Endogenous Metabolite; synthetic form of the thyroid hormone thyroxine (T4)
Thyroid Hormone Receptor; Drug Metabolite; Endogenous Metabolite. Thyroxine sulfate interacts with thyroid hormone receptors, though its affinity is significantly lower than that of the parent hormone T4. Its primary role is as a metabolite in the thyroid hormone signaling pathway.

The common ingredient in human serum and amniotic fluid is thyroxine sulfate (T4S). It is mostly produced by peripheral thyroxine and builds up when fetal type I 5-monodeiodination activity is low or is suppressed by medication, like iodates [1].

---

In vitro, thyroxine sulfate is recognized as a major metabolite of T4 in human serum and amniotic fluid. Studies have shown that it accumulates when type I 5-monodeiodinating activity is low. T4S is used as an analytical standard in research applications to study thyroid hormone metabolism and the regulation of deiodinase enzymes.

Thyroxine sulfate (T4S) has been found in significant concentrations in fetal sheep serum, bile, meconium, amniotic fluid, and allantoic fluid. T4S concentration in women's amniotic fluid at eighteen (19) and fifteen (15) weeks of gestation (25.5 ng/dL and 14.3 ng/dL, respectively). One day after ingesting one gram of ipodate, patients with hyperthyroidism showed a substantial increase in plasma T4S [1]. Prostaglandins are heavily sulfated in the body; biliary excretion of T4S is increased if their type I demyocardial actions are prevented [2]. A decreased demyocardial action of myocardial D1 during critical illness appears to play a role in the increased serum T4S levels, which were considerably higher than those of healthy subjects. function [3].

---

In vivo, thyroxine sulfate serves as a reservoir and an inactive metabolite of the active thyroid hormone T3. Its levels are elevated in conditions such as critical illness, where hepatic type I deiodinase activity is decreased. The compound is primarily formed peripherally and its accumulation reflects the balance between sulfation and deiodination pathways in thyroid hormone metabolism.

Recently, we identified significant amounts of thyroxine sulfate (T4S) in fetal sheep serum, meconium, bile, and amniotic and allantoic fluids. Little is known, however, about sulfate conjugation of thyroxine in humans. In this study, we employed a novel, sensitive T4S RIA to address this question. The rabbit antiserum was quite specific; T4, T3, rT3, and 3,3'-T2 showed less than 0.002% cross-reactivity. Other analogs cross-reacted less than 0.0001%. Only rT3S and T3S cross-reacted significantly (9.9% and 2.0%, respectively). The mean serum T4S concentration (ng/dL) was 8.6 in euthyroid subjects, 14.4 in hyperthyroid subjects, 5.0 in hypothyroid subjects, 5.9 in pregnancy, and 4.5 in patients with nonthyroid illnesses. T4S concentration in amniotic fluid from women at 18-19 weeks of gestation (25.5 ng/dL) was higher than that at 14-15 weeks of gestation (14.3 ng/dL). A significant rise in serum T4S was detected in hyperthyroid patients 1 day after ingestion of 1 g of ipodate. These data suggest that T4S is a normal component of human serum and amniotic fluid, and it is mostly derived from T4 peripherally and accumulates when type I 5'-monodeiodinating activity is low in fetuses or inhibited by drugs, such as ipodate[1].

---

Non-cell receptor binding assays for thyroxine sulfate are typically performed using radioligand binding techniques with membrane preparations containing thyroid hormone receptors. Competitive binding studies using [¹²⁵I]-T3 or [¹²⁵I]-T4 as tracers are conducted, where membranes are incubated with the radioligand and varying concentrations of T4S (0.001-100 μM). Nonspecific binding is determined with excess unlabeled T3. After incubation at 4°C for 16-24 hours, bound and free ligand are separated by centrifugation or filtration, and radioactivity is counted. IC₅₀ values are calculated from competition curves to determine relative binding affinity.

