Drug compounds have included stable heavy isotopes of carbon, hydrogen, and other elements, mostly as quantitative tracers while the drugs were being developed. Because deuteration may have an effect on a drug's pharmacokinetics and metabolic properties, it is a cause for concern [1].

Absorption, Distribution and Excretion
Norethindrone is absorbed from the gastrointestinal tract, metabolized in the liver, and excreted in urine and feces as glucuronide and sulfate conjugates. (14) C-norethindrone was administered to seven subjects, and 43% of the dose was excreted in urine within 5 days; the biological half-life of the radioactivity is 24 hours. Enzymatic hydrolysis releases only 32% of the urinary radioactivity, with another 25% excreted as sulfate conjugates. The metabolites excreted in urine are much less polar than those excreted after administration of the related compounds norethindrone or norethindrone alcohol. The 3αOH,5β and 3βOH,5β isomers of tetrahydronorethindrone (13β-ethyl-17α-ethynyl-5β-gonan-3α,17β-diol) were isolated from urine and identified by mass spectrometry, thin-layer chromatography, and gas-liquid chromatography. Plasma radioactivity decreased more rapidly after administration of norethindrone than after administration of norethindrone. Approximately 2% of the administered dose was converted to acidic compounds. No significant differences were observed in radioactive excretion rates or metabolites after oral or intravenous administration of norethindrone. The binding of different synthetic steroids (used for hormonal contraception) to SHBG was investigated by measuring their ability to displace tritium-labeled testosterone from sex hormone-binding globulin (SHBG) in a competitive protein-binding system. Only 19-nortestosterone derivatives exhibited a significant ability to displace testosterone from SHBG, with dextrogestrol acetate (d-Ng) showing the strongest displacement capacity. In women with previously stable plasma d-Ng levels, increasing SHBG levels resulted in a 2- to 6-fold increase in SHBG levels. This leads to the conclusion that SHBG is the primary carrier protein of d-Ng. The potent testosterone displacement activity of d-Ng may also explain the androgenic side effects observed in oral contraceptives containing d-Ng.

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Metabolism / Metabolites
Seven subjects were given (14) C-norethindrone, and 43% of the dose was excreted in the urine over 5 days…enzymatic hydrolysis released only 32% of the urinary radioactivity, with another 25% excreted as sulfate conjugates. The urinary metabolites were far less polar than those excreted after administration of the related compound norethindrone or acetylene. The 3αOH,5β and 3βOH,5β isomers of tetrahydronorethindrone (13β-ethyl-17α-ethynyl-5β-gonan-3α,17β-diol) were isolated from the urine and identified by mass spectrometry, thin-layer chromatography, and gas-liquid chromatography. The rate of decrease in plasma radioactivity was faster than after administration of norethindrone or acetylene. Approximately 2% of the administered dose was converted to acidic compounds. There was no significant difference in the excretion rate of radioactive substances and their metabolites after oral or intravenous administration of norethindrone. This study compared the metabolism of dl-, d-, and l-norethindrone in African green monkeys (Cercopithecus aethiops). Following a single oral administration of 14C-dl-norethindrone (1 mg/kg), the total urinary excretion of 14C (51.4 ± 5.0%) was significantly higher than that of the d-enantiomer (37.5 ± 5.4%), but not significantly different from the l-enantiomer (44.2 ± 8.9%). In all cases, the radioactive material in urine was predominantly in free form (48–62%), with an additional 13–27% released by β-glucuronidase preparations. No sulfate conjugates were detected. At least one major metabolic pathway (16β-hydroxylation) and one minor metabolic pathway (16α-hydroxylation) exhibit stereoselectivity, meaning they are effective for the I-enantiomer but ineffective for the D-enantiomer. Three metabolites, namely 16β-hydroxynorethindrone, 16α-hydroxynorethindrone, and 16-hydroxytetrahydronorethindrone (considered to be the 16β form), were detected only in urine samples from animals administered 14Cd- and l-norethindrone. Following administration of 14Cd-norethindrone, 3α,5β-tetrahydronorethindrone was found to be the major urinary metabolite. These observations were compared with previously reported results regarding the metabolism of dl-norethindrone in female urine. The in vitro metabolism of norethindrone stereoisomers (d, l, and a racemic mixture dl) by rabbit liver microsomal fractions was investigated. The bioactive levonorgestrel was metabolized more rapidly than the inactive dextro-norethindrone. This is primarily due to the more readily convertible levonorgestrel to the A-ring reducing metabolite. There was no difference in the degree of hydroxylation between the two isomers; after 30 minutes of incubation, approximately 40% of each isomer was converted to the hydroxylated metabolite. However, differences exist between the two isomers: levonorgestrel is primarily converted to 16β-hydroxysteroids, while dextronorgestrel is primarily converted to 16α-hydroxysteroids. The amount of hydroxylation at the C-6 position is similar in both isomers. The metabolism of the racemic mixture falls between that of the levonorgestrel and dextronorgestrel isomers. The rates of in vitro metabolism of 19-nortestosterone-derived synthetic progestins in rabbit liver tissue were compared. Within 1 hour, the metabolic rate of norethindrone was comparable to that of 19-nortestosterone, while the metabolic rates of dextronorgestrel and norethindrone were slightly slower. The metabolic rate of levonorgestrel was less than 5%. In all cases, the reaction product was a tetrahydrosteroid. Norethindrone is metabolized via norethindrone. Skeletal muscle, lung, and small intestine also metabolize norethindrone and dextronorgestrel, but at a slower rate than liver tissue. Adipose tissue metabolizes small amounts of norethindrone, but the heart and spleen do not metabolize it. In any of the extrahepatic tissues studied, neither norethindrone nor levonorgestrel was metabolized. The metabolism of three steroids used in oral contraceptives (OCs) in a small amount of human jejunal mucosal tissue was studied in vitro. This study was conducted because the human gastrointestinal mucosa is known to metabolize a variety of drugs. After incubation, approximately 40% of ethinylestradiol, 9.8% of levonorgestrel, and 7% of ethinylestradiol were metabolized. All of these metabolic responses were significantly different from the control group. The results indicate that ethinylestradiol metabolism is related to the weight of the tissue used. These results are consistent with the known significant first-pass effect of ethinylestradiol. Norethindrone, which is known to have a small or no first-pass effect, has a low intestinal metabolic rate. Under the experimental conditions used, neither ethinylestradiol nor levonorgestrel exhibited significant phase I metabolism.

