A parylene-MEMS ion-exchange Liquid Chromatography (LC) chip is presented here. The chip is integrated with microfluidic I/O ports, a separation column, frits/filters, and a conductivity detector. The column is packed with conventional LC stationary phase support materials, i.e. micro-beads with surface functional groups. To withstand high pressure normally encountered in high performance liquid chromatography (HPLC), a self-aligned, channel-anchoring technique is developed to increase the pressure rating of the parylene microfluidic devices from 30 to at least 800psi. On-chip injection, separation and detection of anions in water, with ~25 ppm concentration, have been successfully demonstrated. To our knowledge, this is the first demonstration of microbeads-packed column ion liquid Chromatography (LC) on a chip

He, Qing

Lee, Terry D.

Pang, Changlin

Tai, Yu-Chong

Caltech Authors - Main

ION LIQUID CHROMATOGRAPHY ON-A-CHIP WITH 
BEADS-PACKED PARYLENE COLUMN 
Qing He, 'Changlin Pang, 'Yu-Chong Tai, and 2Tervy D. Lee 
'Caltech Micromachining Lab, California Institute of Technology, CA, USA 
*Beckman Research Institute, City-of-Hope, CA, USA 
I 
ABSTRACT 
A parylene-MEMS ion-exchange Liquid Chromatography 
(LC) chip is presented here. The chip is integrated with 
microfluidic U0 ports, a separation column, fritsifilters, and 
a conductivity detector. The column is packed with 
conventional LC stationary phase support materials, i.e. 
micro-beads with surface functional groups. To withstand 
high pressure normally encountered in High Performance 
Liquid Chromatography (HPLC), a self-aligned, channel- 
anchoring technique is developed to increase the pressure 
rating of the Parylene microfluidic devices from 30 to at 
least 8OOpsi. On-chip injection, separation and detection of 
anions in water, with -25ppm concentration, have been 
successfully demonstrated. To our knowledge, this is the first 
demonstration of microbeads-packed column ion liquid 
chromatography (LC) on a chip. 
1. INTRODUCTION 
High Performance Liquid Chromatography (HPLC) is one of 
the most popular, powerful, and versatile separation 
techniques for chemical analysis. Although LC column is 
normally made of capillary tubes due to fluidics limitations, 
the miniaturization of the column can actually improve 
separation performance. As shown in Fig. 1 [l] where same 
separation chemistry applies, separated peak width is 
independent of column ID, while peak heights are larger for 
smaller columns and/or smaller beads. In short, further 
down-sizing could be favorable in liquid chromatography. 
3- A 
4 +- .,.: .... ........ :.:;: ._...... ;..:.:.\-. . .... , I. . .. .""* ,. .. 1 
Injection Column Peak in Eluate 
Figure 1. Separated peak shapes obtained with different 
column and bead size. 
Compared to electrophoresis, however, little has been 
published about the integration of HPLC system onto a 
0-7803-826S-X/04/$17.00 02004 IEEE. 212 
single chip. The two main reasons are the lacks of: (1) a 
process to integrate various components for an LC system; (2) 
high-pressure compatibility of the microfluidic devices 
needed for running HPLC. This work deals with both 
problems. 
In terms of introduction and retaining LC stationary 
phase materials into the column, several approaches have 
been proposed, including open-tubular [2], coating 
micromachined posts arrays [3], coating microchannel with 
nanoparticles [4], monolith [5], packing [6], and packing 
without frit [7]. In this work, packing with conventional 
beads is chosen because of the following advantages. First, 
without introducing new surface chemistry, the extensively 
established separation knowledge can he utilized. Secondly, 
extreme flexibility is there to perform different types of 
liquid chromatography and/or to optimize particular 
separations, by choosing head type, size, pore size, porosity, 
