Method for membrane assisted liquid chromatography

Apparatus and method for extracting a component from a sample across a membrane into an extractant and then injecting the extracted sample component into a chromatographic eluent and onto a chromatographic column to chromatographically analyze the extracted sample component. In essence, the advance provided by this invention is that the extractant and the eluent are the same and only one pump is used for pumping the eluent and extractant. In prior apparatus and methods in this field, the extractant and the eluent were separate and each had its own pump.

FIELD OF THE INVENTION 
The invention is in the field of liquid chromatography and more 
specifically in the field of using membranes to pretreat samples to be 
analyzed by liquid chromatography. 
BACKGROUND OF THE INVENTION 
The chemical analysis technique comprising partitioning a sample containing 
a sample component of interest from a liquid extractant with a membrane is 
known. The sample component of interest permeates through the membrane 
into the extractant which can then be analyzed to determine the component 
of interest. The specific membrane and extractant used are selected to 
enhance the extraction of the sample component of interest and to minimize 
or eliminate the extraction of other components of the sample that may not 
be of interest or that may interfere with the determination of the 
component of interest. One means used to determine a sample component of 
interest in the extractant is liquid chromatography and the overall system 
is then termed "membrane assisted liquid chromatography". In membrane 
assisted liquid chromatography a preselected volume of extractant 
containing the extracted sample component of interest is injected into a 
liquid chromatographic system and the sample component of interest is 
thereby determined. 
One example of membrane assisted liquid chromatography is described in U.S. 
Pat. No. 4,529,521 to Hernan J. Cortes and James C. Davis. A sample of 
synthetic latex solution is partitioned from a water extractant by a 
bundle of dialysis type hollow fiber membranes. The water extractant is 
positioned in the bores of the hollow fibers and relatively low molecular 
weight components in the latex solution permeate through the membrane into 
the extractant. A syringe filled with water is used to pump the extractant 
from the bores of the hollow fibers into the injection loop of a liquid 
chromatography injection valve in the load position. A liquid 
chromatography pump is used to pump dilute sulfuric acid eluent through 
the injection valve, through a liquid chromatography column and then to a 
liquid chromatography photometric detector. When the injection valve is 
placed in the inject position, the eluent pumps the extractant in the 
injection loop onto the column and the extracted relatively low molecular 
weight components of the latex solution are chromatographed and eventually 
emerge from the column to be detected by the detector. If the latex sample 
is injected directly onto the column, the latex particles will soon plug 
the column. 
The use of a membrane to pretreat a sample that can not be directly 
injected is a significant improvement in the art of liquid chromatography. 
However, at least one problem remains with this approach at its present 
state of development. This problem is the complexity of known membrane 
assisted liquid chromatography systems in that two solutions are used 
(extractant and eluent) and two pumping means are needed, one for the 
extractant and one for the eluent. The present invention is a solution to 
this problem. 
SUMMARY OF THE INVENTION 
In the method of the present invention, the same solution is used as both 
the extractant and the eluent. In the apparatus of the present invention, 
a single pumping means is used for pumping the eluent and extractant. 
The apparatus of the present invention comprises six elements. The first 
element is a means for pumping a liquid such as a liquid chromatography 
eluent pump. The second element is a membrane having a first side and a 
second side such as a tubular shaped membrane. The third element is a 
channel having a first end and a second end, at least a portion of the 
channel being formed by the first side of the membrane so that the second 
side of the membrane can be exposed to a sample containing a sample 
component. The fourth element is an injection conduit having a first end 
and a second end for containing a preselected volume of the the liquid 
such as an injection loop, e.g., a preselected length of tubing. The fifth 
element is a liquid chromatography column for chromatographing the sample 
component, the liquid chromatography column having an inlet port. The 
sixth element is a means for switching liquid flow between a first flow 
pattern and a second flow pattern, being in liquid communication with: the 
means for pumping the liquid; the first end of the channel; the second end 
of the channel; the first end of the injection conduit; the second end of 
the injection conduit; and the inlet port of the chromatography column. 
