Excerpt: In this Guest Post, Dr. X (name withheld by request), Research Scientist and Proponent of Natural Desiccated Thyroid, discusses stability problems with the current test methods of T4 measurement. He presents an original and improved test method which yields stable test results. He also presents some interesting information about bio-availability of the major USA prescription porcine thyroid product’s new formulation. Finally he discusses his results after testing Thyroid-S using this improved test method. Dr. X has two Doctorate Degrees: Theoretical Physics, and Medicinal Chemistry. We would like to thank Dr. X for his significant contribution to our understanding of the problem, as well as providing the innovative solution.
Analysis of T4 thyroid hormone has been problematic mainly due to the fact that commercially available T4 internal standards are deuterated (that is, combined with hydrogen-2, or heavy hydrogen, a hydrogen ion with an extra neutron) in positions that exchange with hydrogen-1 (that is, normal hydrogen) immediately upon contact with water. Typically, the exchange occurs because water, even at 18 mega-ohm resistance is partially acidic, displacing the unstable hydrogen-2 (due to expanded valence electron orbits) until an equilibrium occurs, usually at about 30% total exchange, at which point the exchange occurs more slowly, over the course of about 24 hours, until the deuterated internal standard has diminished to zero.
Clinical applications measuring T4 in blood serum make the exchange worse, since acids and bases are used in Strong Cation and Strong Anion Exchange Resins. Methanol extractions (protein crashing) causes the same exchange, acetonitrile that is not completely water free is also problematic.
In laymen’s terms, essentially, the extra neutron in the deuterated T4 standard gives it a molecular mass slightly different than the sample T4 to be measured (which is non-deuterated). The sensitive test equipment is able to differentiate and measure the concentrations of two different types of molecules. Since the concentration of the deuterated T4 standard is known, we measure the concentrations of both molecules, and then we can calculate the unknown concentration of the sample T4. The exchanging of hydrogen-2 with hydrogen-1 explained above causes the deuterated hydrogen-2 labeled T4 to appear as normal T4 during the measurement. This has the effect of both diminishing the reference standard and heightening the unknown sample standard, which will result in largely inaccurate, and time variant, results.
T4 typically comes up looking as though it is at ‘normal’ levels, artificially heightened because the internal standard has diminished, and the 2nd order standard curve is adjusted by QC standards that are not isotopically labeled. Thus, symptomatic patients invariably have artificially looking ‘normal’ T4 levels that have been elevated by non-isotopically labeled QC standards in a standard curve against a constantly diminishing deuterium labeled internal standard that is exchanging hydrogen-1 during the entire analysis.
The process of using a heavier atom such as hydrogen-2 in a molecule so that the molecule can be differentiated and measured is called “labeling” by scientists, and is a common technique in analytical chemistry. In this experiment, carbon-13 labeled T4 was used so that hydrogen exchange is not possible. In other words, rather than use hydrogen-2 as the heavy atom to label the reference standard molecule T4, carbon-13, which is a heavy carbon with one extra neutron, is used. The concentrations of the two different molecules can still be differentiated and measured, but the problem of hydrogen exchange in the previous method is effectively eliminated.
Fig. 1 – Molecular Structure of Carbon-13 Labeled T4
THYROID-S (Sriprasit Pharma Co.) brand was chosen because recently the major USA prescription porcine thyroid product has changed their binding agent from sugar (because it requires frequent QC re-analysis because it is a growth media) to inert cellulose. Unfortunately, cellulose is indigestible, and the new formulation works only if the patient chews the tablet and allows sufficient time for the cellulose to be broken down by saliva enzymes. Consequently, many of these patients, since the formulation change, have complained of a sudden, ‘unexplained’ recurrence of hypothyroid symptoms.
T4 Chemical Information:
Formula Weight (HCI Salt) =819.27
Molecular Formula =C15H11I4N04.HCI
Molecular Weight (Free Base) =782.81
Molecular Formula =C15H11I4N04
The manufacturer’s claim is that the 60mg desiccated pig thyroid product has a T4 content in the 30-40mcg range. Since pig thyroid T1-T3 are invariably in balance, there is no need to measure each of the T l , T2, and T3 components, which are known to be proportional to the T4 content.
Procedure:
The procedure was repeated three times for duplication.
Carbon-13C6 isotopically labeled T4 was weighed out and diluted in methanol, in this case, to 169ug/mL.
1 60mg THYROID-S tablet was mortared and digested in 1 mL concentrated sodium hydroxide overnight (releasing T4 from all biological components) followed by sonication for 1 hour.
1 mL of each component, the labeled T4 and the digested tablet, were diluted into 10 mL methanol and passed through a 0.2 micron filter.
This solution was directly infused into a Micromass TQD in Electrospray Positive Ion Mode (Positive Ion Mode is appropriate following a caustic digestion which reduces all porcine components to sub-background levels). Several hundred continuum spectra were collected and averaged together.
Results:
The T4 content of this particular lot, is approximately 39 microgram per tablet, well within tolerance of the manufacturer’s claim for a natural product The Tl, T2, T3 components will therefore be proportionally correct.
Conclusions:
The THYROID-S product will not have the indigestible issues associated with cellulose because it is not formulated as such. In all, the product meets standards or potency, and is a superior product because of formulation bio-availability.
Laboratories interested in Dr. X’s method can contact him at: 