Preparation of Sterile Products

First Division: Product development

The ultimate objective of this department is to develop a sterile product with therapeutic effect in the patient.

Therefore the Product Development parenterals product considering the following factors:

  • Vehicles
  • Solutes
  • Containers
  • Formulation
  • Stability

I. Vehicle

  • Aqueous
    • Water (frequently used) is the vehicle for all natural body fluids. The superior quality requirement is described under “Water for Injection, USP”
    • It is often used as isotonic vehicles
    • Other aqueous vehicles are Sodium chloride injection, Dextrose and Sodium chloride Injection, and Lactated Ringer‘s Injection.
  • Non-aqueous
      The USP provides the following specifications for such vehicles:
    • The fixed oils must be of vegetable origin, in order that they may be metabolized.
    • Must be liquid at room temperature – it must contain ester of unsaturated fatty acids.
    • Oxidation is prevented by the addition of antioxidants, such as tocopherol.
  • Pyrogen-free
    • The vehicle water used in irrigation ad parenterals solutions.
    • Should be rendered pyrogen free.
    • Gram negative bacteria produce the most potent pyrogenic substances.
    Sources of Pyrogens
    • Water
    • Equipment
    • Solute
    • Manufacturing methods
    Ways of Destroying Pyrogens;
    Pyrogens can be destroyed by heating at high temperatures, such as:
    • 650 Celsius for 1 minute
    • 250 Celsius for 45 minutes
    • 180 Celsius for 4 hours
    • Note: Autoclaving will not destroy pyrogens
    • Adequate washing, accompanied by dry-heat depyrogenation will render containers and equipment free from pyrogens.

II. Solutes

  • Therapeutic agents
  • Added substances such as antibacterial agents antioxidants buffers and tonicity agents
Factors to be considered in the choice of solutes:
  • the level of microbial and pyrogenic contamination
  • solubility characteristics, as determined by the physical or chemical form of the compound
  • freedom from gross dirt
  • Added substances
  • Antimicrobial agents
  • Buffers/Tonicity Contributors
  • Antioxidants

III. Containers

  • Glass
  • Rubber
  • Plastic
  • Metal
  • Type I — generally suitable for all parenterals
  • Type II — treated with sulfur dioxide; suitable for parenterals solutions, which is buffered; has a pH below 7; not reactive with the glass
  • Type III — suitable for anhydrous parenterals liquids or dry substances
  • Type NP — not for parenterals use; suitable only for tablets, capsules, suspensions, ointments and extractives.
  • The flexibility of low density polyethylene polymer for ophthalmic preparations without introducing contamination into the remaining contents in the container.
  • The flexible bags of polyvinyl chloride or select polyolefins for large volume parenterals, has the added advantage that no air interchange is required. The bag simply “collapses” as the solution flow out of the bag.
Rubber closures
  • These are used to seal the openings of cartridges, vials, and bottles, providing a material that is soft and elastic enough to permit the entry and withdrawal of a hypodermic needle.
Metal caps
  • For parenterals products, metal is used as aluminum caps to crimp rubber closures in place.

IV. Formulation

  • Solvent system
  • Additives
  • Freeze dried products
  • Suspensions
  • Emulsions
  • Effect of the route of administration
Freeze dried or Lyophilized Parenteral Products
  • Parenteral solutions intended to be freeze dried must be aqueous, because the drying process involves the removal of water from the product, by sublimation after it is frozen.
  • This process is only applicable to thermolabile or unstable drugs in liquid.
Characteristics of Freeze dried/lyophilized products
  1. Of uniform color and texture
  2. Contains a supporting matrix of solids, sufficient to maintain the original volume of the product after drying.
  3. With sufficient strength to prevent crumbling during storage.
  4. Rapid reconstitution.
Parenteral emulsions
  • The principal problem in parenterals emulsions is the attainment and maintenance of uniform size of droplets, ranging from 1 to 5 microns (for the internal phase).
  • Intravenous nutrient emulsions have been made containing cottonseed oil, dextrose and lecithin.
Effect of Route of Administrations
— The intended route of administration has a marked effect on the formulation of a parenterals product.
The factors to be considered are:
  • Volume in which the dose of a drug is given.
    • For intracutaneous route, not more than 0.2 mL is given due to slow absorption.
    • For subcutaneous route is about 1 mL or less.
    • For intraspinal, 10 mL or less.
    • For intravenous, large volume parenterals may be given by setting the infusion set to 250 and above; by syringe, about 20 mL or less.
  • Isotonicity
    • Consider patient comfort:
      — Cause headache and vomiting as a result of disturbances of osmotic pressure

V. Stability

This factor is considered by the Product development Division and should evaluate the effect of the components, particularly if the product is subjected to thermal sterilization.

Second Division: Product Area

Production Area includes all the steps in:

  1. Accumulation of raw materials and packaging materials
  2. Preparation of the product
  3. Filling and Finishing of the final product
Environmental control

Maintenance of the aseptic area and sterile room is done by normal routine of cleaning of these areas at the end of the working day or during the night.

  • Laminar flow enclosures
  • Ultraviolet radiation
  • Filtration
Methods of environmental control tests:
  • Air sampling techniques
  • Exposure of nutrient agar plates to the settling of microorganisms from the air
  • Total Sterility Test
  • Use of instrumental scanners
Air Sampling Technique
Collection of particulate matter from air through a clean, sterile membrane filter
Examined microscopically
Filters be placed on the culture media
Incubated for the detection of microorganisms
Collection of air sample into a measured volume of nutrient broth
Microorganisms from the broth by filtration on the membrane filter
Drawing a measured volume of air
“slit sampler”
causes the air to impinge on the surface of a slowly rotating agar plate.