The first flow pattern being from the pumping means through the channel 
and through the injection conduit. The second flow pattern being from the 
pumping means, through the injection conduit, to the inlet port of the 
liquid chromatography column. The means for switching liquid flow between 
a first flow pattern and a second flow pattern can be one or more 
multi-port valves such as a 10-port valve, an 8-port valve or a pair of 
6-port valves. 
The method of the present invention comprises five steps. The first step is 
to flow a liquid extractant/eluent into contact with one side of a two 
sided membrane, e.g., flowing the extractant/eluent into the bore of a 
tubular membrane. The second step is to contact the other side of the 
membrane with a sample so that a component of the sample permeates through 
the membrane into the extractant/eluent to form a dispersion of the sample 
component in the extractant/eluent. The third step is to flow the 
dispersion of the sample component in the extractant/eluent into an 
injection conduit, e.g., flowing the dispersion of the sample component in 
the extractant/eluent into an injection loop. The fourth step is to flow 
the extractant/eluent into the injection conduit so that the dispersion of 
the sample component in the extractant/eluent in the injection conduit is 
flowed into a chromatographic column which chromatographs the sample 
component so that at least a portion of the sample component eventually 
emerges from the chromatographic column dispersed in the extractant/eluent 
emerging from the chromatographic column. The fifth step is to detect the 
sample component dispersed in the extractant/eluent emerging from the 
chromatographic column, e.g., to detect the sample component dispersed in 
the extractant/eluent emerging from the chromatographic column using a 
photometric liquid chromatography detector or an electrochemical liquid 
chromatography detector.

DETAILED DESCRIPTION OF THE INVENTION 
Referring to FIGS. 1 and 2, therein is shown a schematic drawing of an 
apparatus embodiment of the invention including a two position 10-port 
valve 20 (such a valve is available from the Anspec Co., Ann Arbor, Mich., 
as the Valco 10-port multi-function sampling valve, Catalog no. H1765) 
shown in one position in FIG. 1 and in the other position in FIG. 2. The 
valve 20 is a preferred means for switching liquid flow between a first 
flow pattern and a second flow pattern. The specific means used for this 
switching is not critical to the invention and many different multi-port 
valve constructions can be used as will be discussed in further detail 
below in reference to Figs. 3 and 4. A reservoir 21 is provided for 
containing a liquid extractant/eluent 22. A length of tubing 23 connects 
the reservoir 21 with a liquid chromatography pump 24 (available from 
Anspec, supra, as the ConstaMetric IIIG pump, as Catalog No. F1025). The 
specific pumping means used is not critical to the invention as long as it 
is suitable for the liquid chromatography of the invention. The pump 24 is 
connected (1/16 stainless steel tubing and fittings used generally 
throughout herein for such connections and is available from Anspec, 
supra) to port 1 of the valve 20 by a length of tubing 25. The valve 20 
internally connects the valve ports as shown, i.e., ports 1-2, 3-4, 5-6, 
7-8, 9-10 in FIG. 1, and ports 2-3, 4-5, 6-7, 8-9, 10-1 in FIG. 2. A 
length of tubing 26 is used to connect port 10 to port 5. A sample loop 27 
is connected to port 4 and port 6 of the valve 20. The sample loop 27 is a 
preferred example of a conduit having a preselected volume for injecting a 
liquid contained in the preselected volume into a flowing stream of liquid 
and this conduit can take other forms such as a conduit that is internally 
machined into a valve. A length of tubing 28 is used to connect port 7 of 
the valve 20 and an inlet port 29 of a liquid chromatography column 30. 
Another length of tubing 31 is used to connect port 8 of the valve 20 and 
an outlet port 32 of the column 30. The specific column 30 used is not 
critical to the invention and can include capillary columns, packed 
capillary oolumns, packed microbore columns, packed columns, semi-prep 
columns and prep columns. A 4-port valve, not shown, (for example, Anspec 
Catalog No. A7752) can be installed between the column 30 and the 10-port 
valve 20 to reverse flow through the column 30, if desired. A length of 
tubing 33 is used to connect port 9 of the valve 20 to a liquid 
chromatography detector 34. The specific detector 34 used is not critical 
to the invention and can include photometric detectors and electrochemical 
detectors. 
