 2165 Rauws AG (1974) Thallium pharmacokinetics and its modification by Prussian Blue. 2166 Naunyn Schmiedebergs Arch Pharmacol, 284(3): 294-306. 2167 2168 Rauws AG & van-Heijst AN (1979) Check of Prussian blue for antidotal efficacy in thallium 2169 intoxication. Arch Toxicol, 43(2): 153-154. 2170 2171 Richelmi P, Bono F, Guardia L, Ferrini B & Manzo L (1980) Salivary levels of thallium in 2172 acute human poisoning. Arch Toxicol, 43(4): 321-325. 2173 2174 Richmond CR (1968) Accelerating the turnover of internally deposited radiocesium. In: 2175 Kornberg HA & Norwood WD (eds). Diagnosis and treatment of deposited radionuclides. 2176 Amsterdam, Excerpta Medica Foundation, pp 315-327. 2177 2178 Richmond CR & Bunde DE (1966) Enhancement of cesium-137 excretion by rats 2179 maintained chronically on ferric ferrocyanide. Proc Soc Exp Biol Med, 121(3): 664-670. 2180 2181 Rìos C, Kravsov J, Altagracia M, Lopez-Naranjo F & Monroy A (1991) Efficacy of prussian 2182 blue against thallium poisoning: effect of particle size. Proc West Pharmacol 34: 61-63. 2183 2184 Rìos C & Monroy-Noyola A (1992) D-penicillamine and prussian blue as antidotes against 2185 thallium intoxication in rats. Toxicology, 74: 69-76. 2186 2187 Robb-Smith AHT (1987) Thallium and a pale horse. Lancet, I: 872.

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2188 2189 Sabbioni E, Gregotti C, Edel J, Marafante E, Di Nucci A & Manzo L (1982) Organ/tissue 2190 disposition of thallium in pregnant rats. Arch Toxicol, Suppl 5: 225-230. 2191 2192 Schwartz JG, Stuckey JH, Kunkel SP, Dowd DC, Kagan-Hallet KS.(1988) Poisoning from 2193 thallium. Tex Med. 84(8):46-8. 2194 2195 Spoerke DG, Smolinske SC, Wruk KM & Rumack BH (1986) Infrequently used antidotes: 2196 indications and availability. Vet Hum Toxicol, 28(1): 69-75. 2197 137 86 2198 Stather JW (1972) Influence of Prussian blue on metabolism of Cs and Rb in rats. Health 2199 Phys, 22(1): 1-8. 2200 2201 Stevens W, van Peteghem C, Heyndrickx A & Barbier F (1974) Eleven cases of thallium 2202 intoxication treated with Prussian blue. Int J Clin Pharmacol, 10(1): 1-22. 2203 2204 Stevens WJ.(1978) Thallium intoxication caused by a homoeopathic preparation. Toxicol 2205 Eur Res. 1(5):317-20. 2206 137 2207 Strömme A (1968) Increased excretion of Cs in humans by Prussian blue. In: Kornberg 2208 HA & Norwood WD (eds). Diagnosis and treatment of deposited radionuclides. Amsterdam, 2209 Excerpta Medica Foundation, pp 329-332. 2210 2211 Tang MH, Gong YF, Shen CY, Ye CQ & Wu DC (1988) Measurement of internal 2212 contamination with radioactive caesium released from Chernobyl accident and enhanced 2213 elimination by Prussian blue. J Radiol Prot, 8(1): 25-28. 2214 2215 Thompson DF & Callen ED (2004) Soluble or Insoluble Prussian Blue for Radiocesium and 2216 Thallium Poisoning? Ann Pharmacother, 38: 1509 - 1514. 2217 2218 Thompson DF & Church CO (2001) Prussian Blue for treatment of Radiocesium poisoning. 2219 Pharmacother, 21: (11) – 1364-1367. 2220 2221 Thurgar LD, Singh JM & Thompson MA (2006) Non radioactive caesium toxicity: A case 2222 of treatment using Prussian blue [abstract]. Clin Toxicol, 44: 730-731. 2223 2224 Toohey RE (2003) Internal dose assessment in radiation accidents. Radiat Prot Dosimetry, 105: 2225 329-331. 2226 2227 Trenkwalder P, Bencze K & Lydtin H (1984) [Chronic thallium poisoning. Report of a case 2228 of criminal poisoning] (German). Dtsch Med Wochenschr, 109(41): 1561-1566. 2229 2230 Van Der Merwe CF (1972) The treatment of thallium poisoning: A report of 2 cases. S.A. 2231 Med J, 46: 960-961 2232 2233 Van der Stock J & De Schepper J (1978) The effect of Prussian blue and sodium- 2234 ethylenediaminetetraacetic acid on the faecal and urinary elimination of thallium by the dog. 2235 Res Vet Sci, 25(3): 337-342. 2236 2237 Van Kesteren RG, Rauws AG, de Groot G & van Heijst ANP (1980) Thallium intoxication.