and functional group. Thirdly, packed column can achieve 
reproducible column performance, which is usually a 
problem in other methods. 
2. DESIGN AND FABRICATION 
The LC-on-a-chip device is made using parylene MEMS 
technology. The chip has four access ports, as shown in Fig. 
2. LC mobile phase flows from port #I to #2. Nanolitre 
volume sample injection is achieved with a cross channel 
injection method [SI from #3 to #4. Beads are packed 
externally into the on-chip column from port # I .  Filters 
(channel with a height smaller than the bead size) trap beads 
in the main channel and prevent beads from entering the side 
injection channels. The bead-packed column inlet is near the 
sample output port #4, which is the injected sample plug 
front. The column outlet is at the filterisensor edge (Fig. 
2(b)). This integrated design minimizes dead volume, thus 
minimizing extra-column peak broadening. Off-chip pumps 
and valves are used in testing. 
A conductivity sensor is used as the detector for ion 
sensing. Interdigital electrodes are patterned in the detector 
cell to monitor liquid conductivity. The Conductivity of the 
mobile phase solution provides a baseline signal. When 
separated ion plugs pass by the detector, changes of solution 
conductivity are detected. Chromatogram is then obtained by 
recording conductivity of the solution flowing in the 
detection cell over time. To increase sensitivity, electrode 
width and spacing should be small, and the total electrode 
area should be maximized. However, the electrode design 
should be compromised in considerations of minimizing 
peak broadening and multiple-peak detection. 
Figure 2. (a) Fluorescent overview picture of the integrated 
separation system. The channels arefilled with Fluorescein; 
(b) Opticalpicture afterpacking beads. 
Etch Backside Holes with D R I E  
Pattern Frontride Oxide  
Pattern Two-Level Photoresist Structure 
Etch Oxide using Patterned Photoresist IS Mark 
Etch Special Trench with DRIE 
Deposit  and Pattern Parylene 
&?T* E l  
DRIE Open Backside Holes 
Acetone Dissolves Photoresist 
n - -  - I 
Si  Oxide  Pl lTi  PhotoreSiit Parylrnr 
Figure 3. Simplified fabricationflow. 
The fabrication process (Fig. 3) starts with growing 
thermal oxide on Silicon wafer. Then a two-step DRIE is 
performed on wafer backside to create the backside holes, 
leaving only a SOpm-thick diaphragm. Following frontside 
oxide patterning, a Ti/Pt/Au ( 3 0 0 ~ 2 0 0 0 8 J 1 0 0 0 ~ )  layer is 
evaporated and patterned. Au is left only at bonding pads. Pt 
is used as the sensing electrode and is patterned to be 
interdigital Spm-wide electrodes at Spm intervals. 25pm- 
thick photoresist is spun and patterned with two masks to 
form two-level sacrificial photoresist structure. The 2 5 p -  
high channel part uses unexposed photoresist, while the 
4pm-high filter part is partially exposed. A 15pm-wide 
“moat”, self-aligned-to channels and filters, is fully exposed 
to the bottom oxide, which is subsequently etched away. 
Modified DRIE process [9] is then done to make 40pm-deep 
trench with mushroom bottom profile in the moat, or XeFl is 
used to roughen the moat area. About l0pm-thick parylene 
is conformally deposited filling up the trench or the 
roughened moat and is patterned. Backside holes are open 
with DRIE. Acetone dissolves photoresist and releases the 
device. 
In the above process, a self-aligned, channel-anchoring 
technique is used to increase the pressure rating of the 
parylene chip. Figure 4 shows cross-sectional pictures of 
fabricated trench-anchored channel. Parylene devices 
without anchoring normally have a pressure limit of about 
3Opsi, above which the top parylene layer of the channel 
tends to delaminate from the bottom surface. The testing 
results show the anchored channels do perform much better 
in terms of pressure compatibility. 
