A New Concept of End-capping for Reversed Phase Silica Material:
Conversion of Silanol Groups to Siloxane Bond by Heating
Norikazu Nagae1, Etsuko Shearer2, Tomoyasu Tuskamoto1
1. ChromaNik technologies Inc. Namiyoke, Minato-ku, Osaka 552-0001Japan
2. BioNik Inc. 3397-19 Obuchi, Fuji, Shizuoka 417-0801 Japan*Corresponding author email: nagae@chromanik.co.jp

www.bionikinc.com

www.chromanik.co.jp

A reversed phase column like C18 has been widely used in various fields such as pharmaceutical, medical, environmental, industrial and food area. Especially silicabased C18 column has showed high performance although an end-capping must be done enough in order to decease a peak tailing of a basic compound caused bya residual silanol group on the silica surface. An End-capping has been recognized to be an important factor for a silica based reversed phase column. In the beginning
residual silanol groups after C18 bonding were reduced by bonding with trimethylsilyl reagents (TMS). This end-capping method has been improved by various end-capping
reagents with not only a monofunctional group but also a difunctional group, a high reaction temperature or a long reaction time. In the past ten years some column
manufacturers have developed the unique method that positive charge added on the silica surface suppressed influence of residual silanol groups against a basic compound.
In this study, not only bonding with an end-capping reagent but also conversion of silanol groups to siloxane bond by heating were tried as an end-capping. After bonding with
C18 on a silica gel, a difunctional end-capping reagent like hexamethyldichlorotrisiloxane was used as a first end-capping, then trimethylchlorosilane was used as a second endcapping. These two end-capping reactions were done at over 200 degree Celsius under a nitrogen atmosphere. The C18 with the above end-capping was more hydrophobic andshowed a longer retention time higher than the C18 with the above end-capping done at 40 degree Celsius lower temperature although both C18s showed a good peak shapefor a basic compound. Conversion of silanol groups to siloxane bond by heating made hydrophobicity high. The peak shape of a basic compound and stability under both acidicand basic pH conditions were evaluated.

End-capping withhexamethyldichlorotrisiloxane and TMS on
C18 silica

Another end-capping with heating on C18silica, reduce of silanol groups
Typical no end-capping C18

Final TMS

C18

Comparison of 2 kinds of end-capping
Conventional end-capping

Silanol ActivityControl
Technology
Heating

Both Sunniest end-capping and Silanol Activity Control

O

Stationary phaseOSiOOOSiSiO

SiSiSi OO O

O O SiOC18

Si O

Si

Carbon loading and retention factoroX CO H

Non hydrated silanol group by influence of hydrophobicity of alkyl groups

O HHydrated silanol group without influence of alkyl groups

Relationship between retention factorand carbon loading(reaction at two different temperatures)

Retention
factor a(X-40) oC(more than 200 oC )Carbon loading of only C18

7.0%

7.0%

Carbon loading
after end-capping

7.3%

7.7%

(cut off some C18
chains by heating)

10.4

7.3

125

Ascentis Express C18, 2.7 μm

9.7

8.0

133

PoroShell C18 EC, 2.7 μm(not cut off C18
chain by heating)

Yes
10.6

SunShell C18, 2.6 μm

9.0Cortecs C18, 2.7 μm

No
9.5Accucore C18, 2.6 μm

8.5Kinetex C18, 2.6 μm

7.7

7.3

113

7.4

8.8

130

5.4

4.9

102

b. Measured after C18 packing material was pulled out of a column.
c. Measured using C18 materials sintered at 600 degree Celsius for 8 hours.

X C

Both Sunniest end-capping and Silanol Activity Control

8101214

16

18

20

22

SunShell C18 showed the largest retention factor althoughcarbon loading was not the highest.

24

Retention time/min

X oC was too high to keep carbon loading of initial C18. (X-40)
oC could keep carbon loading. As a result, carbon loading afterend-capping at (X-40) oC was higher than that at X oC.

It is considered that less residual silanol group madehydrophobicity of a stationary phase high, then maderetention factor large.

Regarding retention factor, reversed result was obtained. Lesssilanol group made hydrophobicity of stationary phase high.