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2238 An evaluation of therapy. Intensivmed, 17: 293-297. 2239 2240 Verzijl JM, Joore JCA, van Dijk A, Glerum JH, Savelkoul TJF, Sangster B & van het Schip 2241 AD (1992) In vitro binding characteristics for caesium of Prussian blue, activated charcoal 2242 and resonium A. J Toxicol Clin Toxicol, 30(2): 215-222. 2243 2244 Verzijl JM, Joore HCA, van Dijk A, Wierckx FCJ, Savelkoul TJF & Glerum JH (1993) In 2245 vitro cyanide release of four Prussian blue salts used for the treatment of caesium 2246 contaminated persons. J Toxicology Clinical Toxicology, 31(4): 553-562. 2247 2248 Villa AF, Poupon J, Cantanese V, El Balkhi S, Nikolova N, Dupont A, Garnier R (2009) 2249 Interest of late and prolonged treatment by Prussian blue in acute thallium poisoning 2250 (abstract). Clinical Toxicology, 47(5): 502 2251 2252 Villanueva E, Hernandez.Cueto C, Lachia E, Rodrigo MD & Ramos V (1990) Poisoning by 2253 thallium. A study of five cases. Drug Safety, 5(5): 384-389. 2254 2255 Vogel AI (1954) A textbook of macro and semimicro qualitative inorganic analysis. 4th 2256 Edition London, Longmans. 2257 2258 Volf V, Doerfel H, Krug H, Prech V, Schuppler U, Sontag W & Stahr B (1987) The effect of 2259 Prussian blue on caesium in man after the Tchernobyl reactor accident. In: Radiation 2260 research. Proceedings of the 8th International Congress of Radiation Research, Edinburgh, 2261 July 1987. EM Fielden (ed). International Congress of Radiation Research (8th 1987 2262 Edinburgh, Scotland). London: Taylor and Francis, abstract B10-7V, p57. 2263 2264 Vrij AA, Cremers HM & Lustermans FA (1995) Successful recovery of a patient with 2265 thallium poisoning. Neth J Med, 47:121-126. 2266 2267 Wainwright AP, Kox WJ, House IM, Henry JA, Heaton R & Seed WA (1988) Clinical 2268 features and therapy of acute thallium poisoning. Q J Med, 69(258): 939-944. 2269 2270 Wolsieffer JR, Stookey GK & Muhler JS (1969) Studies concerning the effect of ferric 137 2271 ferrocyanide, beet pulp, and fluoride upon cesium retention in the rat. Proc Soc Exp Biol 2272 Med, 130(3): 953-956. 2273 2274 Yang Y, Brownell CR, Sadrieh N, May JC, Del Grosso AV, Place D et al (2007) 2275 Quantitative measurement of cyanide released from Prussian blue. Clinical Toxicology, 45: 2276 776-781. 2277 2278 Yang Y, Faustino PJ, Progar JJ, Brownell CR, Sadrieh N, May JC, Leutzinger E, Place DA, 2279 Duffy EP, Yu LX, Khan MA & Lyon RC (2008) Quantitative determination of thallium 2280 binding to ferric hexacyanoferrate: Prussian blue. Int J Pharm, 353(1-2): 187-194. 2281 rd 2282 Walker AW (1998) SAS Trace Element Laboratories. Clinical and analytical handbook, 3 2283 edition. Guildford, Royal Surrey Hospital. 2284 2285