Figure 4. Trench-anchored channel cross-sectional pictures. 
3. RESULTS AND DISCUSSION 
A packaging jig (Fig. 5 )  is developed for convenient testing 
of the chips. A fabricated chip is clamped between a PCB 
and the PEEK jig. Squeezed O-rings at chip backside provide 
sealing. Standard fitting receiving ports are made in the jig, 
so fluid connections are easily obtained. Electrical contacts 
are made by wire bonding from the on-chip Au pads to the 
PCB. The jig can access 8 out of the 16 holes (4x4) 
simultaneously on the chip. 
The pressure limits of trench-anchored and roughening- 
anchoring channels with an optional epoxy overcoat (Fig. 6) 
are obtained. Pressurized air is applied to special dead-ended 
testing channels. Bubble appearance in the water drop 
covering the channel indicates leakage. The pressure limit 
results are summarized in Table 1. It should be noted that the 
pressure limits also depend on the moat width, trench 
depthkhape, and parylene thickness. It is clear that trench- 
anchored channels perform much better than roughening- 
anchored ones, and epoxy overcoat can further improve the 
performance. Maximum pressure limit is achieved with 
trench-anchored, epoxy-coated channels. At the maximum 
testing pressure of SOOpsi, no leakage is found after an hour. 
The failure modes for the two anchoring technique are 
different. Roughening-anchored channels leak out working 
fluid through tiny pathways in the roughened moat area at 
213 
high pressure, while the trench-anchored channels don’t leak 
at all until the parylene breaks. 
I R 
PYlD 
T I R + E  I T + E  
-. ... 
with Backside Holes 
PEEK JIG 
I I 
Figure 5. Parts of the testing jig and a picture of the 
assembledpackage. 
Channel Chanoel 
Silicon Silicon + f 
Roughened Moat 
1 (b) 
Figure. 6 Cross section of trench and roughening anchored 
channels with optional epoxy overcoat. 
Table 1. Summary of thepressure testing result. 800psi is the 
marimum pressure of the testing setup. The safe pressures 
are for at least one hour. R: Roughening Anchoring; l? 
Trench Anchoring; E: Epoxy Overcoat. 
Pressure 
Breaking 
Drn......m 
Packing of beads into the on-chip column is achieved 
with standard slurry technique (Fig. 7). In order to get good 
and repeatable separation, packing quality is very important. 
It is found that in order to get uniform and void-free column, 
high pressurelflowrate during packing is necessary. Beads 
used to produce ion-exchange columns for the following 
experiments are 7pm-diameter anion exchange porous PS- 
DVB (Polystyrene-Divinylbenzene) resin with Trimethyl- 
Ammonium groups from Hamilton. The resins are the same 
used in Hamilton’s anion exchange column PRP-XI IO.  
(a) Parylene Beads Sensing 
Inlet Column Filter Outlet 
\ 
Channel 
Moat 
Sensing 
Electrodes 
PtlTi 
(b) Strengthening 
Beads 
Beads Packed 
in Column 
Figure 7. (a) Cross-sectional illustration of the device after 
packing; (b) Picture of column, packed beads, filter, detector, 
and channel-strengthening moat; (c) Fluorescentpicture of a 
densely packed column. 
Packed-column pressure flow-rate curve is measured 
for DI water (Fig. 8). The column is packed at 20Opsi before 
measurement. According to the theory [I] ,  
where 
F:  volumetric flow rate; At’: column presure drop; 
E: porosity of packed column; 7: viscosity of the fluid; 
d,: beads diameter; @; dimensionless flow resistance; 
A,: column cross-sectional area; L,: column length. 
From the fitted linear curve, it is found that @le = 444. 
Assuming the porosity of the packed column is 0.8, which is 
normal for densely-packed porous-spherical-beads column, 
then @ is 355, which is close to but smaller than its empirical 
value of 500 for slurry packed spherical porous-beads in 
conventional LC columns. It is believed that this is partly 
due to the small on-chip column to head size ratio. 
214 
Pressure V.S. Flow Rate for OnChip Packed Column 
