Labeling of Iodine

Principles

  1. The oxidation state of iodine is essential for its ability to bind to other compounds or molecules.
  2. States of Iodine
    1. Oxidized state (I+) in which it is looking for an electron. Considered unstable and will not be found by itself, but in attached to or part of a molecule. It is know to be electrophilic
    2. Reduced iodine (I-) can is considered the stable form of iodine and can be found by itself. Because it is negative it is trying to share with another element, hence it is nucleophilic
  3. These two forms of iodine are found in an aqueous solution
  4. As stated I+ cannot be found as a free species, and forms nucleophilic complexes (as an example, with water).
I2 + H2O H2OI+ + I-
I2 + OH-   >HOI + I-
  1. Electrophilic substitution occurs with the iodonium ion and the above molecules. Consider the terminology
    1. The iodonium is electrophilic and the H-O is nucleophilic
    2. The iodine is looking for an electron while the H-O want to share its available electron space

Electrophilic substitution

                        R–H + H2O 131I+            R–131I + HI + H2O

The above, first example is electrophilic substitution

Nucleophilic substitution*

                                    R–I + Na131I           R–131 I + NaI + H2O

The above, second example is nucleophilic substitution

  1. Electrophilic substitution of the hydrogen ion occurs with the iodonium ion (1st example) or by nucleophilic substitution (2nd example) where the radioactive iodine is exchanged with the stable iodine.
  2. In protein iodination the following are important:  pH 7 to 9, temperature, and duration of the iodinated process.  Degree of iodination can affects the integrity of the protein and alter its composition.  Usually a one for one exchange occurs

Types of iodination

  1. Triiodinde method – Used with organ compound labeling and has only a 10 to 30 percent yield.  In addition stable iodine is present further reduces the quality of the radioactive purity
  2. Iodine monchloride method (ICl) – Radioiodine and stable iodine (ICl) is added to a HCl solution and labeling occurs at a specific pH and temperature.  Fifty to 80 percent binding is common, but the presence of stable iodine tends to lower desired yield
  3. Chloramine-T Method – Chloramine-T and  radioiodine results in oxidation that causes the binding of the radioiodine.  Because of the presence of only radioactive iodine is in solution there is a 90% yield.  Denaturing of proteins can occur in this process, so the final compound must thoroughly tested.  This process is currently used for compounding
  4. Electrolytic method – Requires radioiodine and the compound to be labeled in a mixed solution.  Products are in a electrolytic cell, where there are anode and cathode compartments separated by a dialyzing bag. Additionally, the cathode is immersed in saline and the anode compartment contains the compounding agents.  Electricity is applied releasing the iodine which then labels to the compound.  This yields about an 80%.
  5. Enzymatic method – Used to label proteins and hormones.  Lactoperoxidase, chloroperoxidase, and a nanomolar amount of H2O2 are mixed.  H2O2 oxidizes the radioiodine that results in compounding.  This method can denature proteins and other organic compounds.  Yields 60 to 80 percent.
  6. Conjugation method – Similar to the chloranime-T method and usually has a low yield without denaturing of proteins.
  7. Demetallation method – carbon-metal bond is cleaved by radioiodination in the presence of an oxidizing agent (chloramines-T and iodogen).  Succinimidyl para-tri-n-butylstannyl (SBSB) is the agent of choice for oxidizing.  The tributyl stannyl group substituted with radioiodine.
  8. Iodegen method – First iodogen or choraminde is dissolved in methylene chloride which evaporates in collecting tubes.  The Radioiodine and protein to be labeled is then added. Iodegen oxidizes the iodine and labeling occurs. The remaining solution that has the unlabeled radioiodine is then removed.  Denature of protein is minimal and the labeling yield is between 70 to 80 percent.
  9. Iodo-bead method – Used to tag with different proteins and peptides.  Nonporous polystyrene spheres (2.5 mm) that contain on its surface an oxidant compound, N-chlorobenzensulfonamide.  Four to five beads are added to a mixture of radioiodine with a protein or a peptides.  Oxidation occurs at room temperature and takes about 15-minutes.  The radio-labeled solution is removed and will contain about a 90% yield.  Little denaturing occurs.

Radioiodine compound

  1. After the labeling process any excess material is removed in order to reduce the free radioiodine from the labeled compound.
  2. Sterility is essential, but autoclaving will denature protein and organ compounds.
  3. Iodine best tags to aromatic compounds because the tag is not reversible.  Other types of compounds that can revert back include:  aliphatic compounds, amino and sulfydryl groups, and others.
  4. Double bonds using radioiodine is not recommended because it usually alters the chemical structure resulting in a change of the physiological uptake.
  5. Consider which iodines are best for diagnostic and therapeutic analysis of the human system.

Definitions

*Nucleophilic substitution is a reaction initiated by a species that is electron rich, but has an unshared pair of electrons. It will then replace another element by substituting the unshared pair of electrons and departs with the new element 

*Electrophilic substitution is the main reaction and is looking for electrons. This type is exhibited by aromatic compounds in which an electrophil attacks an aromatic ring (acquiring the electron pair) and replaces the hydrogen bonds.

Here is a chapter article in PDF format that goes into detail about nucleophilic and electrophilic reactions

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