Aim: Preparation of Ascorbic Acid Injection
Objective
The aim of this experiment is to prepare a sterile aqueous solution of ascorbic acid suitable for parenteral administration, ensuring the correct concentration, pH, and sterility, and adhering to pharmaceutical quality standards.
Introduction
Ascorbic acid (Vitamin C) is a water-soluble vitamin with potent antioxidant properties. It plays a critical role in collagen synthesis, wound healing, immune function, and as a cofactor in enzymatic reactions. Deficiency of ascorbic acid results in scurvy, characterized by weakness, gum bleeding, and poor wound healing.
Therapeutic Uses:
- Treatment of Vitamin C deficiency
- Adjunct therapy in wound healing
- Antioxidant in critical illnesses or sepsis
- Supportive therapy in certain cancer or viral infections
Chemical Profile:
- Chemical Name: L-Ascorbic Acid
- Molecular Formula: C₆H₈O₆
- Molecular Weight: 176.12 g/mol
- Appearance: White to slightly yellow crystalline powder
- Solubility: Freely soluble in water, insoluble in ethanol and ether
- Stability: Sensitive to light, heat, and alkaline conditions; hence, preparation must be done under controlled conditions.
Parenteral administration is preferred when rapid vitamin C supplementation is needed, as oral absorption is limited at high doses.
Materials and Equipment
Materials
- Ascorbic acid (USP grade)
- Sterile distilled water for injection
- pH adjusting agents: Sodium bicarbonate or Sodium hydroxide (to adjust pH if needed)
- Preservatives (optional, e.g., benzyl alcohol 0.9% w/v for multi-dose vials)
Equipment
- Analytical balance
- Glass beakers, graduated cylinders, and volumetric flasks
- Magnetic stirrer
- pH meter or pH indicator strips
- Sterilizing-grade filter (0.22 μm) for filtration
- Autoclave (121°C, 15 psi) if heat-stable
- Laminar airflow chamber for aseptic filling
- Sterile ampoules or vials for injection
- Hot air oven (optional for drying ampoules)
Formulation Example (100 mL Injection)
| Ingredient | Quantity | Function |
| Ascorbic acid | 1–2 g | Active pharmaceutical ingredient |
| Sterile distilled water | q.s. to 100 mL | Vehicle / solvent |
| Sodium bicarbonate/NaOH | q.s. to pH 3–4 | pH adjustment |
| Benzyl alcohol (optional) | 0.9 g | Preservative (for multi-dose vials) |
Remarks:
- Typical parenteral concentrations: 1–2% w/v
- pH adjustment is critical; ascorbic acid is acidic (pH 2–3 in water), and too low a pH can cause vein irritation.
Theoretical Calculations
Step 1: Determine weight of ascorbic acid required
For 1% w/v in 100 mL:
Amount required = 1% × 100 mL = 1g
For 2% w/v in 100 mL:
Amount required = 2% × 100 mL = 2g
Step 2: pH Adjustment
- Ascorbic acid solutions are naturally acidic (pH 2–3).
- Slowly add sodium bicarbonate or NaOH to adjust pH to 3–4 for parenteral safety.
- Avoid excessive alkalinity, as it degrades ascorbic acid.
Procedure
Preparation of Solution
- Accurately weigh the required amount of ascorbic acid.
- Dissolve in 80% of the total volume of sterile distilled water using a magnetic stirrer.
- Check pH using a pH meter. Adjust to 3–4 with dilute sodium bicarbonate or NaOH.
- Make up to the final volume with sterile distilled water.
- Mix gently to avoid oxidation.
Sterilization
- Filtration (preferred for heat-sensitive solutions): Pass the solution through a 0.22 μm sterilizing filter into a sterile container under laminar airflow.
- Autoclaving: Only if using heat-stable formulations and properly sealed containers (rarely used due to thermal degradation).
Filling and Sealing
- Fill sterile solution into ampoules or vials under aseptic conditions.
- Seal ampoules by flame or cap vials with sterile rubber stoppers.
- Label with drug name, concentration, batch number, and expiry date.
Quality Control Tests
- Appearance: Clear, colorless to slightly yellow solution, free of particulates
- pH: 3–4
- Sterility Test: As per USP <71>
- Assay of Ascorbic Acid: Titrimetric (using 2,6-dichlorophenolindophenol) or HPLC
- Pyrogen Test: LAL assay or rabbit pyrogen test
- Particulate Matter: According to USP <788>
Storage Conditions
- Store in tight, light-resistant containers to prevent degradation.
- Temperature: 2–8°C (refrigeration) is preferred for prolonged stability.
- Avoid exposure to air, light, and heat.
- Use freshly prepared solutions if possible.
Precautions
- Perform all steps under aseptic conditions to avoid microbial contamination.
- Avoid prolonged exposure to air, as ascorbic acid oxidizes rapidly.
- Adjust pH carefully; avoid over-alkalinization to prevent degradation.
- Use amber-colored vials to protect from light-induced decomposition.
- Label vials clearly, including concentration, preparation date, and expiry.
References
- United States Pharmacopeia (USP 43-NF 38), Ascorbic Acid Injection Monograph.
- Indian Pharmacopoeia (IP), 2020, Volume II.