Column Packed at 2OOpsi 
DI Water as Working Fluid 
Column L 8mmlW 100pmIH -25pm 
c 
: 
- Conditions: 
3.0- 7pm PS-DVB Beads 
2,5- 
- .  ._ E 2.0- 
3 
$ 1.5- 
a 
Measured Data 
0.5 Linear Fit 
0.0 Id
0 20 40 60 80 100120140160180200220 
Pressure (psi) 
Figure 8. Pressure flow-rate curve for an on-chip packed 
column. Beads type, packing condition, column dimension, 
and workingfluid are noted in thefigure. 
The detection cell impedance frequency response for 
ion-exchange chromatography mobile phase solution is 
measured (Fig. 9), which indicates that above ahout IOlCHz, 
the impedance is almost independent of frequency. It is 
adequate for measuring frequency to be in this resistance 
dominant region to minimize electrochemical effects [IO]. 
Detection Cell Impedance Frequency Response 
Solution: 1.7mM NaHC03 
and 1.8mM Na2C03 ”ool io00 1 
g 800- - 
$ 600- 
400- 
200- 
I 
- 
0 20 40 60 80 100 
Frequency (kHz) 
Figure 9. Zmpedancej?equency response of the detector 
Finally, using 1.7mM NaHC09 and 1.8mM Na2C03 
solution as mobile phase pumped at 0.2pl/min, seven-anion 
mixture in water (-25ppm) have been successfully separated 
using ion-exchange chromatography, and the chromatogram 
is obtained with on-chip conductivity sensor (Fig. IO). 
4. CONCLUSIONS 
An IC-processed Ion Liquid Chromatography chip, 
integrated with on-chip column, frits, injector and detector, is 
demonstrated. Channel-anchoring techniques have been 
developed to increase the pressure compatibility of the 
Parylene microfluidic system to at least 8OOpsi. When the 
column is packed with ion-exchange chromatography beads, 
multi-anion mixture (-25ppm) has been successfully 
separated and detected with on-chip conductivity sensor. 
Column: OnChip Packed Column. L 8 m W  I O O p d H  -2%. 
Stationary Phase: Hamilton PRP-Xi10 h i o n  Exchange 7~ Resin 
Mection: Conductivity, Detestion Cell Volume 0.4 nl 
3. Nitrite: 25ppm 
4. Bromide: 25ppm 
0 20 40 60 80 100 120 
Time (secJ 3; 
Mobile Phase: 1.7mM NaHC03, i.8mM a CO Isocratic; 0.ylmin. 
Figure IO .  Ion chromatography separation of seven-anion 
mixture. Separation conditions are noted in thefigure. 
ACKNOWLEDGEMENT 
This work is supported by the NSF CENS Center (CCR- 
0121778), NSF ERC Center at Caltech (EEC-9402726) and 
NIH (ROI RR06217). 
REFERENCES 
[I] V. R. Meyer, “Practical High-Performance Liquid 
Chromatography”, John Wiley & Sons, 1999, pp. 3 10-3 11 
[2] A. Manz, Y. Miyahara, J. Miura, et al., “Design of an 
open-tubular column liquid chromatography using silicon 
chip technology”, Sensors and Actuators BI,  (1990) 249-255. 
[3] B. He, N. Tait, F. Regnier, et al., “Fabrication of 
nanocolumns for liquid chromatography”, Anal. Chem., 1998, 
70,3790-3797. 
[4] J.P. Murrihy, M.C. Breadmore, A. Tan, et al., “Ion 
chromatography on-chip”, J of Chromatography A, 924 
(2001), 233-238. 
[5] A.K. Singh, D.J. Throchorton, J.S. Brennan, et al., 
“Gradient-elution reversed-phase electrochromatography in 
microchips”, Proc. pTAS 2003, pp. 1163.1 166. 
[6] G. Ocvirk, E. Verpoorte, A. Manr, et al., “High 
performance liquid chromatography partially integrated onto 
a Silicon chip”, Analytical Methods and Instrumentation, 
Vol. 2 No. 2,74-82 (1995). 
[7] L. Ceriotti, N.F. Rooij, and E. Verpoorte, “An integrated 
fritless column for on-chip capillaly electrochromatography 
with conventional stationaly phases”, Ana/. Chem., 2002, 74, 
639-647. 
[8] A.P. O’Neill, P. O’Brien, J. Alderman, et al., “On-chip 
definition of picolitre sample injection plugs for miniaturised 
liquid chromatography”, J. of Chromatography A, 924 
(2001), 259-263. 
[9] M. Liger, D.C. Rodger, and Y.-C. Tai, “Robust Parylene- 
to-Silicon mechanical anchoring”, MEMS’03, pp. 602-605. 
[IO] S. Bohm, B. Timmer, W. Olthuis, et al., “A closed-loop 
controlled electrochemically actuated micro-dosing system”, 
J Micromech. Microeng., I O  (2000), 498-504. 
215 


