HPLC Coursework
weight for Noscapine BP is 413.4; the sample is soluble in acetone but not water (British Pharmacopoeia
Commission 2007, p. 1166). Hence normal phase HPLC is suitable for separation of related substance in
Noscapine BP.
Normal phase HPLC allows easy separation of isomers or mixture of compounds with varying
functional groups (Valko 2006, p. 51). If the solute functional groups are in suitable conformation to
interact with the absorbent surface the retention time will be much greater. The majority of
separation as required by BP and retention time. The mobile phase in general is more ploar than the
stationary phase. The use of non-polar stationary phase and a polar mobile phase indicates RP-HPLC.
'
log k =slope+log k 0
log k
'
log k '
'
log k 0
is the
is the intercept
1.2.2 Suitability
The basic criteria for RP-HPLC are that the testing material has molecular weight < 2000 and is
soluble in non-polar to weakly polar solvents (Harris 2000, p. 723). According to BP2007 the molecular
weight for Nifedipine BP is 346.3; the sample is insoluble in water but soluble in acetone (British
Pharmacopoeia Commission 2007, p. 1142). Hence RP-HPLC is suitable for separation of related
substance in Nifedipine BP.
RP-HPLC allows separation of isomers or mixture of compounds with varying functional groups at a
much quicker speed than its normal phase counterpart. The majority of the impurities (specified impurities
A and B) share the same chemical structure as nifedipine with only two function group variations (Figure
2). Minor impurities C and D are sub-fragments of nifedipine (Figure 3; Figure 4). Due to a high number of
impurities the use of normal phase HPLC is ineffective; it is likely that impurity signals overlap with each
other. RP-HPLC is therefore an better method of differentiating these structurally-related compounds.
2 Calculation
2.1 Concentration of unknown paracetamol solution
Let the concentration of unknown paracetamol solution be x.
Due to the same retention time the peak areas (10500mAu and 8750mAu) are
compared.
As same amount of solution is injected, by simple ratio,
10500mAu = x : 8750mAu
500uM
x = 416.67uM
500uM
= 500 umol in 1
3 Bibliography
British
Pharmacopoeia
Commission,
2007.
Monographs.
Pharmacopoeia 2007. 2007th ed.: Stationery Office Books. p. 1670.
In
British
British
Pharmacopoeia
Commission,
2007.
Monographs.
Pharmacopoeia 2007. 2007th ed.: Stationery Office Books. p. 1142.
In
British
British
Pharmacopoeia
Commission,
2007.
Monographs.
Pharmacopoeia 2007. 2007th ed.: Stationery Office Books. p. 1166.
In
British