SARS-CoV-2[1]

For the purification of vaccines, propiolactone (β-propiolactone) can be employed. Following the collection of cells using low-speed centrifugation, Propiolactone (1:1000 v:v) was used to chemically inactivate SARS-CoV. SARS-CoV was incubated with propiolactone for a whole day at 4°C. To hydrolyze any remaining propiolactone, a second incubation is carried out at room temperature. and the vaccine's concentration. after BPL deactivation. To precipitate the inactivated virus, a combination of polyethylene glycol and sodium chloride (PEG-NaCl) was applied. following vaccination and concentration. A combination of polyethylene glycol and sodium chloride (PEG-NaCl) is used to inactivate the virus and precipitate it once propiolactone has been rendered inactive. As a final bacteriostatic agent, 1:10000 v:v propiolactone was applied. In Vero cells, the propiolactone-inactivated virus becomes less contagious [1].

The influenza A virus was inactivated with propiolactone (β-propiolactone) and injected intramuscularly into mice at a dose of approximately 25 mg total protein. Young BALB/c mice treated with propiolactone inactivation are not fatally affected by SARS. Even while the virus continued to grow in the mice's respiratory system, by day five it had disappeared. After the mice were infected, 1.5 μg of total hemagglutinin protein were produced as a result of propiolactone treatment, which was negative [1].

Absorption, Distribution and Excretion
THE LD50 BY SKIN APPLICATION IS LESS THAN 5 ML/KG IN THE GUINEA PIG, INDICATING CONSIDERABLE ABSORPTION.
BETA-PROPIOLACTONE BINDS IN VIVO TO DNA, RNA & PROTEINS OF MOUSE SKIN. DEGREE OF TUMOR-INITIATING ACTIVITY IS PROPORTIONAL TO EXTENT OF DNA BINDING BUT NOT TO EXTENT OF RNA OR PROTEIN-BINDING. MAJOR RNA & DNA BINDING PRODUCT IS 7-(2-CARBOXYETHYL)GUANINE. S-2-CARBOXYETHYLCYSTEINE ... FOUND IN ACID HYDROLYSATE OF PROTEIN ... .

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Metabolism / Metabolites
Propiolactone is completely hydrolyzed after 3 hours of being in an aqueous solution and this time can be even faster in the presence of cellular debris and cell culture media. When in water, the lactone ring opens at the alkyl and acyl bonds. The degradation products of propiolactone are not toxic.
BETA-PROPIOLACTONE CAN REACT WITH CHLORIDE ION TO FORM 3-CHLOROPROPIONIC ACID, ESPECIALLY IN BLOOD PLASMA.
BETA-HYDROXYPROPIONIC ACID, THE HYDROLYSIS PRODUCT OF BETA-PROPIOLACTONE, FAILED TO PRODUCE EITHER LOCAL SARCOMAS IN SC STUDY IN RATS ... OR SKIN TUMORS AFTER APPLICATIONS TO SKIN OF MICE.

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Biological Half-Life
The half-life of propiolactone in water is of 225 minutes.
No reports found; [TDR, p. 1048]

Protein Binding
Propiolactone is highly bound to proteins showing an almost 2-fold binding increase when compared to DNA and RNA.

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Toxicity Data
LC50 (rat) = 25 ppm/6h

