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YMC GENERAL CATALOG
1Reversed-PhaseC18 Columns (ODS)
Types and characteristics of C18packing materials
Guides for selecting C18 packing materials
YMC-Triart C18
YMC-UltraHT series YMC-UltraHT series YMC-UltraHT Pro C18 YMC-UltraHT Hydrosphere C18
Pro series Features of Pro series ODS columns YMC-Pack Pro C18 Hydrosphere C18 YMC-Pack Pro C18 RS
Types and characteristics of C18 packing materialsOctadecyl-silica (ODS) is the industry standard packing material for HPLC applications. YMC Co., Ltd. offers packing materials with an impressive assortment of functional groups for liquid chromatography and a selection of ODS columns that is far ahead of all others in variety. We also offer hybrid-silica based C18 packing materials that are widely applicable to a variety of fields, from microanalysis to large-scale isolation.
第01章.indd 14 11.1.28 10:39:04 AM
15
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
Low molecular weight compounds
High molecular weight compounds
Alkaline mobile phase
General organic compounds
Basic compounds
Coordination compounds
Sugars and glycosides
YMC-Triart C18YMC-Pack Pro C18YMC-Pack ODS-AJ’sphere ODS-M80
YMC-Pack Pro C18 RSJ’sphere ODS-H80
YMC-Pack ODS-A (Wide-Pore)J’sphere ODS-L80
Hydrosphere C18YMC-Pack Pro C18 RSYMC-Pack ODS-ALJ’sphere ODS-H80J’sphere ODS-L80YMC-Pack PolymerC18
YMC-Triart C18YMC-Pack Pro C18Hydrosphere C18YMC-Pack Pro C18 RSYMC-Pack ODS-AMYMC-Pack PolymerC18
YMC-Triart C18YMC-Pack Pro C18Hydrosphere C18YMC-Pack Pro C18 RSYMC-Pack ODS-AMYMC-Pack PolymerC18
Hydrosphere C18YMC-Pack ODS-AQJ’sphere ODS-L80
YMC-Pack ODS-A (Wide-Pore)
YMC-Triart C18YMC-Pack Pro C18 RSJ’sphere ODS-H80YMC-Pack PolymerC18
Chemical structures differ slightly among solutes
Hydrophobicity differs greatly among solutes
Difficult to separate using standard ODS
High molecular weight peptides and oligonucleic acids
Guides for selecting C18 packing materialsYMC Co., Ltd. offers twelve types of ODS columns, each with unique separation characteristics. The proper selection of packing material is therefore a key factor in the establishment of efficient separation conditions. Selection of packing materials without sufficient consideration of the separation characteristics may result in an expense of much time and effort in separating a target material.
第01章.indd 15 11.1.28 10:39:05 AM
16
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
Versatile Hybrid Silica Based HPLC Column
YMC-Triart C18YMC-Triart C18 is a hybrid silica based ODS column emphasizing durability, low operating pressure, and excellent performance.YMC-Triart C18 has been developed and is manufactured using state-of-the-art technologies that define materials, granulation and surface modification.
YMC-Triart C18 -Integration of triple artistic technologies-
<1>Versatile hybrid material
YMC-Triart C18 is a multi-layer hybrid particle consisting of two types of layers: a silica based layer (inorganic) and a hybrid polymer based layer (organic). This layered structure contributes to Triart's combined physical durability and chromatographic performance.
Image structurefor hybrid-silica gel
inorganic-silane layer
organic-silane layer
Particlesynthesis byMicroreaction
Particlesynthesis byMicroreaction
Versatilehybrid silica
material
Versatilehybrid silica
material
Multi-stageendcappingMulti-stageendcapping
Rugged, long lasting, hybrid particle technologyIntegration of triple artistic technologies
● Hybrid silica base ▼ Usable over a wide pH range makes the column excellent for method development.
● Great chemical durability ▼ Long column lifetime over wide pH range. Low cost per analysis.
● Lower back pressure ▼ Ability to use on both older and newer instrumentation.
● Excellent resolution ▼ Rugged, first choice option for analysis of acids, bases and neutral compounds.
Benefit
第01章.indd 16 11.1.28 10:39:08 AM
17
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
YMC-Triart C18 has a very narrow pore size distribution that enables narrow and reproducible peak shapes for analytes.
Narrow pore distribution
Silica particle for conventional product
350300250200Pore size (Å)
150100500
4500
4000
3500
3000
2500
2000
1500
1000
500
0
d V
p/d
logD
Hybrid particle for competitor's product
Hybrid particle for Triart
<2>New granulation process by microreactor technology
Homogeneous and uniform particle
Use of microreactor technology produces homogeneous and uniform particles that results in low operating pressure and very reproducible surface modification making for the ultimate in column-to-column and lot-to-lot reproducibility.
Triart C18
YMC-Triart C18 -Integration of triple artistic technologies-
Endcapping methods employed for production of conventional products typically employ a single endcapping compound in one chemical step to "neutralize" highly reactive silanols. Unfortunately, highly reactive silanols are easily hydrolyzed and contribute to poor durability. Additionally, low reactive silanols endcap stubbornly, producing a surface modification that is often non-reproducible, and leading to peak tailing. YMC-Triart C18 solves this problem by using an innovative, newly developed "measured multi-stage/multi-compound surface modification" technology which results in the thorough endcapping of all silanols that are accessible to analytes.
