23
250
250
200
200
150
150
100
100
50
50
0 0 20 40 60
80 100 120 140 160 180 200 k'(NUCLEOSHELL®
RP18plus) Figure 7. Comparison of NUCLEOSHELL®
160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0
-10 -20
0.00 1.00 2.00 3.00 4.00 5.00 6.00 7.00 min RP 18plus and NUCLEOSHELL® Biphenyl (methanol/water) 450 1 2 3 350 5 Figure 8. Comparison of NUCLEOSHELL®
0 0 5 10 15 20 k'(NUCLEOSHELL® 25 RP8plus) RP 8plus and NUCLEOSHELL® 400 1,3 300 4 1 6 2 5 3 4 7 6
A B
Figure 9: Separation of a test mixture on NUCLEOSHELL® and NUCLEOSHELL®
Biphenyl
Chromatographic conditions: column A NUCLEOSHELL® column B NUCLEOSHELL®
RP 8plus, Biphenyl, 100 mm x 3 mm, acetonitrile/
water 55:45 (v/v), 0.56 mL/min, 40 °C, UV, 254 nm, 1. propiophenone, 2. propylparaben, 3. desoxycorticosterone, 4. biphenyl, 5. fluorene, 6. progesterone, 7. pyrene.
RP 8plus 200 4,5 7 6 150 2 100 1 50 3 4 5 6 0 7
A B
-50 0.0 2.5 5.0 7.5 10.0 12.5 15.0 min
Figure 10: Separation of a test mixture on NUCLEOSHELL® and NUCLEOSHELL®
Biphenyl
Chromatographic conditions: column A NUCLEOSHELL® column B NUCLEOSHELL®
Biphenyl, 100 mm x 3 mm,
acetonitrile/water 55:45 (v/v), 0.56 mL/min, 40 °C, UV, 254 nm, 1. propiophenone, 2. butylparaben, 3. desoxycorticosterone, 4. biphenyldiisobutylphthalate, 5. pyrene, 6. pentylbenzene, 7. chrysene.
Although both phases have biphenyl containing modifications, several reversals of the order of elution occur. The reason for this is the dilution of the p-p interactions by the two isobutyl groups. Mononuclear aromatics, which show rather less interactions with the biphenyl groups, gain in retention in comparison to the polynuclear aromatics and the steroid progesterone.
y phase
: Comparison of interaction strength and dispersion ry phase
Comparison of interaction strength and dispersion results acetonitrile
SHELL® Biphenyl, 2.7µm e column)
NUCLEOSHELL® Biphenyl, 2.7µm (reference column)
SHELL® π², 2.7µm
SHELL® RP18plus, 2.7µm SHELL® RP8plus, 2.7µm
SHELL® π², 2.7µm p², SHELL® RP18plus, 2.7µm
NUCLEOSHELL® 2.7µm
SHELL® RP8plus, 2.7µm
NUCLEOSHELL® RP18plus, 2.7µm
NUCLEOSHELL® RP8plus, 2.7µm
SHELL® Biphenyl, 2.7µm ce column)
′() 1.001.00
0.96 1.23 0.78
0.96 1.23 0.78
0.96 1.23 0.78
results acetonitrile RSD
′() 1.00
RSD ---
0.18 0.24
0.18 RSD ------
0.18 0.24
0.240.24 0.24
0.24
Table 4. Comparison of interaction strength and dispersion. Stationary phase results acetonitrile
Conclusion NUCLEOSHELL®
RP 18plus,
Chromatographic conditions: column A NUCLEOSHELL® column B NUCLEOSHELL®
RP 18plus, Biphenyl, 100 mm x 3 mm, acetonitrile/
water 50:50 (v/v), 0.50 mL/min, 35 °C, UV, 254 nm, 1. biphenyl, 2. fluorene, 3. progesterone, 4. butylbenzene, 5. diisobutylphthalate, 6. pyrene, 7. pentylbenzene, 8. o-terphenyl.
RP 18plus 7 250 2
120 110 100 90 80 70 60 50 40 30 20 10 0
-10 -20
0.0 2.5 5.0 7.5 10.0 12.5 15.0 17.5 min Biphenyl (methanol/water) 30 35
1
2
5 1
3 4 7 2 3 5 8 4 7 6
A B
Figure 11: Separation of a test mixture on NUCLEOSHELL® NUCLEOSHELL®
Biphenyl p² and 6 8
Biphenyl is a new biphenyl modified core shell silica. The introduction
of bulky substituents to the silicon atom improves the stability of this stationary phase against acidic conditions. The selectivity is significantly different from octadecyl and octyl sorbents. The biphenyl ligand plays a crucial role in the interactions of this phase. Due to the influence of the two isobutylgroups, significant differences in the selectivities between NUCLEOSHELL®
Biphenyl and NUCLEOSHELL® results methanol
′() 1.001.00
1.00
1.54 0.91 0.48
1.54 0.91 0.48
1.54 0.91 0.48
results methanol ′()
results methanol RSD RSDRSD --- ---
---
0.340.34 0.36
0.430.36 0.43
0.34 0.36 0.43
p² can be observed. The change
of acetonitrile and methanol makes it possible to influence the strength of the p-p interactions and is therefore another exciting tool for optimising the separation.
References
[1] L.R. Snyder, J.J. Kirkland, and J.L. Glajch, Practical HPLC Method Development, 2nd Ed. (John Wiley & Sons, New York, 1997) pp. 234-240. [2] F. Chan, L.S. Yeung, R. LoBrutto, and Y.V. Kazakevich, J. Chromatogr. A 1069, 217-224 (2005). [3] M.R. Euerby, P. Peterson, and W. Campbell, J. Chromatogr. A, 1154, 138-151 (2007). [4] K. Croes, A. Steffens, D.H. Marchand, and W. Roe, J. Chromatogr. A, 1098, 123-130 (2005). [5] H. Riering, N. Bilmann, and M. Ganin, Int. Labmate 43, 16-18 (2018). [6] J.J. Kirkland, J. Chromatogr. A, 1060, 9-21 (2004). [7] J.J. Kirkland, J.L. Glajch, and R.D. Farlee, Anal. Chem. 61, 2-11 (1989).
WWW.LABMATE-ONLINE.COM
Absorbance [mAU] k'(NUCLEOSHELL® Biphenyl)
Absorbance [mAU]
k'(NUCLEOSHELL®
Biphenyl)
Absorbance [mAU]
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