Sterile preparations

STERILE PHARMACEUTICALS 1. PARENTERALS

Parenteral products are products that are administered to the body by injection. Because this route of administration bypasses the normal body defence mechanisms, it is essential that these products are prepared with a higher degree of care and skills than utilised in preparing conventional oral or topical products. The finished product must be sterile, non-pyrogenic and free from extraneous insoluble materials. These products must satisfy a number of requirements for parenteral products. Formulation of Parenteral products Sterile formulations must meet a number of special criteria such as: a. Sterility b. Particulate material c. Pyrogen free d. Stability e. pH f. Osmotic pressure During the formulation of parenteral products, the following factors are critical: (a) The vehicle in which the drug is dissolved or dispersed (b) Volume (dose) of the injection (c) Adjustment to isotonicity (d) Adjustment of pH (e) Stabilisers (f) Preservatives (g) Adjustment of specific gravity (for spinal anaesthesia) (h) Concentration units The formulation of injections are not different to the formulation of products by other routes except for certain requirements which parenteral products must satisfy. The major ones are limits to the level of pyrogens present and of particulate matter. For formulation of injections, the injection route dictates the volume of the formulation. Hence the solubility of the drug in the selected vehicle is critical in the formulation. The preferred vehicle is water as it is well tolerated by the body, is easy to administer and has a large solvent capacity. Due to the wide variety of contaminants in mains water, water for injections must be used as the vehicle for parenteral products. Water for injections (WFI) must be sterile and free from pyrogens. Solutions for parenteral use and injections with a volume of 100mL or more must comply with the BP test for the absence of particulate matter. Particles >50

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microns can be detected by visual inspection. Examples of such particulate materials are cellulose, glass, rubber cores, cloth or cotton fibres. Suitable filtration media for removal of particulate materials are sintered glass filters or membrane filters with a pore size of 0.45-1.2m. As parenteral formulations are administered directly to tissues and systemic circulation, formulations prepared should not vary significantly from physiological pH, which is about 7.4. In certain cases however, acidic or alkaline solutions may be needed to solubilize drugs. The acceptable pH range is 3-10.5 for IV preparations and 4-9 for other routes. Buffers are included in injections to maintain the pH of the packaged product. However, the buffer used in the injection must allow the body fluids to change the product pH after injection. Acetate, citrate and phosphate buffers are commonly used in parenteral products. The osmotic pressure of blood is approximately 300 milliOsmoles/L and ideally any sterile solution would be formulated to have the same osmolarity e.g., 0.9% w/v Sodium Chloride iv solution has an osmolarity of 308 mOsmole/L and 5% w/v Dextrose iv solution has an osmolarity of 280 mOsmol/L. Intravenous solutions that have larger osmolarity values (hypertonic) or smaller osmolarity values (hypotonic) may cause damage to red blood cells, pain, and tissue irritation. Parenteral solutions which are hypotonic need to be adjusted to isotonicity. Osmolarity adjustment is made usually by using sodium chloride, glucose or mannitol using one of the following methods; The freezing point depression method Sodium chloride equivalent Molar concentrations Serum osmolarity

Adjustment of tonicity using the Freezing Point Depression Injection solutions are often made isotonic with 0.9% w/v sodium chloride solutions. The amount of solute, to the required dilution necessary to make a solution isotonic, can be determined from the freezing point depression. The freezing point depression of blood plasma and tears is 0.52C. Thus solutions which freeze at 0.52C have the same osmotic pressure as body fluids. Hypotonic solutions have a freezing point depression higher than 0.52C and require the addition of a solute to depress the freezing point to 0.52C. The amount of adjusting substance added to these solutions may be calculated from the equation: W = where: W = percentage concentration of adjusting substance in the 0.52 - a b

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a b

= =

final solution freezing point depression of the unadjusted hypotonic solution freezing point depression of a 1% w/v concentration of the adjusting substance

A list of freezing point depression values is detailed in Table 6, page 53 of the Pharmaceutical Codex. Note: that some components of injections, such as buffers and antioxidants, affect the tonicity and these have to be accounted for when adjusting the tonicity of parenteral formulations. Other components, such as preservatives, which are present in low concentration, have little effect on the tonicity. Antimicrobial agents Aqueous preparations which are prepared using aseptic precautions and which cannot be terminally sterilised may contain a suitable antimicrobial preservative in an appropriate concentration. No antimicrobial preservative is added when: The volume to be injected in a single dose exceeds 15mL unless otherwise justified The preparation is intended for administration by routes where for medical reasons an antimicrobial preservative is not acceptable If the drug formulation itself has sufficient antimicrobial activity Antimicrobial agents are added to multiple dose vials to inhibit the growth of microbial organisms which may occur accidentally and contaminate the product during use. The antimicrobial agents selected must be stable and effective in the parenteral formulation. Because they are effective in the free form, their activity can be greatly reduced by interaction with components of the injection. Rubber closures have been shown to take up antimicrobial preservatives from the injection solution. The effectiveness of antimicrobial agents can be tested by challenging the product with selected organisms. The test procedure will evaluate the antimicrobial activity of the preservative in the packaged product (refer BP 1998). Table 1 lists anti-microbial agents commonly utilised in parenteral formulations