Ion liquid chromatography on-a-chip with beads-packed parylene column

Qing He

Changlin Pang

Yu-Chong Tai

Terry Lee

eScholarship - University of California

UCLAPapersTitleIon Liquid Chromatography System On-a-Chip with Beads-Packed Parylene ColumnPermalinkhttps://escholarship.org/uc/item/0fc6s47mAuthorsQing HeChanglin PangYu-Chong Taiet al.Publication Date2004 Peer reviewedeScholarship.org Powered by the California Digital LibraryUniversity of CaliforniaION LIQUID CHROMATOGRAPHY ON-A-CHIP WITH  BEADS-PACKED PARYLENE COLUMN  1Qing He, 1Changlin Pang, 1Yu-Chong Tai, and 2Terry D. Lee  1Caltech Micromachining Lab, California Institute of Technology, CA, USA 2Beckman Research Institute, City-of-Hope, CA, USA  ABSTRACT  A parylene-MEMS ion-exchange Liquid Chromatography (LC) chip is presented here. The chip is integrated with microfluidic I/O ports, a separation column, frits/filters, and a conductivity detector. The column is packed with conventional LC stationary phase support materials, i.e. micro-beads with surface functional groups. To withstand high pressure normally encountered in High Performance Liquid Chromatography (HPLC), a self-aligned, channel-anchoring technique is developed to increase the pressure rating of the Parylene microfluidic devices from 30 to at least 800psi. On-chip injection, separation and detection of anions in water, with ~25ppm concentration, have been successfully demonstrated. To our knowledge, this is the first demonstration of microbeads-packed column ion liquid chromatography (LC) on a chip.   1. INTRODUCTION  High Performance Liquid Chromatography (HPLC) is one of the most popular, powerful, and versatile separation techniques for chemical analysis. Although LC column is normally made of capillary tubes due to fluidics limitations, the miniaturization of the column can actually improve separation performance. As shown in Fig. 1 [1] where same separation chemistry applies, separated peak width is independent of column ID, while peak heights are larger for smaller columns and/or smaller beads. In short, further down-sizing could be favorable in liquid chromatography.     Figure 1. Separated peak shapes obtained with different column and bead size.           Compared to electrophoresis, however, little has been published about the integration of HPLC system onto a single chip. The two main reasons are the lacks of: (1) a process to integrate various components for an LC system; (2) high-pressure compatibility of the microfluidic devices needed for running HPLC. This work deals with both problems.          In terms of introduction and retaining LC stationary phase materials into the column, several approaches have been proposed, including open-tubular [2], coating micromachined posts arrays [3], coating microchannel with nanoparticles [4], monolith [5], packing [6], and packing without frit [7]. In this work, packing with conventional beads is chosen because of the following advantages. First, without introducing new surface chemistry, the extensively established separation knowledge can be utilized. Secondly, extreme flexibility is there to perform different types of liquid chromatography and/or to optimize particular separations, by choosing bead type, size, pore size, porosity, and functional group. Thirdly, packed column can achieve reproducible column performance, which is usually a problem in other methods.   2. DESIGN AND FABRICATION  The LC-on-a-chip device is made using parylene MEMS technology. The chip has four access ports, as shown in Fig. 2. LC mobile phase flows from port #1 to #2. Nanolitre volume sample injection is achieved with a cross channel injection method [8] from #3 to #4. Beads are packed externally into the on-chip column from port #1. Filters (channel with a height smaller than the bead size) trap beads in the main channel and prevent beads from entering the side injection channels. The bead-packed column inlet is near the sample output port #4, which is the injected sample plug front. The column outlet is at the filter/sensor edge (Fig. 2(b)). This integrated design minimizes dead volume, thus minimizing extra-column peak broadening. Off-chip pumps and valves are used in testing.         