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2286 16. Author(s) Name, Address

2287 Initial draft by ANP van Heijst, A von Dijk & J Ruprecht. 2288 2289 Update and revision by Maeve McParland, Paul Dargan, London 2290 2291 November 2009 2292

2293 17. Additional Information 2294 th 2295 Table 1: Results of EMBASE search 4 November 2009 Number Keywords Results 1 Prussian AND Blue 631 2 Thallium 7299 3 Caesium 959 4 Caesium 2718 5 Radiocesium 392 6 Poisoning OR 193447 Toxicity OR Overdose 7 1 AND 2 47 8 Duplicate filtered: 1 47 unique results AND 2 0 duplicate results 9 6 AND 7 33 10 1 AND 4 15 11 1 AND 5 14 12 1 AND 4 AND 6 1 13 1 AND 5 AND 6 4 14 Rubidium 1098 15 1 AND 15 0 2296 2297 th 2298 Table 2: Results of Medline search 4 November 2009 Number Keywords Results 1 Prussian AND Blue 860 2 Thallium 8122 3 Caesium 1138 4 Caesium 3733 5 Radiocesium 297 6 Poisoning OR 240196 Toxicity OR Overdose 7 1 AND 2 49 Duplicate filtered: 1 49 unique results AND 2 0 duplicate results 8 6 AND 7 32 9 1 AND 4 25 10 1 AND 5 14 11 1 AND 4 AND 6 3

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12 1 AND 5 AND 6 4 14 Rubidium 1935 15 1 AND 14 0

th 2299 Results of Cochrane Library Search 4 November 2009 2300 No results found for Prussian blue using all possible MeSH terms.

52 Table 3: Summary of evidence of use of Prussian blue in metal poisoning Metal data Case reports Animal studies Radiocaesium No controlled trials Prussian blue treatment reduced Administration of a single dose the half-life of caesium in 5 of radiocaesium and Volunteer studies: patients accidentally exposed to concomitant oral dosing of 3 g daily of Prussian blue given 137Cs (Ma et al, 1985) Prussian blue resulted in before 137Cs did not reduce reduction of caesium-uptake caesium absorption. The Goiânia incident Of 249 people from the gastrointestinal tract increase in 137Cs excretion was contaminated with 137Cs, 46 were (Brenot & Rinaldi, 1967; small following 0.5 g of treated with Prussian blue and this Dresow et al., 1990, 1993; Giese Prussian blue three times daily significantly increased the rate of & Hantzch, 1970; Nielsen et al., (Madshus & Strömme, 1968). faecal excretion and reduced the 1988b; Nigrović, 1963; whole body burden retention of Nigrović, 1965). In 6 volunteers it was shown caesium (IAEA, 1988). In 15 of that Prussian blue (4 x 0.5 g or these patients the body burden of The biological half-life was 10 x 0.2 g daily for 2-3 weeks) 137Cs was reduced by an average reduced (Madshus et al., 1966; did not fully block caesium of 71% within 2 months of the Nigrović et al., 1966; Havliček uptake from contaminated food exposure (Melo et al, 1994). et al., 1967; Havliček, 1968; (Volf et al., 1987). Müller et al., 1974; Richmond, Comparing the elimination of 1968; Strömme, 1968). Both forms of Prussian blue 137Cs from the bodies of 18 adults were equally effective in after varying regimens of Prussian The reduced whole-body reducing radiocaesium blue therapy, Lipsztein et al retention of caesium after absorption in 2 male volunteers (1991) found that overall the half- treatment with Prussian blue was who ingested meals labelled life of 137Cs was reduced by 32% seen in individual organs: with a tracer dose of 134Cs, 10 and higher dose rates of Prussian muscle, bone, carcass, liver and minutes after ingestion of 1 g of blue gave a greater reduction. kidney (Bozorgzadéh, 1971; Prussian blue (Dresow et al., Bozorgzadéh & Catsch, 1972; 1993). Chernobyl incident Brenot & Rinaldi, 1967; Kostial Fifteen students were exposed to et al., 1983; Müller et al., 1974; DRAFT for comment - not for distribution