Stability under acidic pH condition
▲

▲

▲

▲

Relative retention/%

8060

Durable test condition
Column size: 50 x 2.1 mm
Mobile phase: CH3CN/1.0% TFA,
pH1=10/90
Flow rate: 0.4 mL/min
Temperature: 80 ºCSunShell C18
Company S C18
Company W C1840

Company T C18
Company P C1820

Company A C1800204060Time/h80

100120Measurement condition
Column size: 50 x 2.1 mm
Mobile phase:
CH3CN/H2O=60/40
Flow rate: 0.4 mL/min
Temperature: 40 ºC
Sample: 1 = Uracil
2 = Butylbenzene

Stability under basic pH condition
Durable test condition
Column size: 50 x 2.1 mm
Mobile phase:
CH3OH/20mM Sodium
borate/10mM NaOH=30/21/49(pH10)
Flow rate: 0.4 mL/min
Temperature: 50 ºC

100

Relative plate of butylbenzene /%100N4=16,700, TF4=1.20 SunShell C18 2.6 μm
N4=12,800, TF4=1.89 Company A C18 2.7 μm
4

N4=5,200, TF4=2.61 Company P C18 2.6 μm

4

N4=5,300, TF4=2.73 Company T C18 2.6 μm
4

N4=4,800, TF4=3.12 Company W C18 2.7 μm
41234

567

8

N4=3,400, TF4=3.24 Company S C18 2.7 μm
910Retention time/min

Column dimension, 150 x 4.6 mm; mobile phase,acetonitrile/10mM ammonium acetate pH6.8 = 40/60;
flow rate, 1.0 mL/min; temp.: 40oC; sample, 1=uracil,
2=propranolol, 3=nortriptyline, 4=amitriptyline.

CH3NCH3Amitriptyline(X-40) oC

10% reduce
6240o4143135a. Retention factor of amylbenzene, mobile phase; methanol:water=75:25, 40 oC.

2Morehydrophobic

Comparison of amitriptyline peak

CarbonSpecific surface
loading b (%) area b, c (m2/g)*Base material: Core shell silica, 2.6 μm0C18

A basic compound shows no tailing on Sunrise C18-SAC
because hydrated silanol groups don’t make a basic
compound tailing as well as silica column on HILIC modeshows no tailing for a basic compound.

We named this end-capping method Sunniest end-capping.

Silanol activity control
Retention factor (k)C18

End-capping layer

End-capping reagent moves like a Geometrid caterpillar, so thata functional group on the tip of the arm can bond with a silanolgroup which Is located anywhere.

End-capping reaction
temperature

C18C18SiO
O OHO
SiSiSiOOO
Si OO
SiSiOOSiSiSiO O
HO OOOSiOOOOSiOSiOOOSiOOSiSiSiSiOSiO O
O SiO
Si O OOOOOOOOOOOSiOOO
SiSiC18

Change over tosiloxane bondSiSiC18

End-capping layer

HOEnd-capping
reagent

C18C18C18

Silanol groups withhigh absorption
activity

Stationary phase

Sunrise C18-SAC

80
SunShell C1860

Company S C18

Company T C18
Company A C18

20

0
0

1,000

2,000

3,000

4,000

Elution volume/mL

5,000Conclusion◆ Hexamethyldichlorotrisiloxane was used as an end-capping reagent for a first end-capping step. Thentrimethylchlorosilane (TMS) was used as an end-capping
reagent for a second end-capping step.
◆ Silanol groups were changed to siloxane bonding byheating on C18 silica.

Company P C1840SunShell C18 showed the highest theoretical plate (N) andthe lowest tailing factor. Highly-effective end-capping
including heat treatment (silanol activity control) reduced
tailing for a basic compound. As a consequence, highefficiency for a reversed column was achieved by highlyeffective end-capping.

6,000

Measurement condition
Column size: 50 x 2.1 mm
Mobile phase:
CH3OH/H2O=70/30
Flow rate: 0.4 mL/min
Temperature: 40 ºC
Sample: 1 = Butylbenzene◆ End-capping done by both bonding of a silylation agent

and conversion from silanol groups to siloxane bondleaded to not only high efficiency for basic compounds
but also increase in retention.
◆ Stability under both acidic and basic pH conditions

increased due to proposed end-capping.