Effects of Eluent pH (ODP-50 6D)

A sample of different proteins and peptides was separated with a various pH values of the eluent. Under acidic conditions (pH2.3, pH3.0), Leu-Enkephalin and Leu-Trp-Met have been strongly retained and the elution times for the substances are quite similar; therefore, the separation is not satisfactory. In contrast, when an alkaline according to the fact that, under alkaline condition (pH10) with Asahipak ODP column, a better separation with sharper peaks of the substances has been obtained in a shorter time. In addition, ODP columns have higher theoretical plate numbers under alkaline conditions.
Refer to Analysis of Oligopeptides with Acidic and Alkaline Eluent (ODP-50 6D)

Sample:

  1. 1.Glu-Gly-Phe
  2. 2.Met-Enkephalin
  3. 3.Leu-Enkephalin
  4. 4.Leu-Trp-Met
Scroll
No. Theoretical plate number
pH2.3 pH3.0 pH6.7 pH10
1 6100 1100 3800 -
2 7400 1100 4700 5700
3 7600   600 3800 6700
4 6100   500 2400 8000
Column
：Shodex Asahipak ODP-50 6D (6.0 mm I.D. x 150 mm)
Eluent
：Solution/CH3CN=85/15
(pH2.3) 0.05 % TFA aq.
(pH3.0) 50 mM Acetic acid aq.
(pH6.7) 50 mM Ammonium acetare buffer
(pH10.0) 50 mM Ammonium acetare buffer
Flow rate
：1.0 mL/min
Detector
：UV (220 nm)
Column temp.
：30 ℃

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