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Biological Half-Life
Seven subjects were administered (14) C-norethindrone, and 43% of the dose was excreted in urine within 5 days; the biological half-life of the radioactive material is 24 hours.

Interactions
Concomitant use of substances known to induce drug-metabolizing enzymes (especially cytochrome P450 enzymes), such as anticonvulsants (e.g., phenobarbital, phenytoin, carbamazepine) and anti-infectives (e.g., rifampin, rifabutin, nevirapine, efavirenz), may increase estrogen and progesterone metabolism. Ritonavir and nelfinavir, although potent inhibitors, exhibit induction when used concomitantly with corticosteroids. Herbal preparations containing *Hypericum perforatum* may induce estrogen and progesterone metabolism. Phenytoin and rifampin increase serum sex hormone-binding globulin (SHBG) concentrations; this significantly reduces serum free drug concentrations of certain progestins, a particular concern for patients using progestin for contraception. There are currently no data on drug interactions with rifabutin, but due to its structural similarity to rifampin, similar precautions may be necessary when used concomitantly with progestins.

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Non-Human Toxicity Values
Rats: Oral LD50: 5010 mg/kg; Rats: Intraperitoneal LD50: 11200 mg/kg; Mice: Intraperitoneal LD50: 7300 mg/kg; Mice: Oral LD50: 5010 mg/kg

[1]. Impact of Deuterium Substitution on the Pharmacokinetics of Pharmaceuticals. Ann Pharmacother. 2019;53(2):211-216.

[2]. Oral progestogens vs levonorgestrel-releasing intrauterine system for endometrial hyperplasia: a systematic review and metaanalysis. Am J Obstet Gynecol, 2010. 203(6): p. 547 e1-10.

Norgestrel may cause developmental toxicity depending on state or federal labeling requirements. Norgestrel is a synthetic progestin that acts similarly to progesterone. This racemic or (±)-form is about half the potency of levonorgestrel. Norgestrel is used as a contraceptive, an ovulation inhibitor, and to treat menstrual disorders and endometriosis. See also: Norgestrel (note moved to).

C21H22D6O2

318.48

318.246

2376035-98-0

Levonorgestrel;797-63-7

71751212

Typically exists as solid at room temperature

3.3

1

2

2

23

609

6

[2H]C1=C2[C@](CC(C1=O)([2H])[2H])([C@H]3CC[C@]4([C@H]([C@@H]3CC2([2H])[2H])CC[C@]4(C#C)O)CC)[2H]

WWYNJERNGUHSAO-PWFLSJGKSA-N

InChI=1S/C21H28O2/c1-3-20-11-9-17-16-8-6-15(22)13-14(16)5-7-18(17)19(20)10-12-21(20,23)4-2/h2,13,16-19,23H,3,5-12H2,1H3/t16-,17+,18+,19-,20-,21-/m0/s1/i5D2,6D2,13D,16D

(8R,9S,10R,13S,14S,17R)-2,2,4,6,6,10-hexadeuterio-13-ethyl-17-ethynyl-17-hydroxy-7,8,9,11,12,14,15,16-octahydro-1H-cyclopenta[a]phenanthren-3-one

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

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

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*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.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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