<3>Surface modification technology
Multi-stage endcapping
Reaction image for "multi-stage endcapping"
3STEP
Silanol group
Endcapping (EC) group
1STEP
2STEP
BeforeEC
Evaluation of silanol activity
12
3
Triart C18
Endcappingby traditional method
Peak tailing for basic compounds
min15.010.05.00.0
mAU
60
1
2
3
Q090928E
Q090928C
40
20
0
min15.010.05.00.0
mAU
60
40
20
0
第01章.indd 17 11.1.28 10:39:14 AM
18
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
Versatile Hybrid Silica Based HPLC Column
YMC-Triart C18
Quality Control -Excellent reproducibility-
1. Chlorpheniramine3. Propyl p-hydroxybenzoate
2. Dextromethorphan
Basic compounds
Lot. E
Lot. D
Lot. C
Lot. B
Lot. Amin17.515.012.510.07.55.02.50.0
13
2
J091201Y
YMC-Triart C18 exhibits excellent lot-to-lot reproducibility for all types of compounds including difficult to chromatograph molecules that exhibit basic and coordination compound behavior.
Packing Material
Rigid control of theoretical plate number (N) and tailing factor (Tf) are exhibited for each YMC-Triart C18 packed column.
Packed Column
Lot. E
Lot. D
Lot. C
Lot. B
Lot. Amin14.012.010.08.06.04.02.00.0
1 2
J091201X
1. Hinokitiol2. Methyl benzoate
20000
15000
10000
5000
0
2.5
2.0
1.5
1.0
0.510 20 30 40 50
Theoretical plate number
Tailing factor
Column number
Theo
retic
al p
late
num
ber
Taili
ng fa
ctor
Column :5 µm, 150×3.0 mmI.D.Eluent :20 mM KH2PO4-K2HPO4 (pH 6.9)/acetonitrile (65/35)Flow rate :0.425 mL/minTemperature :40 ℃Detection :UV at 235 nm
Low back-pressure -Free from pressure problem-Column pressure and solvents
YMC-Triart C18 is designed for use in all conditions. Even with higher viscosity methanol, YMC-Triart C18 can operate with lower pressure (typically 30% less than conventional columns).
Standard ODS -Best balance hydrophobicity-Comparison of separation selectivity
Conventional hybrid silica based ODS columns tend to be less hydrophobic than silica based columns. YMC-Triart C18 has a favorable carbon balance and may be used as a "versatile first-choice" method development column.
YMC-Triart C18 shows great chemical stability provided by newly developed hybrid silica. Even in high pH or high temperature conditions, the lifetime of YMC-Triart C18 is more than 10 times greater than conventional ODS columns.
YMC-Triart C18
Time (hr)
Rat
e of
the
oret
ical
pla
te n
umb
er
Silica based C18
Silica based C18alkaline-resistant type
0%
20%
40%
60%
80%
100%
120%
0 100 200 300 400 500 600 700 0 12010080604020
Time (hr)
Rat
e of
ret
entio
n tim
e
0%
20%
40%
60%
80%
100%
120%
YMC-Triart C18
Silica based C18
The columns are kept in acetonitrile/water/TFA (10/90/1), 70 °C,and tested for performance every 20 hours.
Test conditions Column :5 µm, 50×2.0 mmI.D. Eluent :acetonitrile/water (60/40) Flow rate :0.2 mL/min Temperature :37 ℃ Sample :butyl benzoate
Reduced column bleed -Operate at high sensitivity-Bleed test by LC/MS
No column
Triart C18Scan ES+TIC
0.0E+00
4.0E+08
8.0E+08
1.2E+09
min121086420
-
-
Bleeding of stationary phase can often contribute to high background signals when employing MS and other hyphenated detection techniques. In the example cited above, use of a 50×2.0 mmI.D. YMC-Triart C18 column shows no contribution to the signal observed by the MS positive total ion current (TIC) trace. Such performance allows for low limits of detection (high S/N ratio) for MS and other detection techniques. The inherent hybrid particle strength is also superior to conventional silica materials for detection techniques that are sensitive to particle degradation.
Column :5 µm, 50×2.0 mmI.D.Eluent : A) water/formic acid (100/0.1)
B) acetonitrile/formic acid (100/0.1) 5% B (0-1 min), 5-100% B (1-5 min), 100% B (5-10 min), 100-5% B (10-10.1 min), 5% B (10.1-12.5 min)
Excellent resolution -Without adsorption and tailing-Analysis of basic compounds
Hybrid-silica based C18Brand I8
5 µm, 150×4.6 mmI.D.
Silica based C18Brand D5
5 µm, 150×4.6 mmI.D.
Silica based C18Brand N1
3 µm, 150×3.0 mmI.D.
YMC-Triart C185 µm, 150×3.0 mmI.D.
0.0 2.5 5.0 7.5 10.0 12.5 15.0 17.5 min
21
3
21
3
2
1
3
2
13
Q090928C
Q090123F
Q090821C
J091111K
Column :150×3.0 mmI.D. or 150×4.6 mmI.D.Eluent :20 mM KH2PO4-K2HPO4 (pH 6.9)/acetonitrile (65/35)Flow rate : 0.425 mL/min for 3.0 mmI.D.
1.0 mL/min for 4.6 mmI.D.Temperature :40 ℃Detection :UV at 235 nm
Ingredients in a cough/cold medication
N
N
CH3
H3C
Cl
H3C
O
O
OH
O
NH
CH3
CH3
Chlorpheniramine
Propyl p-hydroxybenzoate
Dextromethorphan
1.
3.
2.
YMC-Triart C18's innovative surface modification technology results in excellent peak shapes even for the basic compounds that often exhibit tailing shapes on conventional silica and hybrid silica based ODS columns.
Hybrid-silica based C18Brand I8
5 µm, 150×4.6 mmI.D.
Silica based C18Brand N1
3 µm, 150×3.0 mmI.D.
YMC-Triart C185 µm, 150×3.0 mmI.D.