A conductivity sensor is used as the detector for ion sensing. Interdigital electrodes are patterned in the detector cell to monitor liquid conductivity. The conductivity of the mobile phase solution provides a baseline signal. When separated ion plugs pass by the detector, changes of solution conductivity are detected. Chromatogram is then obtained by recording conductivity of the solution flowing in the detection cell over time. To increase sensitivity, electrode width and spacing should be small, and the total electrode area should be maximized. However, the electrode design should be compromised in considerations of minimizing peak broadening and multiple-peak detection.  1 3 4 2 Injection Column Peak in Eluate               Figure 2. (a) Fluorescent overview picture of the integrated separation system. The channels are filled with Fluorescein; (b) Optical picture after packing beads.  Etch Backside H oles w ith DRIEPattern Frontside O xide Evaporate and Pattern Ti/Pt/AuPattern Two-Level Photoresist StructureEtch O xide using Patterned Photoresist as M askDRIE O pen Backside H olesA cetone Dissolves PhotoresistEtch Special Trench w ith DRIED eposit and Pattern Parylene Si O xide Pt/Ti Photoresist Parylene  Figure 3. Simplified fabrication flow.          The fabrication process (Fig. 3) starts with growing thermal oxide on Silicon wafer. Then a two-step DRIE is performed on wafer backside to create the backside holes, leaving only a 50µm-thick diaphragm. Following frontside oxide patterning, a Ti/Pt/Au (300Å/2000Å/1000Å) layer is evaporated and patterned. Au is left only at bonding pads. Pt is used as the sensing electrode and is patterned to be interdigital 5µm-wide electrodes at 5µm intervals. 25µm-thick photoresist is spun and patterned with two masks to form two-level sacrificial photoresist structure. The 25µm-high channel part uses unexposed photoresist, while the 4µm-high filter part is partially exposed. A 15µm-wide “moat”, self-aligned-to channels and filters, is fully exposed to the bottom oxide, which is subsequently etched away. Modified DRIE process [9] is then done to make 40µm-deep trench with mushroom bottom profile in the moat, or XeF2 is used to roughen the moat area.  About 10µm-thick parylene is conformally deposited filling up the trench or the roughened moat and is patterned. Backside holes are open with DRIE. Acetone dissolves photoresist and releases the device.         In the above process, a self-aligned, channel-anchoring technique is used to increase the pressure rating of the parylene chip. Figure 4 shows cross-sectional pictures of fabricated trench-anchored channel. Parylene devices without anchoring normally have a pressure limit of about 30psi, above which the top parylene layer of the channel tends to delaminate from the bottom surface.  The testing results show the anchored channels do perform much better in terms of pressure compatibility.    Figure 4. Trench-anchored channel cross-sectional pictures.  3. RESULTS AND DISCUSSION  A packaging jig (Fig. 5) is developed for convenient testing of the chips. A fabricated chip is clamped between a PCB and the PEEK jig. Squeezed o-rings at chip backside provide sealing. Standard fitting receiving ports are made in the jig, so fluid connections are easily obtained. Electrical contacts are made by wire bonding from the on-chip Au pads to the PCB. The jig can access 8 out of the 16 holes (4x4) simultaneously on the chip.         