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Interactions
WHILE BETA-PROPIOLACTONE CAUSES LIVER NECROSIS AND RENAL TUBULAR DAMAGE WHEN GIVEN BY ITSELF INTRAVENOUSLY, IF IT IS ALLOWED TO REACT WITH PROTEINS BEFORE INJECTION, THE TOXICITY IS SAID TO BE VERY MUCH REDUCED.
The ability of UV-A light (320-400 nm) to induce cellular transformation in vitro and to modify chemical carcinogen-induced cellular transformation was investigated in BALB/c 3T3 cell cultures. When administered as a series of nontoxic exposures, UV-A alone was found to induce cellular transformation as a linear function of the numbers of UV-A exposures. Possible interactions of UV-A with environmentally encountered chemical carcinogens were studied by examining the effects of UV-A light exposures on cellular transformation in cells exposed to the direct acting carcinogen, beta-propiolactone, an alkylating agent, with a standard initiation/promotion protocol. Twenty-four hours after a single treatment with 2.5 ug/ml of beta-propiolactone, cells were exposed to 3.0 kJ/sq m of UV-A light. UV-A exposures were repeated weekly for up to 5 weeks, after which cells were fixed, stained and dishes were scored for type III transformed foci. Weekly exposures to UV-A alone for 5 weeks induced approximately 3 foci/dish. Treatment with beta-propiolactone alone induced approximately 1 focus/dish (background was 0.17 foci/dish). A combination of the two treatments resulted in a marked increase in the yield of transformed foci/dish, with the UV-A enhancement increasing with increasing numbers of exposures (approximately 10 foci/dish after a single exposure to beta-propiolactone and five UV-A exposures). These results suggest a synergistic interaction between beta-propiolactone and subsequent UV-A exposures in the induction of in vitro neoplastic transformation.
Studies have been initiated to find compounds that can trap direct-acting carcinogens within the lumen of the GI tract and thus prevent these carcinogens from attacking tissues of the host. Sodium 4-mercaptobenzene sulfonate is a potent nucleophile and was found to react rapidly in vitro with the direct-acting carcinogen beta-propiolactone. In further investigations sodium 4-mercaptobenzene sulfonate was shown to inhibit mutagenesis resulting from exposure of Salmonella typhimurium strain TA-100 to beta-propiolactone and a second direct-acting carcinogen, N-methyl-N'-nitro-N-nitrosoquanidine. Subsequent experiments were performed to determine if sodium 4-mercaptobenzene sulfonate would inhibit beta-propiolactone induced carcinogenesis in vivo. In the first of these, sodium 4-mercaptobenzene sulfonate was administered by oral intubation to female A/J mice 5 min before oral administration of beta-propiolactone. Under these conditions inhibitions of carcinogenesis of the forestomach occurred. In a second experiment, sodium 4-mercaptobenzene sulfonate was given by rectal intubation 5 min before beta-propiolactone also administered intrarectally. Administration of beta-propiolactone intrarectally produced adenomatous polyps of the large intestine. The occurrence of these neoplasms was inhibited by the prior administration of sodium 4-mercaptobenzene sulfonate. These results suggest that sodium 4-mercaptobenzene sulfonate has the capacity to trap direct-acting carcinogens and to inhibit the occurrence of beta-propiolactone induced neoplasia.
Using a two-step carcinogenesis protocol, SENCAR mice were initiated with 25 ug 7,12-dimethylbenz(a)anthracene and were then treated twice weekly with either (a) 0.5 mg beta-propiolactone or (b) 1 ug fluocinolone acetonide followed in 30 min by 0.5 mg beta-propiolactone. The tumor incidence for the group receiving fluocinolone acetonide prior to beta-propiolactone was significantly greater than for beta-propiolactone alone (p< 0.0005). Under these experimental conditions beta-propiolactone alone showed neither promoting activity nor complete carcinogenic activity. These results were not anticipated, but the reasons for their occurrence are being explored.
Studies have been initiated to find compounds that can trap direct-acting carcinogens within the stomach. Sodium thiosulfate is a potent nucleophile and in initial experiments was found to inhibit mutagenesis resulting from exposure of Salmonella typhimurium strain TA100 to the direct-acting carcinogens beta-propiolactone and styrene oxide. In in vitro experiments sodium thiosulfate was shown to maintain its nucleophilicity in the acid pH range. It reacted with beta-propiolactone as rapidly at pH 2 as at pH 7.4. Thus sodium thiosulfate has the prerequisite attributes to inhibit the carcinogenic effects of electrophiles in the stomach. Experiments were performed in which sodium thiosulfate was administered by oral intubation to female A/J mice 5 min before oral administration of beta-propiolactone. Under these conditions, inhibition of formation of the forestomach tumors occurred. The data obtained suggest that use of nucleophiles to protect against direct-acting carcinogens is a potential strategy for chemoprevention.

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Non-Human Toxicity Values
LC50 RAT INHALATION 250 PPM/30 MIN
LC50 RAT INHALATION 25 PPM/6 HR
LD50 RAT YOUNG IV 225 + OR - 55 MG/KG, SCORED @ 24 HR
LD50 GUINEA PIG SKIN APPLICATION < 5 ML/KG

[1]. Immunogenicity and protective efficacy in mice and hamsters of a β-propiolactone inactivated whole virus SARS-CoV vaccine. Viral Immunol. 2010 Oct;23(5):509-19.

[2]. Anti-SARS-CoV-2 IgG antibody response among Indian COVID-19 patients using β-propiolactone-inactivated, whole virus-based indirect ELISA. J Virol Methods. 2021 Jan;287:113996.

Drug Warnings
Because production of skin cancer is felt to be the overriding consideration in the toxic potential of BPL, all contact with liquid BPL should be avoided.

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Pharmacodynamics
When employed under conditions of maximum effectiveness, propiolactone is approximately 25 more active as a vapor phase disinfectant than formaldehyde, 4000 times more active than ethylene oxide and 50000 times more active than methyl bromide. It has been shown to be mutagenic by inducing cell transformation, chromosomal aberrations and chromatoid exchange. Propiolactone has been shown to be mutagenic in both somatic and germ cells.

C3H4O2

72.06

72.021

57-57-8

25037-58-5

2365

Colorless to light yellow liquid

1.2±0.1 g/cm3

162.0±0.0 °C at 760 mmHg

−33 °C(lit.)

35.0±16.1 °C

2.2±0.3 mmHg at 25°C

1.445

-1.33

0

2

0

5

57.9

0

O1C(C([H])([H])C1([H])[H])=O

VEZXCJBBBCKRPI-UHFFFAOYSA-N

InChI=1S/C3H4O2/c4-3-1-2-5-3/h1-2H2

oxetan-2-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.)