PA20024 Albert Bolhius Stuff!

Question 0

What is latent heat?

Answer 0

When this heats exchanged it results in a change in physical state, no temperature change occurs, you cant see any temperature change eg converting from liquid to gas releases a lot of latent heat, known as super latent heat.
Latent heat can penetrate the surface of bacteria  therefore is required to be released in moist heat sterilisation, that’s why we use dry saturated steam as this is the only steam that releases this (as well as sensible heat)

Question 1

What is the only steam suitable for autoclaving (moist heat sterilisation?)

Answer 1

Dry saturated steam! 
This sits on the phase boundary between superheated steam (too dry, not enough moisture) and supersaturated steam (too wet, not hot enough)

Question 2

What is sensible heat?

Answer 2

When this heat is exchanged, a change in temperature occurs. Ie heating up a liquid so that it’s more likely to turn into a gas involves sensible heat. You can see the temperature increase.

Question 3

Why is moist heat sterilisation better than dry heat sterilisation?

Answer 3

Moist heat uses dry saturated steam, which penetrates and damages bacteria using its latent heat and by hydration, it’s a more rapid heat transfer and therefore takes less time.
Dry heat sterilisation does not use steam, involves oxidation which is less effective at inactivating bacterial spores than hydrolysis, and it is much slower heat transfer, you have to heat for a lot longer.

Question 4

How much latent heat must be released in order to have enough energy transfer to kill bacteria?

Answer 4

946 BTU/lb latent heat

Question 5

At 121 degrees, 15 PSIG, steam contains about 5 X more latent heat than sensible heat. What is so important about latent heat?

Answer 5

When the dry saturated steam comes into contact with the bacteria that is cooler, it will condense on the surface of the bacteria and release later heat into it which penetrates it, letting in more and more steam, a vacuum is then created which kills the bacteria.

Question 6

Why is condensation of steam good for killing bacteria?

Answer 6

When the steam condenses there’s more contact with the steam and microbe surface.
Steam gets sucked into the microbe through PORINS, more penetration.
You only get condensation with dry saturated steam

Question 7

What type of sterilisation involves the use of autoclaving?

Answer 7

Moist heat sterilisation

Question 8

In dry heat sterilisation, inactivation of microbes is principally by _____. In moist heat sterilisation, inactivation of microbes is principally by_______.

Answer 8

Dry heat: oxidation. Think: no water, only hot air, air=O2

Moist heat: hydrolysis. Think: water involved; OH, hydrolysis

Question 9

What does the BP suggest as temperature and time for dry heat sterilisation?

Answer 9

160 degrees for 1 hour

normally 150-180 degrees for 30 mins- 2 hours

Question 10

What’s better about fan assisted ovens?

Answer 10

Temperature variation is under a lot More control than in a non fan assisted oven

Question 11

What are the two gases used for gas sterilisation?

Answer 11

Ethylene oxide

Formaldehyde (low temperature steam formaldehyde: LTSF)

Question 12

What do you need to watch out for when using ethylene oxide gas to sterilise?

Answer 12

It’s explosive if used over 3.6% concentration in the presence of AIR 
Therefore we need to minimise the amount of air, ethylene oxide must be used in vacuum conditions, a special oven that sucks out all air.
Ethylene oxide is also toxic, and can leach out of products after sterilisation

Question 13

Why do you need to control humidity when using ethylene oxide?

Answer 13

If organisms are in their dried out state then they’re more resistant, therefore humidity needs to be controlled so it doesn’t get too dry, humidity should be 30-70%, this is critical

Question 14

What are the set amounts, temp and time for sterilisation by ethylene oxide?

Answer 14

EITHER
900mg/L at 54 degrees for 3 hours
Or 450mg/L at 30 degrees for 16 hours

Question 15

What are the set amounts, and temp for sterilisation by LTSF (formaldehyde)?

