Contamination Control

Question 1

Why do microorganisms pose a problem to pharmacists?

Answer 1

• small
• abundant and can grow in the most unlikely situations
• may cause disease - pathogenic
• some form spores, which are resistant to killing by heat, chemical, radiation or drying

Question 2

Describe bacteria.

Answer 2

• 0.7-4mcm
• unicellular
• rod shaped or spherical
• gram +/-
• nutrients: C, H, N, O, S, P, water
• aerobic/anaerobic

Question 3

Describe bacterial spores.

Answer 3

• undergo a rpofound biochemical change
• lead to formation of spore
• highly resistant to adverse environemtn

Question 4

Describe viruses.

Answer 4

• 20-250nm
• most pass through bacteria proof filters
• destruction by heat
• most inactivated at 60°C for 30 mins
• all killed by boiling water
• most chemical disinfectants have minimal virucidal activity

Question 5

Describe fungi.

Answer 5

• single or multicellular
• single - 5-10mcm in diameter
• commonly form spores
• yeasts, moulds and mushrooms

Question 6

Describe the air supply in aseptic areas.

Answer 6

areas have to be supplied and continuously flushed with air of suitable quality and at a positive pressure

achieved through High Efficiency Particulate Air (HEPA) filters of appropriate efficiency

Question 7

Describe the cleanroom specifications.

Answer 7

• air wuality standards in aseptic processing areas must meet both US and European requirements

European standards:

• Grades A, B, C and D classifications, instead of 100,1000
• use particle and microbial limits per cubic metre, instead of cubic foot
• require particle measurements at 5 microns in addition to 0.5 microns in grade A and B areas
• differentiate area cleanliness dynamically “in operation” and ‘at rest’

*

Question 8

How can you measure aerial contamination?

Answer 8

• settle plates
• Slit Samplers

Question 9

Describe the use of settle plates.

Answer 9

• nutrient plates which collects aerial contamination >100mcm which settle by gravity
• important hazard in aseptic work
• small particles not represented
• unaware of volume of air introduced

Question 10

Describe the use of Slit Samplers.

Answer 10

• air drawn through a slit or number of small holes
• beneath which rotates a “nutrient” agar plate
• deposition of particles depends on the rate of air flow, slit width, or dimensions of holes and distance between slit/holes and plate
• 99% efficiency of particle - 1mcm
• can count number of organisms associated with particles in a known volume of air drawn through the device

Question 11

What needs to be monitored in aseptic areas?

Answer 11

• particulate contamination
• temp & humidity
• air velocity and pressure

Question 12

What is required from those who work in aspetic areas?

Answer 12

• high standards of personal hygiene and cleanliness
• any medical condition (colds, skin infections) should be reported
• min. number of people should be present when work in progress
• adequate training
• gowning up

Question 13

What happens when viable cells are exposed to a lethal agent?

Answer 13

constant proportion, not a constant number of organisms are killed per unit time

plotted as:

• log number of viable cells
• log % viable cells against time

Question 14

Steriling conditions should produce what probability level?

Answer 14

a probability level of one in one million units being non-sterile (based on linear death kinetics)

Question 15

What is the sterility assurance and what does it mean?

Answer 15

10^-6

degree of assurance with which the process in question renders a population of items sterile

SAL 10^-6 = probability of not more than one viable organism in 1 x 10⁶ sterilised items of final product

Question 16

What is a D value?

Answer 16

decimal reduction value

value of a parameter of sterilisation (duration or absorbed dose) required to reduce the number of viable organisms by 90% or 1 log cycle

Question 17

What is inactivation factor and how do you calculate it?

Answer 17

reduction in the number of viable organisms produced by sterilising process

10^t/D

t = exposure time

D = D value

Question 18

Describe the two approaches to sterilisation.

Answer 18

• overkill method
  
  • sterilisation cycles based on inactivation of a greater N of organisms of greater resistance than the natural bioburden of the product
  • minimum SAL set for cycle
  • basis of traditional BP methods of heat sterilisation
    
    • with recomended combinations of time and temp
• bioburden method
  
  • requires knowledge of bioburden, D value and most resistant organism
  • prior to defining the sterilising cycle
  • allows sterilisation cycles to be run with acceptable SALs and minimal product detoriation

Question 19

What is the bioburden?

Answer 19

number of viable organisms a batch contains

Question 20

What is terminally sterilised vs non terminally sterilised products?

Answer 20

• terminally sterilised
  
  • products sterilised in their final container
  • method of choice whenever possible
  • manufactured in cleanroom
  • pyrogen free product
  • low microbial and particulate counts prior sterilisation
• non-terminally sterilised
  
  • prepared under aseptic conditions from previously sterilised materials
  • processed in cleanrooms
  • until sterilised
  • filled into final container in aspetic areas

Question 21

What are examples of terminally sterilised vs non terminally sterilised products?

Answer 21

terminally sterilised:

• steam sterilisation (heating in autoclave)
• dry heat sterilisation
• ionising radistion sterilisation

non-terminally sterilised:

• filtration

Question 22

Describe heat sterilisation.