0.0 2.0 4.0 6.0 8.0 10.0 12.0 14.0 min
2
1
21
2
1
Q090925G
Q090120A
J091110B
Column :150×3.0 mmI.D. or 150×4.6 mmI.D.Eluent :acetonitrile/0.1% H3PO4 (40/60)Flow rate : 0.425 mL/min for 3.0 mmI.D.
1.0 mL/min for 4.6 mmI.D.Temperature :40 ℃Detection :UV at 254 nm
Hinokitiol
O OCH3
OH
CH3
CH3
O
Methyl benzoateHinokitiol
2.1.
YMC-Triart C18 has an extremely low level of metal impurities, much lower than conventional products.YMC-Triart C18 is able to provide excellent peak shape for coordination compounds.
Column :150×3.0 mmI.D. or 150×4.6 mmI.D.Eluent :acetonitrile/0.1% H3PO4 (5/95)Flow rate : 0.425 mL/min for 3.0 mmI.D.
1.0 mL/min for 4.6 mmI.D.Temperature :37 ℃Detection :UV at 210 nm
YMC-Triart C18 is synthesized utilizing methodology borrowed from microreactor technology. This synthesis technique allows for a reduction of impurities that contribute to peak tailing during the analysis of some types of acidic compounds.
10 mM phosphate buffer (pH 6.7)/organic solvent 10 mM CH3COONH4/organic solvent
Clemastine is a well known basic compound which can easily tail on conventional ODS columns. YMC-Triart C18 can separate clemastine sharply using many buffer/solvent combinations.
Clemastine
N O
Cl
H3CCH3
Column :5 µm, 50×2.0 mmI.D. or 50×2.1 mmI.D.Eluent : A) 10 mM KH2PO4-K2HPO4 (pH 6.7) or 10 mM CH3COONH4 (pH 6.7)
B) methanol or acetonitrile 5-90% B (0-10 min), 90% B (10-15 min)
Flow rate :0.2 mL/minTemperature :25 ℃Detection :UV at 230 nm
Improvement of loadability -Minimizing strong solvent/sample loading effects-Influence on peak shapes by injection volume
1
2
1 2
min1.5 2.0 2.5
Hybrid-silica based C18
Silica based C18
YMC-Triart C181
2
Peak anomaly at 1.5 µL
YMC-Triart C18 can tolerate larger injection volumes of samples containing strong solvents (e.g., acetonitrile) while allowing for better peak shape than traditional columns. This can be important for difficult to solubilize samples that need higher concentrations of organic solvent for solubilization prior to analysis.
Column :5 µm, 50×2.0 mmI.D. or 2.1 mmI.D.Eluent : A) water/formic acid (100/0.1)
B) acetonitrile/formic acid (100/0.1) 5% B(0-0.5 min), 5-100% B (0.5-2.5 min)
Flow rate :0.4 mL/minTemperature :40 ℃Detection :UV at 275 nm
Propranolol (50 µg/mL) Verapamil (50 µg/mL)
1. 2.OCH2CHCH2NHCH(CH3)2
H3CO
H3CO
CH(CH3)2
CN CH3OCH3
OCH3C (CH2)3NCH2CH2OH
acetonitrile 1.0 µL1.5 µL2.0 µL3.0 µL
solvent for sample injection volume
第01章.indd 21 11.1.28 10:39:27 AM
22
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
Versatile Hybrid Silica Based HPLC Column
YMC-Triart C18
YMC-Triart C18 3 µm, 50×2.0 mmI.D.
80% reduction in run time
5%B, 0.4 mL/min, 12.2 MPa
1
2
R091201B
mAU
25
20
15
10
5
0
0 1 2 3 4 5 min
Baseline separation,But long analysis time
5%B, 0.2 mL/min, 5.6 MPa
12
R091127C
mAU
25
20
15
10
5
0
0 5 10 15 20 25 min
YMC-Triart C18 5 µm, 150×2.0 mmI.D.
Fast analysis of paraquat and diquat
Insufficientseparation
10%B, 0.2 mL/min, 5.7 MPamAU
25
20
15
10
5
0
0 2 4 6 8 10 12 14 min
12
R091126J
YMC-Triart C18 5 µm, 150×2.0 mmI.D.
Eluent : A) water/HFBA* (100/0.1) B) acetonitrile/HFBA* (100/0.1)
Temperature :37 ℃Detection :UV at 290 nmInjection :1 µL (0.1 mg/mL)*heptafluorobutyric acid
Eluent : A) water/formic acid (90/10) B) acetonitrile/methanol/water/formic acid (22.5/22.5/40/10) 20-28% B (0-30 min), 28-70% B (30-40 min), 100% B (40-45 min)
Particle Size (µm) 2 2Pore Size (Å) 120 120Specific Surface area (m2/g) 330 330Carbon Content 16% 12%Bonding Type Monomeric MonomericAvailable pH Range 2 - 8 2 - 8Characteristics Standard type For polar analytes
2 µm ODS column ideal for Ultra-Fast LC
Features
● Superior column performance at higher flow rate and higher pressure● Reduced analysis time while maintaining excellent resolution● Excellent resolution with back pressure less than that of sub-2 µm● Applicable to both conventional HPLC and specific ultra pressure LC system● Same selectivity and superior peak shapes as 3 µm and 5 µm Pro series● Simple method transfer from conventional HPLC on Pro series; no need to
change elution conditions
YMC-UltraHTYMC-UltraHT series is a C18 bonded-phase based on highly efficient 2 µm spherical silica particles. YMC-UltraHT series columns are specifically designed for high-speed and high throughput analysis in various fields such as pharmaceuticals, foods, and environmentals.YMC-UltraHT series has two products. Pro C18 as our
standard ODS, and Hydrosphere C18 for hydrophilic compounds. Both of them show the same selectivity and superior peak shapes as 3 µm and 5 µm Pro series, allowing for an easy method transfer from conventional HPLC to ultra-fast HPLC without changing elution conditions.