Answer 15

15-100mg/L at 70-75 degrees
(requires higher temps than ethylene oxide which can be used at room temps)
LTSF also has a longer cycle time than ethylene oxide

Question 16

What do gamma ray sterilisers use as their radioactive source?

Answer 16

Cobalt 60
This needs to be used in a reinforced concrete building, and it should be stored under water when not in use, very radioactive. 

Question 17

What’s the dose specified by the BP for radiation use to sterilise?

Answer 17

25 KGy

Question 18

When using radiation as the sterilisation process, what will the units of your D value be in?

Answer 18

It will be a Dose of radiation, so usually kGy

Question 19

What is radiation used to sterilise?

Answer 19

Disposable plastics

Raw materials

Question 20

If you are we’re using filtration with high temperature, which type of filter would you use?

Answer 20

Cellulose acetate,

This has thermal stability

Question 21

What is breakthrough when applied to the process of filtration?

Answer 21

The number of bacteria after filtration, ie the number of bacteria that have managed to sneakily squeeze through the filter pores some how.
Larger pore size, higher bio burden, and large volume of filtrate all increase the likelihood of breakthrough occurring!

Question 22

What is growthrough with regards to filtration?

Answer 22

This is when bacteria start to divide and grow through the filter membrane if a filtrate is left in contact with a membrane for too long.
To stop this occurring, minimise the time of the filtration process… The EP limits it to 4 hours. 

Question 23

What’s the only practical way to sterilise liquids? 

Answer 23

By filtration
You can’t use heat, radiation or gases
Could potentially use high intensity light (UV light) if transparent 

Question 24

How do we test air for contamination?

Answer 24

Use settle plates and air samplers

Question 25

How do we test surfaces for contamination ?

Answer 25

Use contact plates and swabbing

Question 26

When manufacturing batches in industry, why are you always going to have to make a few more than the number you’re intending to sell?!

Answer 26

Because sterility tests need to be done on a fraction of the batch, and any product tested can’t be used anymore, as sampling itself can introduce contamination. Eg if your batch is 100-500, 10 of these need to be sampled.

Question 27

What are examples physical indicators, used for quality assurance of sterilisation processes?

Answer 27

Digital recording of temperature in coolest region in heat sterilisation process
Leak tests and pressure testing with gas sterilisation
Filtration: use bubble point pressure test (looking for the pressure required to push air through the filter)

Question 28

What is an example of chemical indicators used for sterilisation quality assurance?

Answer 28

Change of colour observed such as brownes tubes, indicator strips, autoclave/Bowie dick tape.
Brownes tubes turn from red to green if autoclaving has worked and the high temp was reached.

Question 29

What are biological indicators for quality assurance of sterility tests?

Answer 29

Bacteria/ test organisms added that would be your worst case scenario, eg brevundimonas diminuta for filtration

Question 30

What is the Attest biological indicator? (hint; we used it in class!)

Answer 30

Consists of: outer tube, paper strip indicated with b.stearothermophilis spores, and a glass ampule inside tube with indicator inside.
After autoclaving, crush glass, if bacteria hasn’t been killed off propley, spores will germinate, and the indicator will turn yellow. If autoclaving has worked it will turn purple

Question 31

What is parametric release?

Answer 31

Where all relevant parameters of that process (eg temp, pressure, time) have been accurately controlled, monitored and logged.data from this is then processed into a system of release that will give assurance that a product is of intended quality!
Only applies to products terminally sterilised in their final containers

Question 32

What are pyrogens?

Answer 32

Endotoxins, so things like LPS on gram negative bacteria. can induce fever

Question 33

Two tests to test for pyrogens?

Answer 33

Rabbit pyrogen test, see if rabbits develop fever when injected with product.  
LAL test (limulus amoebocyte lysate pyrogen test) 
Blood cells are isolated from horseshoe crabs, lysate contains enzymes that are activated by endotoxins, and these enzymes cause clotting