Advantage: This sampler is portable and can be disinfected and hand carried whenever needed.

Exposure of nutrient agar plates to the settling of microorganisms from the air

If pathogenic microorganisms are particularly of interest, blood agar plates may be needed.

Note: A period of one hour may not collect one microorganism under conditions of use in a well controlled aseptic area.

Total Sterility Test
  • The best indication of the efficiency of the aseptic filling process.
  • This is done by filling and sealing sterile fluid thioglycollate or trypticase soy broth in sterile containers
  • The entire lot is incubated and examined for the appearance of growth of microorganisms
Use of instrumental scanners
  • There are instruments currently being utilized to obtain particle counts from a measured volume of air.
  • These instruments operate on the principle of the measurement of light scattered from particles passed through the optical system.
Filtration of Parenteral Solutions

Solutions must be filtered for the following primary objectives:

  1. Clarification – is also termed as “polishing” and a highly polished solution would require the removal of particulate matter down to at least 3 microns in size
  2. Sterilization by filtration – is the removal of approximately 0.2 to 0.45 microns, the size of viable microorganisms and spores; and is used for pharmaceutical solutions that are heat labile
Filter type & its Principal components
  1. ASBESTOS PAD (disposable): Asbestos fibers or wood cellulose
  2. CELLULOSE ESTER MEMBRANE: Cellulose acetate and/ or cellulose nitrate
  3. DIATOMACEOUS EARTH CANDLES: Diatoms of SiO2;Fe2O3,and others (about 80%)
  4. SINTERED GLASS CANDLES/DISC: Borosilicate glass
  5. UNGLAZED PORCELAIN/SELAS CANDLES: Clay, silicon dioxide
Filling of Parenteral Solutions:
  • repetitively forcing a measured volume of liquid through the orifice of the delivery tube, which is introduced into the container
  • The size of the delivery tube will vary from that of about 20 gauge hypodermic needle to a tube of 0.5 in. or more in diameter
  • For large volume parenterals like infusion solutions, filling may be made by gravity;
    • A pressure pump filling machine or vacuum filling machine.
    • Vacuum filling is the best method, since it is faster and can be rendered automatic.
Filling of Sterile Solids
  • Sterile solids eg, antibiotics is more difficult to subdivide evenly into containers.
Types of Ampoule sealing methods
  • Tip sealing method
  • Pull sealing method
Sealing of vials and bottles
  • Vials and bottle are sealed by closing the opening with a rubber closure or stopper..
  • When rubber closures are to be inserted mechanically, the surface of the closure is often “halogenated” or “siliconized” to give it less friction.
  • Mechanical stoppering has been developed to meet the need for high speed production.
  • Rubber closures are held in place by means of aluminum caps.
Sterilization of Parenteral products
  • Thermal
  • Non-thermal
  • Chemical
  1. Thermal Sterilization
    1. Dry heat
      • For substances unaffected at a temp of 148 to 260 degrees in oven, at an exposure time of 45 minutes
      • Kills spores, vegetative forms of microorganism
      • Ideal for sterilizing glasswares, metal wares, anhydrous oils
    2. Moist heat sterilization
      • Effective than dry heat method
      • It destroys spores and vegetative forms of bacteria at 121 degrees Celsius for 20 minutes at 15 psi in an autoclave
      • Ideal for sterilizing rubber stoppers, glasswares, bottles, cellulose membrane filters
    3. Fractional Sterilization Method
      • Tyndallization – makes use of moist heat at 100 degrees Celsius, using free flowing steam. It is normally performed by 2 to 3 exposures, alternated with intervals at room temperature or incubator temperature.
      • Inspissation – a fractional method of sterilization at 60 degrees Celsius in an oven, alternated with intervals at room temperature or incubation for 2 to 3 days.
      • Fractional methods of sterilization are effective for vegetative forms of microorganisms, but not for spores. Bacteriostatic agents may be used to improve this method.
  2. Non-thermal Sterilization Methods
    1. Ultraviolet radiation

      This is used to aid reduction of air borne contamination produced by mercury vapor lamps. This method has poor penetration capability. Its effectiveness depends on:

      1. length of time of exposure
      2. intensity of radiation
      3. susceptibility of the microorganism
    2. Ionization radiation

      This radiation method makes use of high energy emitted from radioactive isotopes such as cobalt 60 (gamma rays) or by cathode or beta rays (mechanical acceleration of electrons to high velocity and energy).

  3. Chemical Sterilization Methods
    1. Gas Sterilization
      • A method of sterilization using:
        1. formaldehyde and sulfur dioxide gas — has a problem of leaving “residual deposits”.
        2. Ethylene oxide and beta propiolactone — limited to dry powders and plastic materials.
      • This is done by autoclaving the substance at 55 Celsius at 27 PSI.
      • The ethylene oxide gas is introduced and exposure is timed at 60 min.
    2. Surface Disinfectant
      • A method of sterilization of work surfaces with the use of chemical disinfectants for environmental control.
      • Phenolic and quaternary ammonium compounds are commonly used.
      • Gamma rays are more reliable because there is no mechanical breakdown, but it has a disadvantage of rare source and cannot be shut off immediately.
      • Accelerated electrons provide higher and more uniform dose outfit and can destroy organisms by stopping its reproduction.
    3. Filtration by Membrane Filtration QC Parenterals
      • Sterility test
      • Pyrogen test
      • Clarity test
      • Leaker test
      • Safety test