第01章.indd 24 11.1.28 10:39:43 AM
25
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
Ultra Fast analysis for flavonoids
This example shows the method transfer from conventional HPLC method to Ultra-fast LC method using a YMC-UltraHT column. Since the column efficiency is not decreased even when the flow rate is 3 times as fast as the conventional condition, it can be shorten analysis time by 90% without losing the separation characteristics.
YMC-UltraHT series can be used for fast analysis with conventional LC system, as its pressure is lower than commercial sub-2 µm columns. Considering the extra column band broadening, the column of 3.0 mml.D. is more applicable for conventional LC system than the column of 2.0 mml.D..
Conventional LC system (8 µL, cell length 10 mm)
Conventional semi-micro LC system (2.5 µL, cell length 5 mm)
Column :YMC-UltraHT Pro C18Eluent :acetonitrile/20 mM KH2PO4
(10/90)Temperature :37 ℃Detection :UV at 260 nm
第01章.indd 25 11.1.28 10:39:45 AM
26
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
Analytical columns
YMC-UltraHT Pro C18● Superior column performance at higher flow rate and higher pressure● Excellent resolution with back pressure less than 60% of commercial sub-2 µm● Same selectivity and superior peak shapes as 3 µm and 5 µm Pro C18● Simple method transfer from conventional HPLC
F i r s t c h o i c e O D S c o l u m n f o r U l t r a - f a s t L CYMC-UltraHT Pro C18 is a C18 bonded-phase based on highly efficient 2 µm spherical silica particles. YMC-UltraHT Pro C18 is designed for high-speed and high throughput analysis, advanced endcapping technology
makes YMC-UltraHT Pro C18 highly appropriate for analysis of basic compounds. The physical properties and separation characteristics are strictly controlled, so it is highly suitable as the first choice column for ultra fast LC.
Eluent :A) acetonitrile/water/TFA (10/90/0.1) B) acetonitrile/water/TFA (35/65/0.1)Flow rate :0.2 mL/minTemperature :37 ℃Detection :UV at 220 nmSample :tryptic degest of Cytochrome c
Analysis of a tryptic digest of cytochrome C is shown below. A 50mm length UltraHT Pro C18 column of 2 µm particle can be shorten the analysis time by one-thirds compared to a 150 mm column packed with 5 µm Pro C18.
YMC-UltraHT Hydrosphere C18● Can be used with 100% aqueous as mobile phase● Superior column performance at higher flow rate and higher pressure● Reduced analysis time while maintaining excellent resolution
T h e b e s t c o l u m n f o r h y d r o p h i l i c c o m p o u n d sYMC-UltraHT Hydrosphere C18 is a C18 bonded phase based on highly efficient 2 µm spherical silica particles.
It is designed to maintain hydrohilicity on the packing surface for superior separation of hydrophilic compounds.
Examples illustrated shows the optimization process of isoflavones separation by 2 µm YMC-UltraHT Hydrosphere C18. It has the same selectivity as 3 µm Hydrosphere C18, so there is no need to change the condition at downsizing. Maintaining resolution, the analysis time can be reduced by using 2 µm packing material.
Features of Pro series ODS columnsPro series high performance reversed-phase columns feature advanced endcapping technology and quality assurance which is ready for validation. The Pro series includes three kinds of ODS, each with unique separation characteristics.
We provide products with excellent performance and stable quality all over the world to fulfill the needs of the pharmaceutical industry, in which production, as well as research and development, are increasingly globalized.
Providing products with stable quality all over the world
Feature
● Silica gel base, low in metal impurities● Processed with advanced endcapping
technology● Superior separation of basic compounds● Useful for LC/MS● Excellent durability● Excellent reproducibility● Two types of test reports are included
with each column
YMC-Pack Pro C18 RS is a high
carbon "polymeric type" ODS bonding,
characterized by high resolution and
high durability. It is applicable to a
wide range of compounds. The
separation of compounds with relatively
high hydrophobicity and those that differ
only slightly in hydrophobicity, such as
structural isomers, is outstanding. It
also has excellent resistance to acid
and alkali, making it useful under
demanding separation conditions.
High carbon ODS column, useful for separation of highly hydrophobic compounds
YMC-Pack Pro C18 RS
YMC-Pack Pro C18 is a high
performance monomeric type ODS
column providing standard
hydrophobicity, high resolution, high
durability and excellent reproducibility.
It is highly versatile and suitable as the
first choice column in almost all fields,
including pharmaceutical products,
agricultural chemicals, foods, and
natural products.
Standard ODS columnwith high versatility
YMC-Pack Pro C18
Hydrosphere C18 is an ODS column
designed to maintain maximum
hydrophilicity without activating the
silica surface. It can be used with
100% water eluent, a problematic
eluent for conventional ODS columns.
Hydrosphere C18 is useful for
separating hydrophilic compounds.
Low carbon ODS column, useful for separation of hydrophilic compounds
Hydrosphere C18
Types of Pro series ODSPro C18 Hydrosphere C18 Pro C18 RS
Particle Size (µm) 2 , 3 , 5 , 10 2 , 3 , 5 3 , 5Pore Size (Å) 120 120 80Specific Surface area (m2/g) 330 330 510Carbon Content 16% 12% 22%Bonding type monomeric monomeric polymericAvailable pH Range 2 - 8 2 - 8 1 - 10Characteristics Standard type Separation for hydrophilic compounds High durability
第01章.indd 28 11.1.28 10:40:32 AM
29
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
The three kinds of ODS columns in the Pro series each have unique carbon content and separation characteristics, including hydrophobicity, plane recognition ability, and hydrogen-bonding capacity. Selection according to the analyte and mobile phase of Pro C18 with standard hydrophobicity, Pro C18 RS with high hydrophobicity, or Hydrosphere C18 with low hydrophobicity, facilitates method development.