Answer 22

• most reliable, versatile and readily available and economic method of sterilisation
• materials must be stable
  
  • heat decomposition of drug, interaction between drug and additives, modification of pharmacological activity of drug
  • stability of containers and closures
• safer compared to chemical sterilisation

net result depends on balance between

1. heat increasing rate of chemical reactions within bacteria cells = increased growth rate
2. destructive effect on more heat sensitive components of cell

Question 23

What is bacteriostasis?

Answer 23

point at which organisms die at the same rate they reproduce

Question 24

What is a further increase in temp past bacteriostasis point called?

Answer 24

destructive bactericidal effect

death rate increased and time to sterilise decreased

Question 25

What is one bad thing about vegetative bacteria?

Answer 25

relatively heat sensitive killed rapidly in hot water at 60-100deg spores usually require temp greater than 100deg

Question 26

What can be used as a biological indicator to test the efficiency of sterilisation processes and why?

Answer 26

bacillus stearothermophilus very heat resistant

Question 27

Describe the mechanism of destruction of moist heat.

Answer 27

* hot air with water vapour * unfolding protein = protein denaturation = uncoiling DNA = enzyme inactivation * changes to cell membrane = leakage of cell contents = death * relatively lower temp than dry heat

Question 28

Describe the mechanism of destruction of dry heat.

Answer 28

* absence of water * proteins much more stable - more enrgy required for unfolding * death occurs by * oxidation of cell components * requiring high temp

Question 29

What sterilisation process is preferred in BP?

Answer 29

saturated steam under pressure saturated steam: water vapour at a temp corresponding to the boiling point of water appropriate to its pressure

Question 30

What temp and time is required for steam sterilisation?

Answer 30

for aqueous preparations: a minimum of 121 deg for 15 mins

Question 31

Define sensible heat.

Answer 31

heat required to raise the temp of water to its boiling point

Question 32

Define latent heat.

Answer 32

additional heat, absorbed when boiling water is converted to steam at the same temp

Question 33

What are advantages vs disadvantages of moist heat?

Answer 33

+ * terminal sterilisation * any dose volume * good safety margin * virsuses killed * relatively short progress * suitable for dressings, rubber and some plastics * suitable for solutions, suspensions, surgical materials - * unsuitable for themrolabile materials, anhydrous materials (oils and powders) * organisms killed but not removed * skilled operators required * may damage glass and metal * batch process

Question 34

Describe dry heat sterilisation.

Answer 34

* terminally sterilised * temp 160-180 for 30 mins-2 hours * used for thermostable items - sensitive to moisture or impermeable to steam * very high temp (250deg) used to sterilise and depyrogenate glassware * anhydrous materials * glass and metal materials * powders * some rubbers

Question 35

Why is terminal sterilisation preferred?

Answer 35

provides a SAL that is possible to calculate, validate and control incorporates a saftey margin

Question 36

What are advantages vs disadvantages of dry heat?

Answer 36

+ * terminal sterilisation * any dose volume * viruses killed * used for anyhdrous material * sutiable for suspensions and solutions * less damage to glass and metal than moist heat - * drastic heat treatment * organisms killed but not removed * unsuitable for dressings, rubber and plastics * suspensions may give different crystal form on cooling

Question 37

What is sterilisation by filtration?

Answer 37

* API and preparations not sufficiently heat stable to allow autoclaving (thermolabile) * passage through a sterile membrane filter of normal pore size0.22mcm or less * organisms removed, instead of killed * doesnt sterilise: suspensions, medicaments highly adsorbed by filters, readily oxidised solutions, products unstable in solution

Question 38

What are advantages vs disadvantages of filtration?

Answer 38

+ * no thermal effects * remove dead and viable organisms - * difficult technique * require sterility testing prior to release = delay * will not remove or inactivate viruses * defects in filters not visible * possibility of adsorption of medicament, liberation of particles or alteration of pH by the filter * unsuitable for suspensions

Question 39

What is ionising radiation sterilisation?

Answer 39

* terminal sterilisation * minimum adsorbed dose of 25 kGy * gamma radiation - very penetrating * beta radiation (high speed electron) - poor penetration * physical and biochemical effects in microorganisms (DNA) * used for: heat sensiitve materials (disposables, dressings, organic compounds (antibitoics, hormones))

Question 40

What are advantages vs disadvantages of ionising radiation sterilisation?

Answer 40

+ * operates at ambient temp * terminal sterilisation * concinuous process * reliable and accurately controlled * treatment time short - few seconds - * expensive * damaging effect on some materials (glass, plastic, PVC) * protective precautions

Question 41

What gas can be used for sterilisation?

Answer 41

ethylene oxide

Question 42

What are the principles of good manufacturing practise?

Answer 42

* qualified personell * adequate premises * suitable production equipment, designed for easy cleaning and sterilisation * validated procedures for all critical production steps * environmental monitoring and in process testing procedures * precautions to minimise pre sterilisation bioburden