The pressure limits of trench-anchored and roughening-anchoring channels with an optional epoxy overcoat (Fig. 6) are obtained. Pressurized air is applied to special dead-ended testing channels. Bubble appearance in the water drop covering the channel indicates leakage. The pressure limit results are summarized in Table 1. It should be noted that the pressure limits also depend on the moat width, trench depth/shape, and parylene thickness. It is clear that trench-anchored channels perform much better than roughening-anchored ones, and epoxy overcoat can further improve the performance. Maximum pressure limit is achieved with trench-anchored, epoxy-coated channels. At the maximum testing pressure of 800psi, no leakage is found after an hour.          The failure modes for the two anchoring technique are different. Roughening-anchored channels leak out working fluid through tiny pathways in the roughened moat area at #1: Eluent In#2: Eluent Out#3: Sample In#4: Sample OutFilter & SensorFilterFilter500μm(b) Packed Column Filter & Sensor Filter (a)  Empty Column ~ 40µmSi ChannelAir Air Si Parylene Channel Parylene high pressure, while the trench-anchored channels don’t leak at all until the parylene breaks.     Figure 5. Parts of the testing jig and a picture of the assembled package.             Figure. 6 Cross section of trench and roughening anchored channels with optional epoxy overcoat.  Table 1. Summary of the pressure testing result. 800psi is the maximum pressure of the testing setup. The safe pressures are for at least one hour. R: Roughening Anchoring; T: Trench Anchoring; E: Epoxy Overcoat.   R T R + E T + E Safe Pressure 250 psi 600 psi  700 psi  800 psi  > Safe Pressure Bubbles Appear No Bubble until Break Bubbles Appear NA Breaking Pressure ~ 350 psi ~ 700 psi ~ 800psi > 800 psi                  Packing of beads into the on-chip column is achieved with standard slurry technique (Fig. 7). In order to get good and repeatable separation, packing quality is very important.   It is found that in order to get uniform and void-free column, high pressure/flowrate during packing is necessary. Beads used to produce ion-exchange columns for the following experiments are 7µm-diameter anion exchange porous PS-DVB (Polystyrene-Divinylbenzene) resin with Trimethyl-Ammonium groups from Hamilton. The resins are the same used in Hamilton’s anion exchange column PRP-X110.         Figure 7. (a) Cross-sectional illustration of the device after packing; (b) Picture of column, packed beads, filter, detector, and channel-strengthening moat; (c) Fluorescent picture of a densely packed column.          Packed-column pressure flow-rate curve is measured for DI water (Fig. 8). The column is packed at 200psi before measurement. According to the theory [1],  2where: volumetric flow rate; : column presure drop;: porosity of packed column; : viscosity of the fluid;: beads diameter; : dimensionless flow resistance;                       p ccpF Pdd AF PLε ηεη∆Φ= ∆Φ          : column cross-sectional area;  : column length.c cA L        From the fitted linear curve, it is found that Φ/e = 444. Assuming the porosity of the packed column is 0.8, which is normal for densely-packed porous-spherical-beads column, then Φ is 355, which is close to but smaller than its empirical value of 500 for slurry packed spherical porous-beads in conventional LC columns. It is believed that this is partly due to the small on-chip column to bead size ratio.   (a)(b)(c)Beads Packed in ColumnSensing Electrodes Pt/TiBeadsChannel Strengthening MoatColumn Filter OutletInletParylene Sensing Electrodes Pt/TiBeads100μm100μmSilicon PR Parylene Oxide Silicon PR Parylene Oxide Epoxy Epoxy (a) (b) Roughened Moat Trench Moat Channel Channel 0 20 40 60 80 100 120 140 160 180 200 2200.00.51.01.52.02.53.0 Measured Data Linear FitFlow Rate (µl/min)Pressure (psi)Pressure v.s. Flow Rate for On-Chip Packed ColumnConditions:7µm PS-DVB BeadsColumn Packed at 200psiColumn L 8mm/W 100µm/H ~25µmDI Water as Working Fluid Figure 8. Pressure flow-rate curve for an on-chip packed column. Beads type, packing condition, column dimension, and working fluid are noted in the figure.          