Three kinds of ODS, each with different separation characteristics
Hydrophobicity and plane recognition ability of Pro series ODS
3 types of ODS are available to separate the para-benzoates shown in the left, but the best separation between peaks 5 and 6 is achieved on Pro C18 RS.
1
2
3 4
65
10 20 300 min(J020827E)
Pro C18 RS
1
2
3 4
65
10 200 min(J020827D)
Pro C18
1
2
3 4
65
10 200 min(J020827C)
Hydrosphere C18
Methyl p-hydroxybenzoate
OH
COOCH3
Ethyl p-hydroxybenzoate
Propyl p-hydroxybenzoate
Butyl p-hydroxybenzoate
Isopropyl p-hydroxybenzoate
Isobutyl p-hydroxybenzoate
COOCH
CH3
CH3
OH
OH
COOCH2CH3
COO(CH2)2CH3
OH
COOCH2CH
CH3
CH3
OHCOO(CH2)3CH3
OH
1.
2.
3.
4.
5.
6.
Flavonoid glycosides
Pro C18 RS
Pro C18
Hydrosphere C18
5 10 150 min
1
2
3
J020828F
4
1
2
3
4
J020828A
1
2
3
4
J020828C
O
O
HO
OH
HO
OR
HO
Hydrogen-bonding interactions significantly contribute to the retention of hydrophilic compounds, including glycosides. Therefore, the retention is longer in the Hydrosphere C18, which has strong hydrogen-bonding capacity, than in Pro C18 RS, which has highly hydrophobic packing materials. To separate such hydrophilic compounds, the Hydrosphere C18 is suitable for precise separation of multicomponent samples, while Pro C18 RS and Pro C18 are suitable for rapid separation of simple components.
Column :150×4.6 mmI.D.Eluent :acetonitrile/10 mM H3PO4 (25/75)Flow rate :1.0 mL/minTemperature :40 ℃Detection :UV at 260 nm
1. Rutin2. Isoquercitrin3. Quercitrin4. Quercetin
R = rutinoseR = glucoseR = rhamnoseR = H
第01章.indd 30 11.1.28 10:40:39 AM
31
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
Terphenyl isomers
1
2
3
10 200 minJ020719E
Pro C18 RS
1.
2.
3.
m-Terphenyl
o-Terphenyl
p-Terphenyl
Since Pro C18 RS has excellent hydrophobic selectivity and shape selectivity, it works well in the separation of structural isomers that are not easily separated by other ODS.
Peaks 2 and 3, which are not easily separated by other ODS, can be completely separated by Hydrosphere C18. It is thought that hydrogen-bonding interaction of hydrophilic functional groups in sugar chain and terpene skeleton as well as hydrophobic interaction has great contribution to the separation of saikosaponins.Hydrosphere C18 is the most appropriate in the separation of this kind of class of compound.
● Excellent reproducibility● Utilizes highly pure silica gel base● Two types of test reports are included
with each column
P e r f e c t l y e n d c a p p e d O D SYMC-Pack Pro C18 includes more advanced endcapping technology for strictly controlled processing of residual silanol groups that are likely to affect quality. Pro C18 is highly appropriate for basic compounds, including pharmaceutical products and agricultural chemicals.
Two test reports are included with each product; a Column Test Report, indicating the column performance, and a Packing Material Test Report, detailing strictly controlled physical properties and separation characteristics of the packing material.
Column :YMC-Pack Pro C18 (5 µm, 120 Å) 150×4.6 mmI.D.Eluent :20 mM KH2PO4-K2HPO4 (pH6.9)/ methanol (25/75)Flow rate :1.0 mL/minTemperature :37 ℃Detection :UV at 254 nm
Tricyclic antidepressants
(A961015G)0 10 20 min
1
2 3
4Advanced endcapping technology results in good peak shapes even for basic compounds that often elute with poor tailing shapes on competitive columns.
CHCH2CH2NCH3
CH3
HCI・N
CH2CH2CH2N
CH2CH2CH2NHCH3
CH3
CH3
HCI・
NHCI・
1.
Desipramine hydrochloride
2.
Acenaphthene(I.S.)
3.
Imipramine hydrochloride
4.
Amitriptyline hydrochloride
Basic peptides
(A990123B)
1
23
4
56
0 5 10 15 20 min
Pro C18YMC-Pack Pro C18 is processed using advanced endcapping technology and produces a better chromatogram than that obtained using ODS-A for separating basic peptides.Similar chromatograms have been obtained with YMC-Pack Pro C18 and ODS-A for separation of neutral peptides.
(A990121A)
1
2
3
4
56
0 5 10 15 20 min
ODS-A
Neutral peptides
(A990121B)0 5 10 15 20 min
1
2
3 45
6
Pro C18
(A990119B)0 5 10 15 20 min
1
2
34
56
ODS-A
1. Bradykinin2. Dynorphin A (Human,1-13)3. Eledoisin related peptide4. Neurotensin5. Bombesin6. Substance P
Column :YMC-Pack Pro C18 and ODS-A (5 µm, 120 Å) 75×4.6 mmI.D.Eluent :A) water/TFA (100/0.1) B) acetonitrile/TFA (100/0.1) 20-40% B (0-20 min,linear)Flow rate :1.0 mL/minTemperature :37 ℃Detection :UV at 220 nm
A three-lot comparison is shown on the left.Excellent reproducibility is achieved in separating hydrophilic, basic and acidic compounds, as well as hydrophobic compounds.
Advanced packing technology results in high column efficiency. All packed columns are inspected and controlled according to the strictest quality control criteria for column performance ever, including theoretical plate number testing.