FIGS. 1 and 2 also schematically show a tubular membrane 35 immersed in a 
sample 36 which is contained in a cup 19. The bore of the tubular membrane 
35 defines a channel having a first end 37 and a second end 38. The 
outside of the membrane 35 is exposed to the sample 36. A length of tubing 
39 connects port 2 of the valve 20 to the first end 37 of the channel and 
another length of tubing 40 connects port 3 of the valve 20 to the second 
end 38 of the channel. 
Referring to FIG. 1, a specific flow pattern is shown and critically 
includes a flow pattern of from the pump 24, through the valve 20, through 
the bore of the membrane 35, and then into the sample loop 27. This flow 
pattern allows the extractant/eluent 22 to be flowed into contact with the 
inside of the membrane 38. The outside of the membrane 38 is contacted 
with the sample 36 so that generally more than one component of the sample 
(and critically at least one component of the sample) can permeate through 
the membrane into the extractant/eluent in the bore of the membrane 38 to 
form a dispersion of the sample components in the extractant/eluent in the 
bore of the membrane 38. Continuing flow of the extractant/eluent 22 in 
the tubing 39 flows the dispersion of the sample components in the 
extractant/eluent in the bore of the membrane 35 into the tubing 40 and 
then into the sample loop 27. Since the components of the sample generally 
continuously permeate through the membrane 38, it is often preferable to 
continue pumping the extractant/eluent 22 into the bore of the membrane 38 
so that the dispersion of the sample components in the extractant/eluent 
continues to flow into and then through the sample loop 27, through the 
tubing 26 and 33 to the detector 34. In this event, it is often possible 
to monitor the sample components in the extractant/eluent with the 
detector 34 to insure that a generally preferable steady state of 
permeation of the sample components has been achieved before switching 
valve 20 to the position shown in FIG. 2. 
Referring to FIG. 2, a specific flow pattern is shown and critically 
includes a flow pattern of from the pump 24, through the valve 20, through 
the sample loop 27, through the column 30, through the valve 20 and then 
to the detector 34. This flow pattern allows the extractant/eluent to flow 
into the sample loop 27 so that the dispersion of the sample components in 
the extractant/eluent in the sample loop 27 is flowed into the column 30 
for chromatographic retention of at least one sample component and 
preferably a chromatographic separation of all of the sample components so 
that at least a portion of one of the sample components eventually emerges 
from the column 30 in the extractant/eluent emerging from the 
chromatographic column 30 (from the port 32) and then flows through the 
valve 20 to the detector 34. The detector 34 detects at least one sample 
component and generally outputs to an integrator recorder 41 as is well 
known in the art. 
When the valve 20 is in the position shown in FIG. 2, there is essentially 
no flow of extractant/eluent in the bore of the membrane 35 and a 
relatively high concentration of the permeated sample components can build 
up in the extractant/eluent in the bore of the membrane 35 over an 
extended period of time, e.g. 10 minutes. If desired, this characteristic 
can be beneficially used to increase detection sensitivity of the 
permeated sample components by switching the valve 20 to the position 
shown in FIG. 1 only long enough to move the relatively high concentration 
of the permeated sample components into the sample loop 27 and then switch 
the valve 20 back to the position shown in FIG. 2 for a chromatographic 
separation of the components. 
Referring to FIG. 1, the column 30 is isolated from the flow pattern so 
that the membrane 38 will not be subjected to the relatively high 
pressures often generated when pumping a liquid eluent through a liquid 
chromatography column. Referring to FIG. 2, the membrane 38 is isolated 
from the flow pattern so that, as in above, the membrane 38 will not be 
subjected to the relatively high pressures often generated when pumping a 
liquid eluent through a liquid chromatography column. The embodiment of 
the invention shown in Figs. 1 and 2 demonstrates one of the principal 
advantages of the present invention in that the extractant is the same as 
the eluent and only one pump is used. The invention requires that the 
extractant perform satisfactorily as an eluent and vice versa but the 
extensive knowledge available to the liquid chromatographer about the many 
different eluent compositions used in liquid chromatography (including 
reverse phase, normal phase, ion exchange, size exclusion, and 
hydrodynamic chromatography) should be an advantage when one skilled in 
the art of liquid chromatography uses the present invention. 
