Visible Spectrophotometric Method for the Estimation of Molnupiravir Using 1,10 Phenanthroline Reagent

The most commonly employed reagents for the determinations are the following:

• N-1-Naphthyl ethylene diamine dihydrochloride called as Bratton–Marshall reagent (BM reagent) undergoes diazotization for the determination of sulpha drugs, local anaesthetics, etc.
• 3-Methyl-2- benzothiazolinone hydrazone hydrochloride, which is generally known as MBTH. The color production is depending upon the oxidative coupling of this reagent with phenols, amines, carbonyl compounds, etc.
• Para dimethyl amino benzaldehyde (PDAB) and para dimethyl amino cinnamaldhyde (PDAC) - certain amines condenses with a variety of aldehydes in acidic media to give a product which are colored and oxidisable.
• Phosphomolybdotungstic acid called as Folin – Ciocalteu reagent (FC reagent) undergoes oxidation, reduction (or) hydrolysis based on the functional group to be determined such as amines, phenols etc.
• 1,2-Naphthoquinone-4-sulfonate sodium (NQS) is a chromogenic agent used for the determination of 1° aromatic amines.
• 2,6-Dichloroquinone chloroimide, which is commonly known as GIBB'S reagent used for the identification and estimation of phenols.
• Oxidation followed by complexation. Ex: Bathophenanthroline, 1,10 - phenanthroline, 2,2'-Bipyridine.
• Oxidation followed by charge transfer complex formation ex: Metol-KIO3.

Oxidation Followed by Complexation

1,10-Phenanthroline   Synonym for 1,10-phenanthroline is o-phenanthroline. It is extensively chromogenic reagent used for the colorimetric or visible spectrophotometric   determination of a number of drugs and pharmaceutical substances. It is also known as ortho- phenanthroline (O- phenanthroline). It is constantly utilized in combination with a popular oxidant or oxidizing agent namely ferric chloride.

MECHANISM

The analysis of drug by using 1,10- phenanthroline involves two steps. Initially the solution of the drug under investigation is allowed to react with 1,10- phenanthroline and Ferric chloride (at elevated temperature). During this process, the drug gets oxidized and results in the generation of ferrous iron [i.e., ferric chloride is converted to ferrous chloride, i.e., (II)]. The ferrous now complexes with 1,10- phenanthroline to form an orange red colored complex. In this method, the reason of the addition of phosphoric acid is to stop the photochemical reduction. The following are the chemical reactions involved:

Drug+ FeCl₃                         

Reduced form of Fe⁺³ + 1,10-Phenanthroline → (Fe⁺²-1,10-Phenanthroline) complex

MATERIALS AND METHODS

Chemicals and reagents: Molnupiravir (pure drug) was obtained as a gift sample from Hetero Labs. Ferric Chloride(fecl₃) Grade (Thermo    Fisher Scientific India Pvt.Ltd), 1,10 Phenanathroline, Grade (Thermo Fisher Scientific India Pvt.Ltd) were purchased from Qualigens Company, Methanol, Rankem(Laboratory Reagent)  and All chemicals and reagents used were of analytical grade. Tablets containing drug Molnupiravir (Molnuvid (200mg) were procured from local market.

Instruments: Electronic Precision balance (Infra Instruments Pvt. Ltd., Chennai), PC Based Double Beam Spectrophotometer 2202 (Systronics, India).

Drug name: Molnupiravir

Structure:   

Fig.3.1: Chemical structure of Molnupiravir

Iupac Name: [(2R,3S,4R,5R)-3,4-dihydroxy-5-[(4Z)-4-(hydroxyimino)-2-oxo-1,2,3,4 tetrahydropyrimidin-1-yl] oxolan-2-yl] methyl 2- methyl propanoate

Category: Anti -viral drug.

Molecular weight: 336.35 g/mol

Molecular formula: C₁₃H₁₉N₃O₇

Melting point: 156-157?

Dosage form: Capsules

Description: White solid

pH: 7.2

Solubility:  Soluble in methanol.

Route of administration: Oral route.

Storage: Stored in a closed container at room temperature

Dose: Movfor(200mg)

Principle:                                                                 

Molnupiravir by using 1,10-Phenanthroline involves two steps. Initially the   solution of the drug Under investigation is allowed to react with 1,10-Phenanthroline and Ferric chloride (at   elevated Temperature). During this process, the drug gets oxidized and results in the generation of ferrous Iron [i.e., ferric chloride converted to ferrous chloride. The ferrous   now complexes with 1,10- Phenanthroline to form an orange red colored complex

Step:1

Step:2

Experimental Work

UV-Visible Spectrometry for Molnupiravir

Preparation of Solutions

Preparation of Stock solution: 10mg of Molnupiravir was taken in a 10ml volumetric flask, dissolved in 2ml of Methanol and made upto 10ml with Distilled water. (1mg/ml or 1000µg/ml)

Preparation of Standard solution: 1ml of stock solution was taken in a 10 ml volumetric flask, and made upto 10 ml with Distilled water (100µg/ml)

 Preparation Of 0.5% 1,10-Phenanthroline: 500mg 0f 1,10-Phenanthroline was taken in a 100ml volumetric flask , dissolved in 2 ml of methanol and made upto 10ml with Distilled water.

Preparation Of 0.3% Fecl₃ : 300mg of Fecl₃ was taken in a 100ml volumetric flask , dissolved in 2 ml of methanol and made upto 10ml with Distilled water.

Method Development

Determination Of Λ Max of Molnupiravir

Preparation of solution: 1ml of standard drug solution (100µg/ml) was taken in a 10ml volumetric flask, 1ml of ferric chloride and 1,10-phenanthroline reagent (0.5%) was added. It was heated for 15mins in a water bath for colour development and was allowed to cool. Then it was made up to 10ml with distilled water.

Preparation of blank solution: 1ml of ferric chloride and 1,10-phenanthroline was taken in 10ml of volumetric flask, then it was heated for 15mins. Then it was allowed to cool and made upto 10ml with distilled water.