24000
21000
18000
15000
12000
9000
6000
300020 40 60 80 100
Column No.
Theoretical plate number
150×4.6 mml.D. theoretical plate number
Two types of test reports〈Packing Material Test Report〉
Each YMC-Pack Pro C18 includes both, a Column Test Report and a Packing Material Test Report, to certify lot-to-lot and column-to-column reproducibility.
〈Column Test Report〉
This report describes basic physical properties of silica gel, including particle size, pore size and metal content, carbon loading after modification, and the specifications and results of separation characteristics tests.
The Column Test Report shows the theoretical plate number and asymmetry factor to indicate column performance. For convenience in deterioration evaluation, this report includes all analytical conditions including sample concentration as well.
第01章.indd 33 11.1.28 10:40:51 AM
34
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
Analytical columns
YMC-Pack Pro C18
1
2
3 4 5
6
min0 5 10 15 20
F040119B
Pro C18
Column :150×4.6 mmI.D.Eluent :A) 20 mM KH2PO4-H3PO4 (pH 2.5) B) methanol 20-90% B (0-15 min), 90% B (15-20 min)Flow rate :1.0 mL/minTemperature :37 ℃Detection :UV at 260 nm
A guard cartridge pack includes a cartridge holder (1 set), a guard cartridge column, and a column coupler (produced by PEEK).
Guard columnsS-5 120 10×30 AS12S05-0310WTG
20×50 AS12S05-0520WTG
S-10 120 20×50 AS12S11-0520WTG
When columns packed with material from different lots are needed for validation purposes, please contact us.
Ordering information
第01章.indd 35 11.1.28 10:40:56 AM
36
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
Analytical columns
Hydrosphere C18● Strong retention of hydrophilic compounds● Can be used with 100% water mobile phase● Superior separation of basic compounds● Excellent reproducibility
● Utilizes highly pure silica gel base● Two types of test reports are included with
each column
H y d r o p h i l i c O D SHydrosphere C18 is designed to maintain adequate hydrophilicity on the packing surface for superior separation of hydrophilic compounds. Hydrophilic compounds are retained with much greater affinity than with conventional ODS columns, permitting the product to be used
with 100% water eluent, a problematic eluent for conventional ODS columns. Hydrosphere C18 is useful for separating a wide range of compounds, including nucleic acids and their derivatives, organic acids, saccharides, glycosides and peptides.
Appropriate for separation of hydrophilic compoundsSeparation of organic acids
0 5 10 minA000310E
1
2
34
5
Hydrosphere C18High hydrophilicity of the Hydrosphere C18 packing material surface enables longer retention and superior separation of citric acid and fumaric acid as compared to a conventional ODS column (Brand Ⅰ).
Column :150×4.6 mm I.D.Eluent :20 mM NaH2PO4-H3PO4 (pH 2.8)Flow rate :1.0 mL/minTemperature :30 ℃Detection :UV at 210 nm
Reproducibility of retention time when used with 100% water mobile phase
S0515A10 200 min
※
21
3
4 5
Hydrosphere C18When conventional ODS columns are used with 100% water mobile phase, the apparent hydrophobicity decreases due to repulsive interactions between water and hydrophobic groups on the surface of the packing materials, reducing retention of compounds. In the experiment illustrated here, columns filled with 100% water mobile phase were left to stand overnight. Retention times of nucleic acid bases were analyzed before and after standing and compared. The retention times of adenine after standing decreased to 89% of the original value for Brand II and to 71% of the original Brand III value. By contrast, Hydrosphere C18 retained 98% of its original retention for adenine. Hydrosphere C18 is designed to maintain adequate hydrophilicity on the packing surface so that hydration can be achieved, and when Hydrosphere C18 is used with mobile phase containing no organic solvent, the retention time is not significantly shortened and highly reproducible chromatograms can be obtained.
Column :150×4.6 mm I.D.Eluent :20 mM KH2PO4-K2HPO4 (pH 6.9)Flow rate :1.0 mL/minTemperature :37 ℃Detection :UV at 254 nm
Afte
r sta
ndin
gBe
fore
sta
ndin
g
After completion of analysis, the flow was stopped and the columns were left to stand overnight (ca. 15 hours).
第01章.indd 36 11.1.28 10:41:01 AM
37
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
Differences in separation characteristics from conventional ODS columns
0 10 20 30 40 minS000309A
1
2
34
5 6
Hydrosphere C18The chromatograms shown were obtained in tests conducted to compare separation characteristics using a mixed sample containing compounds with different characteristics. The packing material surface of Hydrosphere C18 has higher hydrophilicity and lower C18 ligand density than those of Pro C18. This results in shorter retention of hydrophobic compounds, including n-propylbenzene and n-butylbenzene, and planar compounds like acenaphthene. By contrast, retention of testosterone, a compound with large hydrogen-bonding properties, is longer on Hydrosphere C18 and on Brand IV, which also has a highly hydrophilic packing surface.Amitriptyline, a basic compound, caused tailing peaks in Brand IV and Brand V. By contrast, Hydrosphere C18 and Pro C18 produce good peak shapes for such basic compounds. Although the retention times of amitriptyline on Hydrosphere C18 and Pro 18 are similar, there is a big difference between the two columns with regard to resolution of n-butylbenzene and amitriptyline. This difference is considered to be due to the hydrophilicity of the Hydrosphere C18 surface.These results demonstrate that Hydrosphere C18, having different separation characteristics from conventional ODS columns, enables separation of compounds that cannot be separated using other ODS columns.0 10 20 30 40 min
A guard cartridge pack includes a cartridge holder (1 set), a guard cartridge column, and a column coupler (produced by PEEK).