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Table 1: Commonly utilised anti-microbial agents in parenteral preparations Anti-microbial agent Benzalkonium Chloride* Benzyl alcohol Chlorobutol Phenol Chlorocresol Phenylmercuric salts Methylhydroxybenzoate Concentration used (% w/v) 0.01 1-2 0.25-0.5 0.5 0.1-0.3 0.002 0.1-0.2

Antioxidants Many drugs in aqueous solutions are easily degraded by oxidation. Small volume parenteral containing these drugs often contain an antioxidant. Bisulphites and metabisulphites are the commonly used antioxidants in aqueous injections. Antioxidants must be carefully selected for use in injections to avoid interaction with the active. Antioxidants have a lower oxidation potential than the active and hence are either preferentially oxidized or block oxidative chain reactions. Injection formulations may, in addition also contain chelating agents, such as EDTA or citric acid, to remove trace elements, which catalyse oxidative degradation. SINGLE DOSE PREPARATIONS The British Pharmacopoeia (B.P.) 1998 define single dose preparations as: The volume of the injection in a single dose container is sufficient to permit the withdrawal and administration of the nominal dose using a normal technique. MULTIPLE DOSE PREPARATIONS (BP 2004) Multidose preparations are multidose aqueous injections which contain a suitable antimicrobial preservative at an appropriate concentration except when the preparation itself has adequate antimicrobial properties.

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CONTAINERS Containers for parenteral products are produced from one of 3 types of glass or from one of a variety of plastic materials. Types of glass Type I: Commonly known as neutral glass. It has a high resistance to hydrolysis and withstands autoclaving, weathering and solution of pH of up to 8. Type II (sulphated glass): Containers may be treated with moist sulphur dioxide at high temperature to create a neutral surface film with high hydrolytic resistance. Lower resistance to autoclaving than for type I glass. Type III (soda glass): This offers very little resistance to hydrolysis and should only be used for powders for reconstitution prior to injection and for non aqueous preparations. Type II and Type III glass containers should be used once only for parenteral preparations. Types of containers Glass ampoules (Fig. 1(a)) are the most commonly used single dose containers and can range from sizes of 1 to 50mL. For aqueous solutions, neutral glass is used. After filling, glass ampoules are sealed by fusion of the glass and hence there is no danger of entry of micro-organisms. Amber glass ampoules are available for light sensitive products. Clear ampoules may be used provided that the ampoules are packaged in a light-resistant box. Glass vials sealed by rubber closures (Fig. 2(b)) are commonly used as multi-dose containers. The rubber closure is held in place by an aluminium sealing ring. The rubber closure permits the penetration of a syringe needle to allow the withdrawal of a dose of injection. Filling and hand sealing of glass ampoules Ampoules must first be rinsed out using water for injections to remove any dust, particulate matter and/ or glass fragments. Using a syringe, gently draw the volume required to be filled plus an excess volume. Invert the syringe to allow the air to rise towards the needle and push up the plunger to remove all air. Attach the membrane filter and needle to the syringe (Fig. 1(c)). Discard the first 0.5-1mL of filtrate. Invert the ampoule over the needle and expel the required volume of liquid into the ampoule taking care not to splash the liquid into the neck of the ampoule.

