Download A Formula for Success

A Formula for Success
Scenario 1:
• You are a medicinal chemist for a small
biopharmaceutical company. One of the
drugs that they are interested in
developing is a lysozyme tablet that can
be taken to help fight certain bacterial
infections. Your job is to develop a
formula for the excipients in the tablet.
Where do you begin?
Excipients
• Inactive substance of a drug that holds the
active ingredients.
– Holds drug together (binder)
– Coatings –addresses taste and size
– Preserving – extends expiration date
– Dilution of dosage/fillers
– Stability/buffers
• Lysozyme – enzyme that disrupts cell wall
of bacteria, found saliva, tears,
Scenario 2:
As part of the development team, you want
to focus your formula to address pH, taste,
and fillers. The plan has now changed to
formulate this for capsules. You have not
received the active ingredient data yet so
you must move on in your development of
the formula. What ingredients will you use
and at what concentrations?
Buffer Design
• Design an experiment to help you decide the
best buffer and concentration to use. This may
require multiple experiments.
• Remember to:
– Decide on buffer and concentration first
– Do the calculations to help you weigh right amount
– Use distilled water and add less water than needed to
give you room to add acid or base as you adjust pH
(near 7)
– Test buffer efficacy by adding acid in increments and
checking pH
Other components
• Taste – sugars (sucrose, dextrose,
fructose)
– Which one? Mass/volume”
• Fillers – starch or gelatin
– Which one? %?
Scenario 3:
The folks working with the lysozyme have
been pulled over to into a new project and
now you have been given the job to
determine what is the best concentration
to use in the proposed drug. You will need
to design an experiment to determine this.
Making Media
Day 1
• Make up 100 ml of Nutrient Agar and 50 ml of Nutrient
Broth. Each can go in a separate, labeled 100 ml media
bottle. Add distilled water to correct volume.
• Autoclave to kill all living organisms.
Day 2
• Melt agar and pour plates
• Inoculate plates with bacteria and incubate overnight
Day 3
• Pick colonies and inoculate 25 ml of Nutrient Broth in a
sterile culture tube and incubate overnight.
Sterile Technique
• Follow SOP for Sterile Technique in Lab
Manual
• Pour a starter plate and allow to solidify
• Triple Z streak on your plate to inoculate it
and incubate overnight
• Transfer 20 ml of Nutrient Broth into sterile
tube
• Pick colony and inoculate broth. Incubate
overnigth
Lysozyme Assay
• Make enzymes up in desired concentrations in buffer.
Concentrations of 1-10 mg/ml work well.
• Set Spec at 450nm
• Blank spec 20 with buffer
• Pour 3 ml of bacteria culture into each tube. Check to
see the OD450 is around 0.8. If not, add buffer to bring
it down to that range.
• After recording the absorbance,
add 0.1 ml of enzyme to tube
and monitor.
• Repeat for other enzyme concent.
Scenario 4:
It has been discovered that the acid
concentration of the stomach has a major
impact on the activity of the active
ingredient. You now need to determine
the concentration of the acid and then
adjust the buffer to improve the maximum
activity of the lysozyme
Titration to Determine Acid Molarity
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Pour 25 ml of the “stomach acid” into a beaker.
Add two drops of phenolphthalein to the solution
Pipet 1 ml of 1M NaOH using a 1ml pipet.
Add the NaOH to the beaker one drop at a time while swirling the beaker.
You should see the solution turn pink for a second and then disappear.
As you get near to the end point (neutralization), it will take longer for the
pink to disappear.
At some point, the pink will not go away as you have passed neutral and the
solution is now basic.
Record the total amount of NaOH needed to reach the end point .
To calculate the molarity of the solution use the following equation:
MA x VA = MB x VB
where M is molarity, V is volume, A is acid (HCl in this case) and B is base
(NaOH in this case)
Scenario 5
• The senior staff likes the data you have
generated so far and wants to move
forward with the development of the drug.
However, marketing is asking that the drug
be given a distinguishable color. They are
concerned however about the dyes that
are used due to adverse reactions. You
need to research the dyes and find a
single, negatively charged dye to use.
Scenario 6:
You and your team have completed our
formulations for the new drug. You are to
submit at least two capsules to quality
control for testing. How close you are to
meeting project goals determines how
large of a bonus your team gets. Your
team may also get to name the new
product.