Guard columnsS-5 120 10×30 HS12S05-0310WTG
20×50 HS12S05-0520WTG
Ordering information
第01章.indd 39 11.1.28 10:41:08 AM
40
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
Analytical columns
YMC-Pack Pro C18 RS● Excellent acid resistance and alkali
resistance (pH 1 to 10)● Superior separation of structural isomers● Superior separation of basic compounds
● Excellent reproducibility● Utilizes highly pure silica gel base● Two types of test reports are included
with each column
H i g h l y d u r a b l e O D SYMC-Pack Pro C18 RS is a polymeric type high carbon ODS column characterized by high resolution and high durability. It is applicable to a wide range of compounds, providing good separation of basic compounds that easily cause
tailing peaks. The separation selectivity for compounds that differ only slightly in hydrophobicity is outstanding. It also has excellent resistance to acid and alkali, making it useful under demanding separation conditions.
YMC-Pack Pro C18 RSAcid-resistant column(manufactured by other company)
Monomeric ODS
Hours exposure to acetonitrile/water/TFA (10/90/1), change in packing material hydrophobicity at 70 ℃.
%
Retention
of
butyl
benzoate
The Pro C18 RS packing material hydrophobicity is hardly changed under the harsh conditions of pH1 and column temperature 70 ℃.
pH 1
0
20
40
60
80
100
0 200 400 600 800(hours)
Pro C18 RSMonomeric ODS
Sequential analysis using 20 mM H3BO3-NaOH (pH 9.8)/methanol (50/50)
%
Theoretical
Plate
Number (
N)
of
methyl
benzoate.
Unlike conventional ODS columns, Pro C18 RS can be used with an alkali mobile phase.
pH 10
Pro C18 RS with excellent acid and alkali resistance can be used over a wide pH range, and it is easy to optimize the separating conditions. In addition, by downsizing from the column (particle 5 µm, length 150 mm) to a shorter column (particle 3 µm, length 50 mm), with high separation ability, it becomes possible to shorten the analysis time by one third without affecting the separation.
Column :YMC-Pack Pro C18 RSEluent :20 mM acetate buffer/methanol (35/65)Flow rate :1.0 mL/minTemperature :30 ℃Detection :UV at 220 nm
A wide choice of mobile phases facilitates method development
0 5 10 15 20 min 0 5 10 15 20 min F040714EF040716A
3,4,5
1
2 21
3
5
4
0 5 10 15 20 minF040714C
21
3
5
4
150×4.6 mmI.D. 5 µm
pH 3.2 pH 6.8 pH 8.8
0 5 10 minF040714D
21 3
5
4
50×4.6 mmI.D. 3 µm
pH 8.8
Downsizing
第01章.indd 40 11.1.28 10:41:12 AM
41
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
Separation of basic compounds
Amitriptyline is a basic compound that can easily tail on some silica based columns. In comparison with competing acid-resistant columns, Pro C18 RS produces good peak shapes without tailing.
Tf(4) 1.16
10 200 minJ011114B
1
2
3 4
YMC-Pack Pro C18 RS
Tf(4) 2.87
Acid-resistant column manufacturedby other company(Brand D3)
Column :150×4.6 mmI.D.Eluent :20 mM KH2PO4-K2HPO4 (pH 6.9) /methanol (25/75)Flow rate :1.0 mL/minTemperature :37 ℃Detection :UV at 254 nm
Separation of structural isomers
Pro C18 RS with a high C18 density is excellent in hydrophobicity recognition ability and plane recognition ability, and can be applied to separate structural isomers which are difficult to separate with other ODS. In addition, by using 3 µm column packing, resolution can be increased, and it becomes possible to achieve a baseline separation of linolenate isomers.
A guard cartridge pack includes a cartridge holder (1 set), a guard cartridge column, and a column coupler (produced by PEEK).
Guard columnsS-5 80 10×30 RS08S05-0310WTG
20×50 RS08S05-0520WTG
Ordering information
第01章.indd 43 11.1.28 10:41:20 AM
44
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
Analytical columns
YMC-Pack ODS-A● Conventional ODS column● Currently in use worldwide
S t a n d a r d O D SYMC-Pack ODS-A has a highly endcapped surface structure and appropriate hydrophobicity for separation of a wide range of compounds. It is produced under strict quality control with respect to 50 or more parameters in
order to ensure stable quality.This product is highly regarded as the standard YMC-Pack packing material in various countries around the world.
The graph indicates lot-to-lot reproducibility with respect to hydrophobic interaction for 10 lots, an essential column performance characteristic. Extremely stable lot-to-lot reproducibility of the hydrophobic surface is achieved by strict control of variables.In addition to measurement of the physical properties, a wide range of compounds including hydrophobic, acidic, basic and coordination compounds are analyzed under optimal conditions to evaluate column performance.
YMC-Pack ODS-AM● Similar to ODS-A in selectivity● Excellent reproducibility● Useful for quality control purposes
O D S w i t h o u t s t a n d i n g l o t - t o - l o t r e p r o d u c i b i l i t yYMC-Pack ODS-AM is a product which especially emphasizes lot-to-lot reproducibility of the packing material. Strict control is performed on all aspects of quality, including physical properties of silica gel base
and surface modifying processes, in order to ensure stable quality. Double endcapping is conducted to ensure that lot-to-lot reproducibility is maintained.
Special attention is given to lot-to-lot reproducibility
α (M
ethy
lpar
ben
/2,6
-Dim
ethy
lpyr
idin
e)
10
8
6
4
2
01 2 3 4 5 6 7 8 9 10
Lot Number
The separation factor (α) of methylparaben/2,6-dimethylpyridine for 10 lots of ODS-AM packing material is plotted on the left graph. Excellent lot-to-lot reproducibility is achieved even in separation of basic compounds.
Secondary interaction of the stationary phase surface may affect peak shapes of acidic compounds.ODS-AM can provide chromatograms without tailing peaks as shown above.