Metal Clinical trial data Case reports Animal studies In volunteer studies with 2 male 134Cs and 137Cs following the Stather, 1972; Wolsieffer et al., adults, the ingestion of Prussian Chernobyl accident. In 3 1969). blue ten minutes before eating a volunteers the biological half-life meal containing 134Cs reduced of caesium ranged from 42 to 71 In piglets soluble and insoluble the caesium absorption more days. Insoluble Prussian blue was Prussian blue reduced the uptake than the simultaneous admini- given from day 114 to 145 post- of 134Cs by more than 97% stration of Prussian blue along exposure and this was reported to (Nielsen et al., 1988b). with the labelled test meal. reduce the half-life of caesium Administration of Prussian blue and enhance elimination (Tang et Prussian blue reduces the body prior to the meal reduced al., 1988). content of 137Cs fed chronically absorption of the radiocaesium to rats (Stather, 1972) to 3-10% of the ingested dose whereas simultaneous ingestion of Prussian blue and the test meal only reduced absorption to 38-63% (Nielsen et al., 1991).

In volunteer studies involving self-dosing by the study authors, ingestion of Prussian blue 180 days after ingestion of 137Cs reduced the biological half-life of caesium from the pre-treatment values of 110 and 115 days to 40 days (Madshus et al., 1966).

In five cases where Prussian blue was given several months after caesium ingestion the

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Metal Clinical trial data Case reports Animal studies biological half-life of caesium was reduced on average to one third of its original half-life (Madshus & Strömme, 1968; Strömme, 1968).

A 37-year-old male was given oral 137Cs followed by Prussian the biological half-life of caesium was reduced from 140 days to approximately 50 days (Richmond, 1968). Non-radiocaesium No controlled clinical trials Prussian blue therapy has been reported to reduce the half-life of non-radioactive caesium in two separate case reports (Chan et al, 2009; Thurgar et al, 2009). Rubidium No human data No Human data Prussian blue reduced the biological half-time of retention of Rubidium-86 in rats (Stather, 1972) Thallium No volunteer studies. A total of 30 published reports of Concomitant oral administration thallium poisoning treated with of thallium and of Prussian blue Bhardwaj et al, 2006 (thallium- Prussian blue were found, in rats resulted in a lower uptake 201) studied the effect of involving 144 cases. These of the metal and lower Prussian blue in 2 patients reports varied in terms of amount concentrations found in organs following 201Tl myocardial of detail provided on treatment. In (Dvořák, 1969; Heydlauf, 1969; scintigraphy. In patient-1 whole addition there were a number of Rauws, 1974). body radioactivity was reduced other variables that makes by 18 and 30% after 24 and 48 comparison difficult. These Reduced retention and increased