Referring to FIGS. 3 and 4, therein is shown a schematic drawing of another 
apparatus embodiment of the invention including a pair (A and B) of two 
position 6-port valves 42 and 43 (such as valves available from Anspec, 
supra, as a pair of Rheodyne Type 70 switching valves, Catalog No. F1131, 
preferably mounted on a Catalog No. H1687 tandem solenoid actuator so that 
each valve can be switched at essentially the same time). An understanding 
of this embodiment is readily apparent from the above discussion of FIGS. 
1 and 2 except that the valve port numbers may be different. It should be 
understood that many specific valve systems incorporating one or more 
valves could be used in the invention as the means for switching between 
two flow patterns. For example, the valve 42 in FIGS. 3 and 4 could be a 
4-port valve and the valve 20 in FIGS. 1 and 2 could be an 8-port valve. 
It is believed that the 10-port valve 20 is the best since it is 
commercially available and involves only one valve body. An air actuated 
8-port valve suitable for use in the invention is available from the Valco 
Co., Houston, Tex., as part number AC8W. In any event, a critical feature 
of the means for switching the extractant/eluent flow between a first flow 
pattern and a second flow pattern is that the means be in liquid 
communication, e.g., by 1/16 inch diameter stainless steel tubing, with 
the means for pumping the extractant/eluent, with each end of the membrane 
35, with each end of the sample loop 27 and with the inlet port 29 of the 
column 30. 
The specific type of membrane used is not critical to the invention. The 
membrane can be flat in shape and form a portion of a channel cut, for 
example, in a stainless steel or Teflon.RTM. block. The membrane can be 
tubular in shape and the tube can be relatively small in diameter, e.g., 
0.025 inches or smaller, or relatively large in diameter, e.g., 0.1 inches 
or larger. The membrane can be of the porous type or the non-porous type. 
The membrane can be hydrophilic or hydrophobic. Critically, the membrane 
should not rapidly deteriorate, e.g., dissolve, in the extractant/eluent 
or the sample. Critically, the membrane should permeate at least one 
component of the sample to the extractant/eluent when the membrane 
partitions the two. A preferred membrane is a tubular silicone rubber 
membrane. 
EXAMPLE 1 
The system shown in FIGS. 1 and 2 (except that a different membrane 
configuration is used as described below) is assembled and includes a 
Hewlett Packard Hypersil ODS 5 micron liquid chromatography column 30 (2.1 
mm.times.100 mm), a Kratos Spectroflow 773 variable wavelength liquid 
chromatography detector 34 (set at 254 nanometers) and a Spectraphysics 
4270 integrator-recorder 41. The eluent/extractant 22 is 50% acetonitrile, 
50% water, 0.02M in phosphoric acid pumped at a flow rate of 200 
microliters per minute. The sample loop 27 contains a fixed volume of 100 
microliters. 
A membrane cell 50, preferred for polar eluent/extractants, is assembled as 
shown in FIG. 5. A 150 mm long 2 mm internal diameter glass tube 51 is 
provided with a sample inlet neck 52 and a sample outlet neck 53. The 
membrane 54 used is Dow Corning Silastic.RTM. Medical Tubing (0.012 inches 
internal diameter, 0.025 inches external diameter, 100 mm long) and is 
spiral wound on a 60 mm long length of 1/32 inch outside diameter 
Teflon.RTM. tubing 55. One end of the tubing 55 is the eluent/extractant 
inlet 57. The other end of the tubing 55 is joined to one end of the 
membrane 54 by first swelling the end of the membrane 54 in xylene, 
inserting the end of the tubing 55 into the end of the swollen membrane 54 
and then allowing the xylene to evaporate to shrink the membrane 54 onto 
the tubing 55 so that a leak tight joint is formed. The other end of the 
membrane 54 is similarly joined to an eluent/extractant outlet 56 which is 
also made from 1/32 inch diameter Teflon.RTM. tubing. The ends of the 
tube 51 are each sealed with Dow Corning.RTM. RTV Silicone Rubber Sealant 
58. The membrane cell 50 is highly preferred for polar eluent/extractants, 
e.g. water based eluent/extractants, because it is easily made and 
provides excellent contact between the sample and the membrane 54 when the 
sample is flowed into the sample inlet neck 52. In non-polar 
eluent/extractants, e.g., toluene, the membrane 54 can swell excessively 
and cause blocked flows. In this event a cell is preferred wherein the 
silicone rubber membrane is assembled in a stretched condition so that 
when it swells the degree of stretch is substantially reduced or a cell 
where the length of membrane can be subsequently adjusted to accommodate 
the swollen membrane. Alternatively, a cell can be assembled with the 
membrane already in the swollen condition. 
