Beruflich Dokumente
Kultur Dokumente
(HPLC)
RAJEEV KUMAR
M.PHARM, Ist Sem,
M.I.E.T. MEERUT
High Performance Liquid Chromatography
(HPLC)
A convenient way to treat solvents is to filter them through a milipore filter under
vacuum. This treatment remove both gases and suspended matter.
Instruments – Pumping System
• Pumping system requirement
1.High P (6kpsi),
2.pulse-free out put,
3.Flow rate 0.1 ~10ml/min.
4.flow reproducibility,
5.resistant to corrosion
• Two major type of pumps are :
1. Displacement pump (Screw-driven
syringe pump)
I. Adv: pulse free,
II.Dis adv: small capacity(250 ml),
no gradient elution
I. D. (mm) 4 – 10 1 – 4.6
packing size (µ m) 3, 5, 10 3, 5
Advantage of “new”:
1. less HPLC-grade solvents needed (less expensive)
2. Columns can be coupled to increase the number of plates.
1.adequate sensitivity
3. fluorescence detector
6. electrochemical detector
– Fluorescence detectors:
-Hg or Xe lamp
-Fluorescing species or fluorescent
derivatives
-high sensitivity
Instruments – Detectors
DA:
- limited sensitivity (1 ng/μl),
-highly Tem dependent (0.001°C),
- no gradient elution
– Electrochemical detectors:
It is based on Amperometry,
voltammetry, coulometry and
conductometry
Adv:
- simplicity,
- high sensitivity,
- convenience
- wide-spreading application
Instruments – Detector
- computer controlled
- computer re constructed chromatogram obtain
- spectra of eluted peaks
- major problem is gas phase sample is needed in mass
spectrometry
Reference
H.H. Willard, J.A. Dean’ L.L. Merritt “Instrumental method of analysis”
www.orbigen.com
www.pharma.uka.edu
www.waters.com
www.laboratorytalk.com
www.chromspec.com
www.capsandstems.com
www.springerlink.com
THANK YOU