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Metal Clinical trial data Case reports Animal studies hours, respectively, of oral reports are tabulated below. excretion of thallium by Prussian blue therapy. Patient- Prussian blue results in a 2 developed constipation and There are single case reports: decrease of the thallium content did not pass any stools after (Atsmon, 2000; Chandler 1990; in liver, kidney, skeleton, blood, oral Prussian blue for 48 hours. DeBacker, 1982; Fred & Accad, heart and muscles (Dvořák, The whole body radiation 1997; Hologittas et al, 1980; 1969; Günther, 1971; Heydlauf, counts were similar to those Malbrain et al; Niehues et al 1969; Kravzov et al., 1993; when Prussian blue was not 1995; Pau, 2000; Pedersen et al Manninen et al., 1976; Rauws, given but there was a 1978; Richelmi, 1980; Rob-Smith, 1974; Rìos et al., 1991; Rìos & concentration of radioactivity in 1987; Schwartz et al 1988; Monroy-Noyola, 1992; Sabbioni the colon suggesting that the Stevens 1978; Villa et al, 2009; et al., 1982). radioactivity was unavailable Vrij, 1995; Wainwright, 1988; for resorption. Rìos & Monroy-Noyola (1992) and multiple case reports: demonstrated that Prussian blue Van Kesteren, (1980) (CDC, 2008; Diaz et al 1990; increased LD50 of thallium by retrospectively assessed the Ghezzi et al, 1979; Kamerbeek et 31%. efficacy of different therapies al 1971; Meggs et al 1994; Moore used in conjunction with et al 1993; Pai, 1987; Pelclova et Meggs et al (1997) found that Prussian blue for thallium al 2009; Rangel-Guerra et al Prussian blue decreased the poisoning in 18 patients. Two 1990; Stevens, 1974; Van mortality from thallium patients died, and for the other Kesteren, 1980; Villanueva, 1990) poisoning in mice 16 mean half-life of thallium was 3.0 ± 0.7 days whilst when The extent and source of thallium In an evaluation of the effect of Prussian blue was combined exposure is often unknown, Prussian blue in thallium-fed with forced diuresis this was diagnosis delayed and hence a rats Lehman & Favari (1985) reduced to 2.0± 0.3 days delay in starting Prussian blue observed that whilst the control therapy. Therapy was started group eliminated only 53% of Kamerbeek et al, (1971) are within 24 hours in only 14 of the the administered thallium dose credited with the first use of above reports (see case summary over the study period, the rats Prussian blue in thallium table below). In other cases there that received Prussian blue

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Metal Clinical trial data Case reports Animal studies poisoning, demonstrating an were varying delays: Villa et al, eliminated 82% of the dose. approximately 7-fold increase (2009) - 42 days; CDC, (2008) - 8 in faecal elimination with its days; Atsmon, (2000) -10-11 Kamerbeek, (Kamerbeek, 1971; use. weeks; Pau, (2000) - 6 weeks; Kamerbeek et al., 1971) showed Vrij, (1995)> 4 months; Chandler that after four days of Prussian A reduction in thallium half-life (1990) - 35 days; Villanueva, blue therapy the concentration of was observed in thallium (1990) > 2 weeks; Wainwright, thallium in the brain of the poisoned patients when (1988) approx. 5 days; Pai, (1987) treated groups was less than half compared with no therapy at all - 4 to 7 days; Rob-Smith, (1987) - that of the control group. The (De Groot & van Heijst, 1988) 5 days. muscle thallium concentration in the treated group was almost Dosing patterns for Prussian blue one-fourth of that of the control varied from case to case (amount and a dose dependent given and duration of treatment) relationship was determined. as did use of concomitant therapy. (Günther, 1971) demonstrated From the limited case studies that soluble Prussian blue Prussian blue appears to be well increased excretion of thallium tolerated and reported to be in rats and reduced the LD50 effective in enhancing the however only if started within excretion of thallium. 24 hours of exposure. Constipation is reported in earlier case studies (Wainwright, 1988; Richelmi, 1980), less so in later cases however many have given mannitol along with the Prussian blue.