A sample containing 1 ppm (parts per million) each of the sample components 
benzene, toluene, styrene and ethyl benzene in water is flowed into the 
sample neck 52 at a flow rate of about 1 ml per minute with the system 
flow pattern as shown in FIG. 1. The integrator/recorder 41 initially 
shows that the absorbance of the extractant/eluent flowing through the 
detector 34 is not increased. However, the absorbance of the 
extractant/eluent flowing through the detector 34 soon increases and then 
reaches a steady state. The valve 20 is then rotated to the position shown 
in FIG. 2 and the integrator/recorder 41 then traces a chromatogram over 
the next 10 minutes showing a separate peak for benzene, toluene, styrene 
and ethyl benzene each of a given peak area and peak height. 
This example teaches how to make a preferred membrane cell for use in the 
invention and how to determine sample components with the invention. 
EXAMPLE 2 
The system of Example 1 is used in this example and the experiment of 
Example 1 is continued. The valve 20 is left in the position shown in FIG. 
2 for 11 minutes (during which time the eluent/extractant in the bore of 
the membrane 54 is not flowing which generally increases the concentration 
of the permeated sample components therein) and then is switched to the 
position shown in FIG. 1 for a time needed to transport the permeated 
sample components from the bore of the membrane 54 into the sample loop 
27. Then the valve 20 is rotated back to the position shown in FIG. 2 and 
the integrator/recorder 41 then traces a chromatogram over the next 10 
minutes showing a separate peak for benzene, toluene, styrene and ethyl 
benzene each of a peak height and peak area greater than in Example 1. The 
peak height and area of the benzene peak is about 3.5 times greater. The 
peak height and area of the toluene peak is about 4.2 times greater. The 
peak height and area of the styrene peak is about 4.5 times greater. The 
peak height and area of the ethyl benzene peak is about 6.3 times greater. 
In this example the sample is fed into the sample inlet port 52 of the 
membrane cell 50 of FIG. 5. Alternatively, water could have been fed into 
the sample inlet port 52 of the membrane cell 50 of FIG. 5 and the sample 
could have been injected into the water and carried by it into contact 
with the membrane 54. In this event, the detector sees a "peak" resulting 
from injection of a sample if the valve 20 is not rotated when the 
permeated sample component(s) flow through the sample loop 27 and this 
mode of analysis can be used if a chromatographic separation of the 
permeated sample component(s) is not desired. 
This example teaches how to gain better sensitivity of analysis with the 
invention. 
EXAMPLE 3 
The system of Example 2 is changed so that the Column 30 is a Brownlee 10 
micron PRP-1 (4.6 mm.times. 30 mm), the eluent/extractant 22 is 0.01N NaOH 
containing 2.5% acetonitrile, and the detector 34 is changed to an 
LDC/Milton Roy e c Monitor electrochemical detector set at +0.55 volts, 
and the sample is changed to a sample containing 100 ppb (parts per 
billion) of the sample component phenol and 20 ppb of the sample component 
2-chlorophenol, in water. The valve 20 is placed in the position shown in 
FIG. 1 until steady state permeation is indicated and then the valve 20 is 
placed in the position shown in FIG. 2 for 15 minutes. Then the valve 20 
is switched to the position shown in FIG. 1 for a time needed to transport 
the permeated sample components from the bore of the membrane 54 into the 
sample loop 27. Then the valve 20 is rotated back to the position shown in 
FIG. 2 and the integrator/recorder 41 then traces a chromatogram over the 
next 10 minutes showing a separate peak for phenol and 2-chlorophenol. 
This example teaches the use of an electrochemical detector in the 
invention. This example also teaches the determination of relatively low 
concentrations of two phenolic compounds.