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Each ampoule must also contain a slight excess volume of product. This is necessary to allow the nominal injection volume to be drawn into a syringe. Refer to table 1 for excess volumes required. Using a twin-jet burner (Fig. 2), adjust the platform height and flame intensity. Try an empty ampoule first before sealing your products. Position the ampoule between the flames. Grip the end of the neck with a blunt-nosed forceps and when the glass is soft enough, pull off the top of the ampoule vertically and gently. Leave the tip in the flame for a second or two longer and then remove. The tip should now be smoothly and evenly rounded (Fig. 3a) Ampoules should have a reliable seal which can be readily leak tested. A good seal will not deteriorate during the lifetime of the product. Filling and hand sealing of glass vials Glass vials are filled using the same rinsing and filling procedure as for ampoules. In the case of vials containing a fixed number of dose units, an excess volume is required to allow the stated number of doses to be withdrawn (Table 2). Vials are sealed using rubber closures which are held in place by an aluminium sealing. A hand crimper is available in the laboratory to seal the rubber closures and aluminium sealing onto the vials. INSPECTION OF FILLED INJECTABLE PRODUCTS Injection ampoules and vials should be checked for defects, cracks, chips, damage to the seals and closures. Injection solutions when examined under suitable conditions of visibility should be free from particles. BP1998 Appendix XIII A & B describes tests for particulate contamination. STORAGE AND LABELLING Previously ampoules were labelled by use of paper labels. Presently this has been superseded by direct printing on the glass. For dispensing purposes in the practical lab, a paper label is used and should contain the following information. The strength expressed in terms of the amount of active ingredient in a suitable dose volume The name and concentration of any added substance e.g., antioxidant, antimicrobial preservative Indication that a single dose preparation should be discarded after first use Special labelling requirements for particular product e.g., powders for reconstitution prior to use, concentrated solutions requiring dilutions prior to use The date after which the preparation is not intended to be used (expiry date) The conditions under which the preparation should be stored

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Note: In practice, the expiry date should be determined for each product by conducting suitable stability studies. In hospital, sterile preparations are often given an expiry date of 1 week from date of manufacture/ reconstitution. Units of concentration The concentration of the components in parenteral products may be expressed in various ways: Percentage weight/volume. Examples include: Magnesium sulphate injection 50%w/v, sodium chloride intravenous infusion 0.9% w/v Weight per unit volume. Examples include: Atropine sulphate 600g/ml or ephedrine hydrochloride injection 30mg/ml Millimoles per unit volume. Examples include: Potassium chloride solution, strong (sterile) contains 2mmol each of K+ and Cl- per ml; Calcium chloride injection BP contains 2.5 mmol of Ca2+ and 10mmol of Cl- in 5ml. ESSENTIAL READING B. P. 1998/2004 The Pharmaceutical Codex Pharmaceutical Dosage Forms and Drug Delivery Systems, H C Ansel, L V Allen Jr and N G Popovich

Fig. 1 (a) glass ampoule

Fig. 1 (b) Multi dose glass vial

Fig. 1(c) Syringe with membrane filter and needle attachment

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Fig. 2 Twin jet burner

Fig. 3 sealed ampoules with good seal (a) and faulty seals (b-e)

Table 2:

Recommended overages for official injectable products

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2.

OPHTHALMIC PREPARATIONS

Ophthalmic preparations are sterile, liquid, semi-solid or solid preparations intended for administration upon the eyeball and/or to the conjunctiva or to be inserted in the conjunctival sac. Several categories of eye preparations may be distinguished: -eye drops -eye lotions -semi-solid eye preparations -ophthalmic inserts

PREPARATION OF EYE DROPS Extemporaneous preparation of eye drops involves the following: Preparation of the solution Clarification Filling and sterilization Preparation of the solution The aqueous eye drop vehicle containing any necessary preservative, antioxidant, stabilizer, tonicity modifier, viscosity modifier or buffer should be prepared first. Then the active ingredient is added and the vehicle made up to volume. Clarification The BP has stringent requirements for the absence of particulate matter in eye drop solutions. Sintered glass filters or membrane filters of 0.45 1.2 m pore sizes are suitable. The clarified solution is either filled directly into the final containers which are sealed prior to heat sterilization or filled into a suitable container prior to filtration sterilization. Clarified vehicle is used to prepare eye drop suspensions which are filled into final containers and sealed prior to sterilization. Sterilization This can take the form of: Autoclaving at 115C for 30 minutes or 121C for 15 minutes Heating at 98 - 100C for 30 minutes together with either benzalkonium chloride 0.01% w/v or chlorhexidine acetate 0.01% w/v or phenylmercuric acetate or nitrate 0.002% w/v or thiomersal 0.01% w/v. This method is described in the BP (1980) but is no longer a pharmacopoeial recommended method.

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Filtration through a membrane filter having a 0.22m pore size into sterile containers using strict aseptic technique. Filling should take place under Grade A laminar airflow conditions. A suitable filter holder for extemporaneous preparation is illustrated in Figure 4. The filter assembly is sterilized by autoclaving before use. Dry heat sterilization at 160C for 2 hours is employed for non-aqueous preparations such as liquid paraffin eye drops. Silicone rubber teats must be used. Immediately following sterilization the eye drop containers must be converted with readily breakable seal, such as a viskring, to distinguish between opened and unopened containers. Fig. 4: Filter holders for bacterial filters (A) 13mm and 25mm types fitted to syringes (B) Cutaway view of 25mm type

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Labelling requirements are summarised in Tables 3 and 4