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Table 4: Summarized case reports of thallium poisoning (NB - please complete the table where there are "?" if you have access to the full papers) Author No cases Amount thallium Thallium level on Interval Form of PB Dosing Other treatments Outcome presentation between regimen onset of illness and treatment Villa et al 1 NK Urine: 5118 µg/g 42 days Insoluble 6g tds Died (final Tl 2009 creatinine (18g/day) for level 3 µg/g 24 d, second creatinine course for 31d Atsmon et al 1 NK Renal excretion >9 days ? 250 mg/kg/d mannitol Neurological 2000 7mg/24 hours sequelae Pau, 2000 1 Urine at 35d: 8.56 ~42 days Colloidal 4g every 8 Activated charcoal, Persistent µmol/L (NR = soluble hours Succimer, weakness 0.003 µmol/L) (12g/day)

Blood at 42d: 0.15 µmol/L (NR <0.07 µmol/L) Vrij et al 1995 1 NK Urine (24 hr): 4300 ~128 days Colloidal 250 mg/kg per Mannitol, cisapride, Neurological µg/L soluble day for 14 forced diuresis sequelae (NR <10 µg/L) days

Blood: 300 µg/L (NR <10 µg/L) Malbrain et al 1 8.75g thallium Urine: 69,600 µg/L 2 hours Colloidal 3g then 0.5g Emesis & gastric lavage Neurological 1997 sulphate soluble 6xday Mannitol, lactulose sequelae Blood: 5,240 µg/L (3g/day) for Potassium, 20 days haemodialysis

Chandler 1 NK 35 days Colloidal 5g every 6 hrs Plasma exchange, i.v. Neurological 1990 soluble (20g/day) for potassium, sequelae 2 mths Wainwright et 1 NK Urine: 6000 µg/L 3 days Colloidal 5g every 6 hrs Mannitol, forced Neurological

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Author No cases Amount thallium Thallium level on Interval Form of PB Dosing Other treatments Outcome presentation between regimen onset of illness and treatment al 1988 soluble (20g/day) diuresis, haemodialysis, sequelae Blood 5750 µg/L haemofiltration, (estimated faecal diethyldithiocarbamate excretion of Tl 2g over 20d; by forced diuresis 820mg; by dialysis 225mg) Robb-Smith 1 (21 mth NK ? >10 days ? NK, for 3 Potassium Neurological 1987 child) weeks sequelae De Backer et 1 100 mg thallium Blood: 415 µg/L 6 hours Colloidal 250mg/kg Gastric lavage, Recovered al 1982 sulphate soluble (per day?) combined haemoperfusion- haemodialysis for 4 hrs Richelmi et al 1 1 g thallium sulphate Urine: 3000 µg/L 4 days Colloidal 5g every 6 Mannitol Recovered 1980 soluble hours Gastric aspirate: (20g/day) for 10800 µg/L 20 days Fred & Accad 1 ? ? ? ? ? ? ? 1997 Hologgitas et 1 ? ? ? ? ? Kayexelate (sodium died al 180 polystyrene) Niehues et al 1 (2 NK Urine: 9000 µg/L 0.5 hours ? 0.5 mg (sic Gastric 1995 admissions) 2nd adm.: 3700 µg/L NK /day for 6 decontamination days Haemodialysis

2nd adm: aemodialysis, Same dose forced diuresis (1 l urine/h), orthograde intestinal infusions, potassium Nogue et al 1 750 mg thallium Blood: 950 µg/L >1.5 days Not stated 1g 8 hourly Mannitol

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Author No cases Amount thallium Thallium level on Interval Form of PB Dosing Other treatments Outcome presentation between regimen onset of illness and treatment 1982 sulphate (approx) (30g/day) Forced diuresis with days 1-4 and potassium 16-19 Haemodialysis IV and oral Diethyl dithiocarbamate Oral Diphenylthiocarbazone (dithizone)

Pedersen et al 1 2g ? ? ? ? Forced diuresis 1978 haemodialysis Schwartz et al 1 ? ? ? ? ? ? ? 1988 Stevens 1978 1 ? ? ? ? ? ? recovered