The detection cell impedance frequency response for ion-exchange chromatography mobile phase solution is measured (Fig. 9), which indicates that above about 10kHz, the impedance is almost independent of frequency. It is adequate for measuring frequency to be in this resistance dominant region to minimize electrochemical effects [10].  0 20 40 60 80 100020040060080010001200Impedance (kΩ)Frequency (kHz)Detection Cell Impedance Frequency ResponseSolution: 1.7mM NaHCO3        and 1.8mM Na2CO3 Figure 9. Impedance frequency response of the detector.          Finally, using 1.7mM NaHCO3 and 1.8mM Na2CO3 solution as mobile phase pumped at 0.2µl/min, seven-anion mixture in water (~25ppm) have been successfully separated using ion-exchange chromatography, and the chromatogram is obtained with on-chip conductivity sensor (Fig. 10).   4. CONCLUSIONS  An IC-processed Ion Liquid Chromatography chip, integrated with on-chip column, frits, injector and detector, is demonstrated. Channel-anchoring techniques have been developed to increase the pressure compatibility of the Parylene microfluidic system to at least 800psi. When the column is packed with ion-exchange chromatography beads, multi-anion mixture (~25ppm) has been successfully separated and detected with on-chip conductivity sensor. 0 20 40 60 80 100 120234.4234.6234.8235.0235.2235.4235.6235.8236.0236.2SO4-2PO4-3NO3-NO2-Br-Cl-F-Mobile Phase: 1.7mM NaHCO3, 1.8mM Na2CO3; Isocratic; 0.2µl/min. 1. Fluoride: 12.5ppm2. Chloride: 25ppm3. Nitrite: 25ppm4. Bromide: 25ppm5. Nitrate: 25ppm6. Phosphate: 25ppm7. Sulfate: 25ppmColumn: On-Chip Packed Column. L 8mm/W 100µm/H ~25µm.Stationary Phase: Hamilton PRP-X110 Anion Exchange 7µm ResinDetection: Conductivity, Detection Cell Volume 0.4 nlInjection: 4 nl; Retention: ~3 min after injection before Time 0Signal Voltage (mV)Time (sec)Figure 10. Ion chromatography separation of seven-anion mixture. Separation conditions are noted in the figure.   ACKNOWLEDGEMENT  This work is supported by the NSF CENS Center (CCR-0121778), NSF ERC Center at Caltech (EEC-9402726) and NIH (R01 RR06217).   REFERENCES  [1] V. R. Meyer, “Practical High-Performance Liquid Chromatography”, John Wiley & Sons, 1999, pp. 310-311 [2] A. Manz, Y. Miyahara, J. Miura, et al., “Design of an open-tubular column liquid chromatography using silicon chip technology”, Sensors and Actuators B1, (1990) 249-255.  [3] B. He, N. Tait, F. Regnier, et al., “Fabrication of nanocolumns for liquid chromatography”, Anal. Chem., 1998, 70, 3790-3797. [4] J.P. Murrihy, M.C. Breadmore, A. Tan,  et al., “Ion chromatography on-chip”, J. of Chromatography A, 924 (2001),  233-238. [5] A.K. Singh, D.J. Throckmorton, J.S. Brennan, et al., “Gradient-elution reversed-phase electrochromatography in microchips”, Proc. µTAS 2003, pp. 1163-1166. [6] G. Ocvirk, E. Verpoorte, A. Manz, et al., “High performance liquid chromatography partially integrated onto a Silicon chip”, Analytical Methods and Instrumentation, Vol. 2 No. 2, 74-82 (1995).  [7] L. Ceriotti, N.F. Rooij, and E. Verpoorte, “An integrated fritless column for on-chip capillary electrochromatography with conventional stationary phases”, Anal. Chem., 2002, 74, 639-647.  [8] A.P. O’Neill, P. O’Brien, J. Alderman, et al., “On-chip definition of picolitre sample injection plugs for miniaturised liquid chromatography”, J. of Chromatography A, 924 (2001), 259-263. [9] M. Liger, D.C. Rodger, and Y.-C. Tai, “Robust Parylene-to-Silicon mechanical anchoring”, MEMS’03, pp. 602-605. [10] S. Böhm, B. Timmer, W. Olthuis, et al., “A closed-loop controlled electrochemically actuated micro-dosing system”,  J. Micromech. Microeng., 10 (2000), 498-504. 

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Ion Liquid Chromatography System On-a-Chip with Beads-Packed Parylene Column

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Ion liquid chromatography on-a-chip with beads-packed parylene column

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