Table 3: General Labelling requirements for eye drop containers

Requirement Fully identify the product

Specify storage conditions State product expiry date Warning label Specify volume Ensure correct use

Include on label Title;either name and concentration of active ingredients or reference to official monograph giving these details. If monograph allows more than one concentration then state the one used Store in a cool place or Protect from light Month and year of expiry Not to be taken e.g. 5mL e.g. Shake the bottle for a suspension

Table 4: All locations Hospital Wards Operating Theatres Clinics Domiciliary

Additional labelling requirements for use in specific locations Include concentration of active ingredient and name and concentration of any antimicrobial present Patients name. The eye to be treated. Date of opening of bottle and/or date to discard Single dose for once-only use. Marked with indication and concentration of active ingredient. No preservative. Outer package fully labelled Single dose or multidose used once only Avoid contamination of contents during use Discard 4 weeks after opening Keep out of reach of children Plus instructions on how to use

PREPARATION OF EYE LOTIONS The purpose of eye lotions is to assist in the cleaning of the external surfaces of the eye. This might be to help remove a non-impacted foreign body or to clean away conjunctival discharge. Eye lotions intended for use in surgical or first-aid procedures should not contain antimicrobial preservatives and should be in singleuse containers. There is no intention to use an eye lotion to deliver any active ingredient to the eye but rather to remove unwanted gross contaminants from the

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eye. Thus these preparations should be very simple and the most common eye lotion consists of sterile normal saline. This preparation typifies the requirements of an eye lotion which are: Sterile and usually containing no preservative Isotonic with lachrymal fluid Neutral pH Large volume but not greater than 200mL Non-irritant to ocular tissue Labelling These should include: Title identifying the product and concentration of contents Sterile until opened Not to be taken Use once and discard remaining solution Expiry date Preserved eye lotion would need the additional labelling: Avoid contamination of contents during use Discard remaining solutions not more than 4 weeks after first opening The lotions should be supplied in coloured fluted bottles and sealed to exclude microorganisms. PREPARATION OF EYE OINTMENTS Eye ointments are popular and duplicate many of the therapeutic options offered by eye drops. Ointments have the disadvantage of temporarily interfering with vision, but have the advantage over liquids of providing greater total drug bioavailability. However, ointments take a longer time to reach peak absorption. Eye ointments must be sterile and may contain suitable antimicrobial preservatives, antioxidants and stabilizers. USP ophthalmic ointments packaged in multi-dose containers are to contain an antimicrobial substance unless otherwise directed in the monograph or if the formulation itself is bacteriostatic. The most commonly used agents include the two mercurials, phenylmercuric nitrate and thiomersal, the parabens and chlorobutanol. It is necessary also that such ointments are free from particulate matter that could be harmful to the tissues of the eye. The EP and BP have limits for the particle size of incorporated solids which will be met if all particles have been reduced to <25m.

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The basic components of an eye ointment are given below: Liquid paraffin 1 part Wool fat 1 part (to facilitate incorporation of water) Yellow soft paraffin 8 parts Hard paraffin as required to produce required consistency in hot climates Preparation of eye ointments Eye ointments are normally prepared using aseptic techniques to incorporate the finely powdered active ingredient or a sterilized concentrated solution of the medicament into the sterile eye ointment basis. Immediately after preparation the eye ointment is filled into the sterile containers which are then sealed so as to exclude microorganisms. The screw cap should be covered with a readily breakable seal. All apparatus used in the preparation of eye ointments must be scrupulously clean and sterile. Certain commercial eye ointments may be sterilized in their final containers using ionising radiation.

Preparation of eye ointment basis

The paraffins and the wool fat are heated together and filtered, while molten, through a coarse filter paper in a heated funnel into a container which can withstand dry heat sterilization temperatures. The container is closed to exclude microorganisms and together with contents is maintained at 160C for 2 hours. Containers for eye ointments Eye ointments should be supplied in small sterilized collapsible tubes made of metal or in a suitable plastic. The tube should not contain more than 5g of preparation and must be fitted or provided with a nozzle of a suitable shape to facilitate application to the eye and surrounds without allowing contamination of the contents. The tubes must be suitably sealed to prevent microbial contamination. Eye ointment may also be packed in suitably designed single-dose containers. Labelling This includes the following: The names and percentages of the active ingredients The date after which the eye ointment is not intended to be used

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The conditions under which the eye ointment should be stored normally at a temperature not exceeding 25C The name and concentration of any antimicrobial preservative or other substance added to the preparation A statement to the effect that the contents are sterile providing the container has not been opened Essential Reading BP 1998 Section on Eye Preparations page 1428-1431

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