CDC 2008 10 (5 NK Blood: 289 µg/L 11 days Insoluble 250 mg/kg/d 2 children died children) (median); range 53 - (Antidotum- before treatment; 1408 µg/L Thallii-Heyl & 2 adults died Radiogardase) (already Urine (24hr): 3063 comatose before µg/L (median); treatment started) range 542 - 12556 µg/L De Groot et al Case 1 NK Blood 2800 µg/L <48 hrs Not stated 10g twice per Gastric lavage Recovered 1985 day Mannitol Case 2 NK Blood 5800 µg/L Forced diuresis Recovered Case 3 NK Blood 1900 µg/L Haemoperfusion Recovered Díaz et al Case 1 NK Urine: 11400 µg/L 5 days Not stated 12 days Magnesium sulphate, Recovered 1990 potassium, furosemide, Case 2 NK Urine: 6300 µg/L 12 days Not stated Potassium Recovered Diuretics

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Author No cases Amount thallium Thallium level on Interval Form of PB Dosing Other treatments Outcome presentation between regimen onset of illness and treatment Case 3 NK Urine: 3500 µg/L 7 days Not stated Magnesium sulphate, Recovered potassium, furosemide, Case 4 15 mg Urine: 3090 µg/L 15 days Not stated Not stated Potassium Recovered Diuretics Note: 6000µg/L thallium level at 45 days after intoxication. No new ingestion of thallium reported.

Villanueva et 4 (2 NK 10yr old: 18400 >14 days ? 250mg/kg 6 Recovery al 1990 children) µg/L hourly

1 adult NK 2400 µg/L 250 mg/kg/d Recovery

Van der Case 1 700 mg thallium 12 hrs ? 250 mg/kg/d Activated charcoal Recovered? Merwe 1972 sulphate (4 x Mannitol 3.75g/day) for Potassium 13 days Multivitamins High fluid intake Case 2 700 mg thallium >6 hours ? 250 Gastric lavage Recovered? sulphate mg/kg/day in Mannitol 4 doses for 10 High fluid intake days Pai 1987 14 4-7 days Colloidal 2g every 8 hrs Magnesium sulphate, 5 died before Case 1 NK Urine 2200 µg/mL soluble (6g/day) for 6 analgesia, diuretics, treatment (sic) weeks vitamin B complex 9 recovered Blood 16 µg/mL (sic) Case 2 Urine 48 µg/mL

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Author No cases Amount thallium Thallium level on Interval Form of PB Dosing Other treatments Outcome presentation between regimen onset of illness and treatment (sic) Blood 4 µg/mL (sic) Case 3 Urine 250 µg/mL (sic) Blood 4 µg/mL (sic) Case 4 Urine 250 µg/mL (sic) Blood 4 µg/mL (sic) Case 5 Urine 660 µg/mL (sic) Blood 6 µg/mL (sic) Case 6 Urine 1100 µg/mL (sic) Blood 11 µg/mL (sic) Case 7 Urine 37 µg/mL (sic) Blood 5 µg/mL (sic) Case 8 Urine 87 µg/mL (sic) Blood 6 µg/mL (sic) Case 9 Urine 23 µg/mL (sic) Blood 4 µg/mL (sic) Van Kesteren 18 (details Urine concentration Colloidal 10g twice per et al 1980 below): (mg/L) soluble day (20g/day) A 500 mg 2.1 28 days B 2.4 g 47.4 2 days Gastric lavage Died C 350 mg 8.8 1 day Gastric lavage D 1 g 20.2 4 days E 1 g 71.1 2 days Gastric lavage

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Author No cases Amount thallium Thallium level on Interval Form of PB Dosing Other treatments Outcome presentation between regimen onset of illness and treatment F 480 mg 1.0 ½ day Gastric lavage G unknown 2.0 2 days Gastric lavage H 1 g 84.0 1 days Gastric lavage Survived but died 6hrs after second overdose with thallium I 1 g 40.0 2 hours Gastric lavage J unknown 3.0 ? K 750 mg 24.6 4 days Forced diuresis L 875 mg 54.0 1 day Gastric lavage, Forced diuresis M 1.5 g 79.8 14 days Forced diuresis N 1.5 g 80.0 1 day Gastric lavage Forced diuresis, haemoperfusion O unknown 8.0 ? Forced diuresis P unknown 10.0 ? Forced diuresis Q unknown 2.2 2 days Gastric lavage Forced diuresis R 3 g 50.0 1 Gastric lavage Forced diuresis, haemoperfusion Stevens et al Case 1 NK Urine 1430 µg/L 30 days Not stated 10g twice per Activated charcoal Recovered with 1974 day every 2 Potassium some muscular days. Duration weakness at 3 not stated mth Case 2 1000mg Not stated Few hours 10g twice per Gastric lavage Recovered day every 2 days for 11 days Case 3 NK Urine: 7100 µg/L 42 days 20g per day Neurological