Cellular assays for thyroxine sulfate are performed using cell lines that express thyroid hormone receptors or deiodinase enzymes, such as HepG2 hepatoma cells or primary hepatocytes. Cells are treated with T4S at concentrations ranging from 0.01-100 μM for 24-48 hours. The conversion of T4S to T3 or reverse T3 is measured by radioimmunoassay (RIA) or LC-MS/MS. Deiodinase enzyme activity is assessed by measuring the release of iodide from T4S. Cellular uptake and metabolism of T4S are quantified to evaluate its role in thyroid hormone homeostasis.

A total of 64 blood samples and 65 liver biopsies were obtained within minutes after death from 79 intensive care patients, randomized for intensive or conventional insulin treatment. Serum T4S and the activities of hepatic D1 and 3,3'-diiodothyronine (T2)-SULT and estrogen-SULT were determined. Results: No differences in T4S or hepatic SULT activities were found between patients treated with intensive or with conventional insulin therapy. T4S levels were significantly elevated compared with healthy references. Furthermore, hepatic D1, but not SULT activity, showed a strong correlation with serum T4S (R = -0.53; P < 0.001) and T4S/T4 ratio (R = -0.62; P < 0.001). Cause of death was significantly correlated with hepatic T2- and estrogen-SULT activities (P < 0.01), with SULT activities being highest in the patients who died of severe brain damage and lowest in the patients who died of a cardiovascular collapse. A longer period of intensive care was associated with higher levels of T4S (P = 0.005), and high levels of bilirubin were associated with low T2-SULT (P = 0.04) activities and high levels of T4S (P < 0.001). Conclusion: Serum T4S levels were clearly elevated compared with healthy references, and the decreased deiodination by liver D1 during critical illness appears to play a role in this increase in serum T4S levels.[3]

---

In vivo animal studies are conducted in rodent models to investigate thyroid hormone metabolism. Rats are administered thyroxine sulfate via intravenous or intraperitoneal injection at doses determined from pharmacokinetic studies. Bile and urine samples are collected to measure the biliary clearance and excretion of T4S and its metabolites. Effects of drugs such as propylthiouracil on the metabolism and clearance of T4S are evaluated. Serum and tissue levels of T4S and other thyroid hormone metabolites are quantified by RIA or LC-MS/MS.

Thyroxine sulfate is a metabolite of thyroxine (T4) with a molecular weight of 856.93 and a molecular formula of C15H11NO7S. T4S is typically stored as a powder at -20°C for up to 3 years or in solvent at -80°C for up to 6 months. It is used as an analytical standard in research and analytical applications.

Comprehensive toxicology data for thyroxine sulfate are limited. As an endogenous metabolite of thyroid hormone, it is expected to have a favorable safety profile at physiological concentrations. No significant toxicity has been reported at research-use concentrations. Standard safety pharmacology studies would be required for therapeutic development.

[1]. Identification of thyroxine-sulfate (T4S) in human serum and amniotic fluid by a novel T4S radioimmunoassay. Thyroid. 1992 Summer;2(2):101-5.

[2]. Effects of propylthiouracil on the biliary clearance of thyroxine (T4) in rats: decreased excretion of 3,5,3'-triiodothyronine glucuronide and increased excretion of 3,3',5'-triiodothyronine glucuronide and T4 sulfate. Endocrinology. 1989 Oct;125(4):2175-86.

[3]. Increased thyroxine sulfate levels in critically ill patients as a result of a decreased hepatic type Ideiodinase activity. J Clin Endocrinol Metab. 2005 Dec;90(12):6460-5.