Coordination compounds
ODS with a high degree ofmetal impurities
ODS-AM
1
23 1
2
3
1. Uracil2. Acetylacetone3. 8-Quinolinol
The peak shapes of coordination compounds, including 8-quinolinol and acetylacetone, may be affected by the presence of metal impurities on the surface of the silica. The distribution level of metal impurities on the column is strictly controlled for ODS-AM, providing excellent peak symmetry.
YMC-Pack ODS-AQ● Useful for separation of hydrophilic compounds● Can be used with 100% water in mobile phase● Different selectivity from conventional ODS
H y d r o p h i l i c O D SYMC-Pack ODS-AQ has moderate hydrophobicity and hydrogen-bonding capacity. It shows different retention behavior from that of YMC-Pack ODS-A, for samples with relatively high hydrophilicity.
It is useful in fields including carbohydrate chemistry for oligosaccharides and glycosides, pharmacognosy and natural product chemistry.
Chromatograms of purine alkaloids are shown on the left.As compared with ODS-A, ODS-AQ and ODS-AL show longer retention of caffeine and theophylline relative to phenol, an internal standard.
Change of separation factor (α) of caffeine/phenol according to temperature change in ODS-AQ, ODS-A and ODS-AL is plotted on the graph shown on the left. Although caffeine is considered to have stronger hydrogen-bonding properties than phenol, the hydrogen-bonding capacity of stationary phase may affect the α of caffeine/phenol. The smaller the hydrogen-bonding capacities are, the higher α of phenol/caffeine. ODS-AQ shows different retention behavior from that of ODS-A since it has moderate hydrogen-bonding capacity.
YMC-Pack ODS-AL● ODS with residual silanol groups● Utilizes secondary interaction caused by silanol groups
N o n - e n d c a p p e d O D SYMC-Pack ODS-AL uses not only hydrophobic interaction but also secondary interaction caused by silanol groups that affect separation. This results in a different selectivity from conventional ODS columns.
When ionic interaction is utilized, it is preferable to use a buffer in the mobile phase to achieve reproducibility of chromatograms.
The separation factor (α) of internal standards methylparaben/2,6-dimethylpyridine for ODS-AM is different from that of ODS-AL, which is not endcapped. ODS-AL, which is affected by residual silanol, shows relatively great retention of pyridines.
The figure shows separation of disinfectants under the conditions described in the Japanese Pharmacopeia. The object is to select the column permitting the elution of benzoic acid, salicylic acid and theophylline, in this order, insuring complete separation of these peaks. With ODS-AQ, separation of salicylic acid and theophylline is incomplete. By contrast, ODS-AL provides excellent separation. Thus, ODS-AL may provide excellent separation when the separation conditions cannot be optimized on other ODS columns.
ODS-AQ
4
2,3
1
0 5 10 15 20 min
Column :YMC-Pack ODS-AL and ODS-AQ (5 µm, 120 Å) 150×4.6 mmI.D.Eluent :100 mM KH2PO4-Na2HPO4 (pH 7.0)/methanol (75/25)Flow rate :0.8 mL/minTemperature :30 ℃Detection :UV at 270 nm(Conditions described in Japanese Pharmacopoeia 15th ed.)
PhenolTheophylline (I.S.)
4.3.
OH
Salicylic acid
2.1. COOH
OHBenzoic acid
COOH
HO
O
CH
CH
N N
NN3
3
Disinfectants
第01章.indd 50 11.1.28 10:41:35 AM
51
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
0 10 20 30 40 min
1
3
2Phenacyl esters of bile acids from pig gall1. Cholic acid (CA)2. Hyodeoxycholic acid (HDCA)3. Chenodeoxycholic acid (CDAC)
J’sphere ODS-H80, ODS-M80, ODS-L80● Useful for method development● High theoretical plate number
T h r e e t y p e s o f O D S , e a c h w i t h d i f f e r e n t l i g a n d c o v e r a g eJ’sphere offers a choice from three kinds of ODS made from the same silica gel base, each with different ligand coverage.Differences in ligand coverage considerably affect the hydrophobic retention behavior of solutes, as well as the separation behavior
resulting from solute functional groups or tertiary structure.J’sphere is useful for the optimization of separation conditions, since there is almost no need to consider interactions other than hydrophobic and hydrogen-bonding interactions (e.g., ionic or coordinate interaction).
ODS columns with different ligand coverage have different hydrophobicity, hydrogen bonding capacity and steric selectivity for tertiary structures. ODS-H80 has exceptionally high hydrophobicity and low hydrogen bonding capacity, ODS-M80 has moderate hydrophobicity and hydrogen bonding capacity and ODS-L80 has low hydrophobicity, comparable to that of C8 and high hydrogen bonding capacity. J’sphere can be used with confidence to identify structural differences of solutes and is useful for improving the efficiency of separation optimization.
ODS-H80 ODS-M80 ODS-L80
Image of stationary phase
第01章.indd 52 11.1.28 10:41:39 AM
53
1
Reversed-P
hase C1
8 C
olumns
(OD
S
)
Analytical columnsPacking material
Particle size(µm)
Pore size(Å)
Column sizeinner diameter×length(mm) Product number
YMC-Pack PolymerC18● Utilizes polymer base which is not affected by silanol● Different separation characteristics from silica-based ODS● Excellent pH stability
P o l y m e r t y p e C18PolymerC18 is a C18 column made from methacrylate polymer. It has excellent pH stability and it is useful for separation of basic compounds because there are no silanol or metal impurities to cause secondary interaction.
Since π electrons of the carbonyl group or the hydroxyl group on the surface of the base material show particular interactions with solutes, PolymerC18 can show different separation characteristics from silica-based ODS.