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Author No cases Amount thallium Thallium level on Interval Form of PB Dosing Other treatments Outcome presentation between regimen onset of illness and treatment for 15 days sequelae Case 4 300 mg Urine elimination 10 hours 5g four times Gastric lavage Recovered 3220 µg/24 hrs per day for 19 mannitol days Case 5 NK Urine elimination 94 days 5g four times mannitol Neurological 120 µg/24 hrs per day. sequelae Duration not stated Case 6 NK Urine 570 µg/L 28 days 5g four times mannitol Recovered per day for >50 days Case 7 NK Urine 232 µg/L 62 days 5g four times mannitol Recovered 2nd admission: urine per day for 18 230 µg/L days

Case 8 NK Not stated 151 days 5g four times Mannitol, Recovered per day for 45 Potassium, antibiotics days Case 9 300 mg Urine elimination 2 hours 5g four times Gastric lavage Recovered 2310 µg/24 hrs per day for 25 Potassium days Case 10 225 mg Urine elimination 9 days 1g four times mannitol Recovered (child 6yrs) 840 µg/24 hrs per day for 35 days Case 11 NK Urine elimination 1 day 1.7g four Lactulose, furosemide Recovered (child 2yrs) 180 µg/24 hrs times Ghezzi & 5 (1 ? ? ? ? ? Dithizone (1 patient) Died Bozza neonate) 4 recovered Marrubini, 1979 Kamerbeek et Case 1 400 mg Not stated Not stated Colloidal ? ? ?

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Author No cases Amount thallium Thallium level on Interval Form of PB Dosing Other treatments Outcome presentation between regimen onset of illness and treatment al 1971 soluble? Case 2 400 mg Not stated Not stated Colloidal soluble? Case 3 2000 mg Not stated 14 days Colloidal soluble? Meggs et al 4 NK Urine: 10837 µg/L 2 days ? 2g tds Multiple dose activated Recovered 1994 and 9569 µg/L (6g/day) charcoal, haemodialysis, analgesia Moore et al 2 NK Blood: 600 µg/L >28 days Colloidal 250 mg/kg/d sequelae 1993 (approx) soluble Pelclova et al Case 1 NK Urine: 8.5 µg/L NK - had Insoluble 6g per day for Neurological 2009 Blood 0.3 µg/L probably be 5 days sequelae poisoned several times over 2 years Case 2 NK Urine: 2800 µg/L 4 weeks Insoluble 6g per day for Neurological Blood: 770 µg/L approx 21 days sequelae Rangel- 50 NK Ranges: Not stated Colloidal 3g/day for 10- Gastric lavage 1 death Guerra et al Urine 100 - 28000 soluble? 14 days Forced diuresis 1990 µg/L 3 patients also received Blood: 100 - 1360 , µg/L 5 patients also received potassium iodide

Case 1 NK Urine: 420 µg/L 4 weeks Colloidal 3 g/day? Forced diuresis, Recovered soluble? painkillers Case 2 NK placental exposure Urine: 60 µg/L 14 days Colloidal Not stated Not stated Neurological (neonate soluble? sequelae from Case 1)

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Author No cases Amount thallium Thallium level on Interval Form of PB Dosing Other treatments Outcome presentation between regimen onset of illness and treatment Case 45 NK Urine: 950 µg/L 2 months Insoluble 3 g/day? Not stated Neurological sequelae

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