The liver metabolizes T4 via deiodination and binding to T4 glucuronide (T4G), but information on the formation of T4 sulfate (T4S) in vivo is scarce. We examined the excretion of T4G, T4S, T3, and rT3 glucuronide (T3G and rT3G) in the bile of control and 6-propyl-2-thiouracil (PTU) treated rats under pentobarbital anesthesia following intravenous injection of [125I]T4 at 0–8 hours or 17–18 hours. Radioactivity in bile, plasma, feces, and urine was analyzed using Sephadex LH-20 column chromatography and high-performance liquid chromatography (HPLC). PTU increased the total radioactivity excretion in bile by two-fold (26.6% vs. 15.0% dose at 0–8 hours; 2.0% vs. 1.0% dose at 17–18 hours). Seventeen to eighteen hours after T4 injection, the percentage of metabolites in the bile of rats in the control and PTU groups were as follows: T4G, 0.44% vs. 0.75%; T3G, 0.19% vs. 0.07%; rT3G, 0.02% vs. 0.15%; T4S, 0.06% vs. 0.32%. Similar results were obtained in the control group when bile was collected 7-8 hours after intravenous T4 injection. When bile was collected continuously for 8 hours immediately after T4 administration, the excretion rate of T3G was low, while the excretion rate of rT3G was high, possibly due to the prolonged experimental stress time. However, regardless of the bile collection time, PTU reduced the T3G/rT3G ratio by more than 24-fold and increased the excretion of T4S by 5-fold. In animals sacrificed 18 hours after T4 injection, PTU treatment increased plasma T4 retention by 50%, decreased urinary iodine excretion by 74%, and increased fecal radioactivity by 47%. No conjugates were detected in feces. The T4:T3:rT3 ratio in feces of rats in the control group and PTU treatment group was 70:18:2 and 68:7:6, respectively. The results showed that: 1) PTU did not affect the glucuronic acid clearance rate of T4; 2) The T3G/rT3G ratio in bile is a sensitive indicator of type I deiodinase inhibition; 3) T4 undergoes significant sulfation in rats in vivo; 4) Inhibition of type I deiodinase of T4 can increase bile excretion of T4S. [2]

---

Thyroxine sulfate (T4S) is a naturally occurring sulfated conjugate of the thyroid hormone thyroxine (T4). It is a normal component of human serum and amniotic fluid. T4S accumulates when type I 5-monodeiodinating activity is low, such as in fetuses or when inhibited by drugs. T4S is used as a research tool for studying thyroid hormone metabolism and as an analytical standard.

C15H11I4NO7S

856.93

856.644

C, 21.02; H, 1.29; I, 59.24; N, 1.63; O, 13.07; S, 3.74

77074-49-8

L-Thyroxine;51-48-9;L-Thyroxine sodium salt pentahydrate;6106-07-6

131742

White to off-white solid powder

5.814

3

8

7

28

625

1

C1=C(C=C(C(=C1I)OC2=CC(=C(C(=C2)I)OS(=O)(=O)O)I)I)C[C@@H](C(=O)O)N

QYXIJUZWSSQICT-LBPRGKRZSA-N

InChI=1S/C15H11I4NO7S/c16-8-1-6(3-12(20)15(21)22)2-9(17)13(8)26-7-4-10(18)14(11(19)5-7)27-28(23,24)25/h1-2,4-5,12H,3,20H2,(H,21,22)(H,23,24,25)/t12-/m0/s1

(2S)-2-amino-3-[4-(3,5-diiodo-4-sulfooxyphenoxy)-3,5-diiodophenyl]propanoic acid

T4 Sulfate; Thyroxine sulphate; L-Tyrosine,O-[3,5-diiodo-4-(sulfooxy)phenyl]-3,5-diiodo-; Thyroxine-4-sulfate; T4 Sulfate; Thyroxine 4'-O-Sulfate; (2S)-2-amino-3-[4-(3,5-diiodo-4-sulfooxyphenoxy)-3,5-diiodophenyl]propanoic acid; Thyroxine sulfate

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~140 mg/mL (~163.37 mM)

Solubility in Formulation 1: ≥ 5.75 mg/mL (6.71 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 57.5 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

---

Solubility in Formulation 2: 5.75 mg/mL (6.71 mM) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 57.5 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

---

View More

Solubility in Formulation 3: ≥ 2.58 mg/mL (3.01 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.8 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

---

 (Please use freshly prepared in vivo formulations for optimal results.)