Adsorption Phenomenon and Development of Low Adsorption
Vials for LC and LC/MS
Motoaki MURAKOSHI1, Kosuke FUKUZAWA 1, Yuki SATO1, Naoki ASAKAWA 2
1: R&D Group, Shimadzu GLC Ltd; 2: Shimadzu Corporation
1. Introduction
In recent years, LC/MS systems capable of high-sensitivity and high-selectivity analysis have become an essential means of analysis in many fields. As the concentration of samples used for LC/MS analysis have
become increasingly lower, there is concern that sample adsorption to containers could seriously compromise the reliability of analytical results. For glass containers, ionic adsorption, due to silanol groups on the
glass surface, and hydrophobic adsorption, due to siloxane, both occur at the same time. In contrast, only material-based hydrophobic adsorption occurs for polypropylene (PP) containers. Therefore, we verified
the phenomenon of adsorption to containers using various types of basic compounds and peptides (trypsin digested myoglobin and BSA) as models. The results showed that the adsorption phenomenon was
different for each type of containers (glass versus PP vials) and confirmed that the vial adsorption phenomenon was one of factors that compromised the reliability of analytical results obtained. This article
describes the adsorption behavior of newly developed low-adsorption HPLC vials with respect to basic compounds and peptides.
2. Current Status of Adsorption to Vials and
Methods for Inhibiting Adsorption
Basic compounds and other substances with a high acid dissociation constant (pKa) adsorb
readily to glass containers by ionic adsorption, whereas compounds with a large octanol-water
partition coefficient (logP) readily adsorb to both glass and PP containers by hydrophobic
adsorption. Therefore, ionic adsorption to silanol groups in the glass containers can be
effectively inhibited by adding a salt to sample solutions, and similarly hydrophobic adsorption
to glass or PP containers can be effectively inhibited by adding an organic solvent or surfactant.
However, in the case of high-sensitivity quantitative analysis using HPLC or LC/MS systems,
the effects of the additives on separation and ionization in the MS unit must be considered,
which can make it quite complicated to set sample preparation conditions optimized for
inhibiting container adsorption .
4. Effectiveness of LabTotal Vials in Inhibiting Basic Compound Adsorption
Four types of basic compounds were analyzed to compare the adsorption inhibition effects using LabTotal vials with commercial LCMS
vials and LC/GC vials. The resulting chromatograms are shown in Fig. 4, with corresponding area values (recovery rates) listed in
Table 1. The results confirmed that the LabTotal vials inhibited adsorption significantly compared to competing vials.
Table 1 Comparison of Basic Compound Recovery Rates for Three Types of Vials
LabTotal Vial
H Silanol
-Si-O
pKa
Ionic adsorption
➖
+
H +
logP
-Si
Basic
compound
O
-Si
Imipramine
Propranolol
64,990 (100 %)
32,249 (100 %)
Competitor Vial A (for LCMS)
45,376 (88 %)
7,620 (88 %)
55,531 (85 %)
31,496 (97 %)
21,788 (42 %)
6,137 (71 %)
24,131 (37 %)
27,327 (84 %)
mAU
20
mAU
LabTotal Vial
Competitor vial A
Competitor vial B
Amitriptyline
10
Inhibition of silanol dissociation
under acidic conditions
Glass Container
Add cations
-Si-O

Add nonionic
surfactant
1.5
Hydrophilic group
+
N
Hydrophobic
group
H
logP
-Si
Hydrophobic adsorption

O
-Si
0
0
pKa
Ionic adsorption
- Na +
1
Basic
compound
Hydrophobic
adsorption

PP Container
-Si-O
Atenolol
2
Aqueous
solution
H
LabTotal Vial
Competitor vial A
Competitor vial B
4
3
Hydrophobic
adsorption
Hydrophobic adsorption
Siloxane
Atenolol
8,638 (100 %)
Competitor Vial B (for LC/GC)
PP Container
Glass Container
-Si-O
Amitriptyline
51,376 (100 %)
mAU
30
LabTotal Vial
Competitor vial A
Competitor vial B
1.0
2.5 min
2.0
Imipramine
mAU
1.5
LabTotal Vial
Competitor vial A
Competitor vial B
20
2.0
min
Propranolol
20
Siloxane
10
Add organic solvents (CH3OH and CH3CN)
10
Fig. 1 Mechanism of Adsorption to Vials and
Methods for Inhibiting Adsorption
0
0
1.5
3. Development of Low-Adsorption Vials
2.0
2.5 min 0.0
0.5
1.0
min
Fig. 4 Chromatograms for Four Types of Basic Compounds (1 mg/L)
 Glass Vial (LabTotal Vial)
Due to the roughness of typical glass vial surfaces, there is a large surface area in contact
with samples. Therefore, the surface area was minimized by optimizing the molding
parameters during manufacture.
5. Effectiveness of TORAST-HTM Bio Vials in Inhibiting Peptide Adsorption
Surface area minimized with
optimized molding parameters
We confirmed that adsorption of highly polar peptides (with retention times between about 7 and 8 minutes) was most prominent on
glass vials, whereas adsorption of highly hydrophobic peptides (with retention times between about 12 and 16 minutes) was most
prominent on PP vials.
TORAST-HTM Bio vials showed effectiveness in inhibiting adsorption for both types of peptides.
Glass surface
Glass surface
LabTotal Vial
LSM Height image (3D)
LSM Height image (3D)
Height
:
1 µm
Height
:
1 m
43 m
43 m
43 m
43 m
AFM Height image (3D)
AFM Height image (3D)
uV
150000
-: TORAST-H Bio
140000 -: Glass
130000 -: Polypropylene (PP)
Adsorbed especially to PP and glass vials
120000
Height: 60.0
nm
5.0
m
Height: 10.0
nm
5.0
m
5.0
µm
110000
Adsorbed to glass vials
100000
Results from Observing Vial Surfaces with a Scanning Probe Microscope (SPM)
90000
Fig. 2 Overview of Low-Adsorption Glass Vials
80000
70000
60000
50000
 PP Vials (TORAST-HTM Bio Vials)
40000
A non-ionic superhydrophilic base was chemically bonded to the PP vial surfaces.
30000
20000
Surface
treatment
PP surface
Superhydrophilic polymer (non-ionic)
Chemical bond
PP surface
10000
0
-10000
-20000
5.0
6.0
7.0
8.0
9.0
10.0
11.0
12.0
13.0
14.0
15.0
16.0
TORAST-H™ Bio Vial
Fig. 3 Overview of Low-Adsorption PP Vials
Fig. 5 Chromatograms of Trypsin Digested Myoglobin (1.9 pmol/mL)
6. Summary and Discussion
We developed vials (LabTotal and TORAST-HTM Bio vials) with low adsorption characteristics required for high-sensitivity LC and LC/MS analysis.
1) The low-adsorption glass vials (LabTotal vials) inhibit the adsorption of basic compounds.
2) The low-adsorption PP vials (TORAST-HTM Bio vials) inhibit the adsorption of peptides.
These low-adsorption vials are expected to help ensure the reliability of high-sensitivity analysis.
17.0
18.0min