Note: Descriptions are shown in the official language in which they were submitted.
10602Z~
This application is a division of Canadian Application
231,834 filed July 18, 1975. ¦ -
This invention relates to liquid chromatographic
detectors and more particularly to improved fiow cell
constructions for liquid chromatography.
A chromatographic detector is a device which supplies ~-
an output signal related to the amount or rate of change of
the amount of a sample to be detected in the effluent of a
chromatographic column. It indicates elution of the
separated components of the input substance by the column
and provides a measure of the amount of each component. It
is usually the most sophisticated and one of the most
expensive components in a chromatograph. The majority of high
performance liquid chromatograph detectors in use today are
UV or visible light absorption and refractive index detectors.
Light rays are directed through the sample and the effect
of the sample on the rays, e.g., light absorption, is
detected by a photocell.
Good detectability, i.e., ability to detect a small
sample, is desired so that small samples and small column
capacity can be used, resulting in shorter analysis times.
Some new, highly efficient, column packing materials have
inherent low capacity and require detectors capable of
detecting small samples. Also, low sample solubility in the
mobile phase may limit the amount of sample available for
detection.
With small flow cell volumes, present flow cell geome-
tries offer low light transmission, poor flow geometry, and
become expensive to manufacture. Parallel light rays are used
with flat or cylindrical cell entry windows. Also, convergent
light rays have been used with flat windows, and typically
the convergent light rays are focused on the entry window for
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maximum transmission to the ceIl. The path lengths of
parallel light rays passing through a cylindrical cell differ
across the width of the cell. Also, the path lengths of
convergent light rays passing through a flow cell with flat
entrance and exit windows differ across the area of the
flow cell. Since light absorbance is a function of the path
length through the cell, the output signal from such cells
is nonlinear; that is, the detector response does not change
linearly with the amount of sample present. Linearity is
desirable because it facilitates quantitation of the results,
eliminating the need of functional callibration curves to
determine the quantity of the component present.
In particular, there is a need in the prior art for a
high light transmission cell with a small aperture to acco- -
mmodate a small sample for use in high performance liquid
chromatography and which affords equal path length for the
light rays directed through the cell.
According to the present invention there is provided
for use in liquid chromatography~ a detector comprising:
a flow cell, said flow cell comprising a union connectable
at one end to a column of a liquid chromatography apparatus,
said union having a flow passage from said one end thereof
to the other end thereof for effluent from said column,
entranee and exit windows disposed in said union on diametri-
cally opposite sides of said effluent flow passage, each of
said windows being in the form of a cylinder segment
providing a light acceptanee angle of at least 45; a light
source emitting a eollimated beam of light rays and having
a light emitting area larger than the cross-sectional area
of said passage; a lens for focusing all rays from said
light souree to said passage through said entranee window in
paths normal to the eylindrical surface of said entranee
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window; and means for detecting light rays that emerge from
said exit window.
A flow cell for liquid chromatography is described
that has a small cell volume and that utilizes convergent
light rays and a cylindrical window so that the path length
for all rays focused on the cell is the same. This arrange-
ment assures high light transmission even though only a
small aperture for the light is available, and assures good
linearity of the output. A large light acceptance angle is
afforded by the construction; and, when used with a broad
light source, the construction averages any local light
intensity fluctuations, thereby enhancing the accepta~ility
of a signal beam detector. The construction can also be
used in conjunction with beam condensing lenses in a spectro- -
photometer without loss of linearity in response.
The described flow cell of the present invention is
constructed for direct coupling to a chromatograph column,
is capable of ligh pressure and-high temperature operation,
is chemically inert, and is of versatile construction to
facilitate different apertures. Economical fabrication is
assured because this cell can bé fabricated from a standard
compression fitting union without intricate machining.
In one embodiment of the invention, dual beam flow
cells are provided with a common exit for both the detection
and the reference cells. This allows~a compact cell
construction and keeps the cells close together for thermal
equilibration.
I More specifically, the present flow cell constructionutilizes a quartz cylindrical window surrounding a small
capacity flow passage. Light stops are provided within the
window structure and passage to limit the light passing
through the cell to convergent rays, but at the same time
106~Z7
providing a large acceptance angle for the rays. With this
construction, rays transmitted through the flow cell are of
a high intensity notwithstanding the small aperture
necessitated by the size of the flow passage. The cell is
housed in a compression fitting union directly coupled
to the chromatographlc column. Where a reference cell is
- desired to factor out solvent effects and light fluctuations,
a feature of this invention includes the axial alignment of
two flow cell passages that are fed from opposite directions
and which discharge into a common zone centrally between
the two passages. The flow is then discharged through a
single port in a direction perpendicular to the flow through
the cells.
It is an object of the described embodiments of this
invention to provide a convergent light flow cell that is
compact, inexpensive and capable of high performance, and
which is particularly well suited for high performance liquid
chromatography.
Embodiments of the present invention will now be
described, by way of example, with reference to the
accompanying drawings in which:-
Figure 1 is a partial perspective view, with parts
cut away, showing one embodiment of a flow cell;
Figure 2 is a longitudinal section of the flow cell
of Figure l;
Figure 3 is a side elevation of the bottom tube
portion of the flow cell of Figures 1 and 2;
Figure 4 is a top plan view of the bottom tube of
Figure 3;
Figure 5 is a transverse section taken along the line
5-5 of Figure 2;
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Figure 6 is an optical schematic view of a liquid
chromatography detector;
Figure 7 is a top plan view of a flow cell, diagram-
matically indicating the relationship of optical stops and
the acceptance angle for light rays;
Figure 8 is an optical schematic of an alternative
embodiment of a liquid chromatography detector; and
Figure 9 is a longitudinal sectional view of a dual
beam flow cell.
With reference now to the drawings, a miniature flow
cell assembly 10 is shown for a single light beam detector.
It is formed from a standard compression fitting union 12
secured to the end of a packed liquid chromatcgraphic column
14. The flow cell assembly has open portions 16, 17 in the
- union and a cylindrical window 18 through which components
of a sample that flows through the columns can be detected.
Detection is accomplished through a light source 20 and a
photocell 22, as best shown in Figure 6. Sample components
carried by a solvent are separated by the packing in the
liquid chromatographic column 14. Because certain sample
- components absorb light, the presence of the components can
be detected by a photocell sensitive to the degree of
absorption of light by the components. An output signal
from the photocell 22 is related to the amount or rate of
change of sample in the column effluent. A dual beam flow
cell assembly 25 is shown in Figure 8, which provides two
paths and two windows for light transmission, for measuring
and comparing the sample to a standard, and thereby factoring
out solvent affects and variations of light intensity of the
source. With the basic construction of the various
embodiments a constant length light path and hig~ light
transmission is achieved through the flow cell.
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A flow cell portion 28 of the assembly 10 is loc~ated
between adjacent but spaced ends of a tube 29 forming the
column 14 and an exit tube 32, and is aligned with the
openings 16, 17 formed in the union 12, which are made by
cutting slots through the nut portion 34 of the union in a
common transverse plane to a depth that intersects a central
passage 35 of the union. This is best shown in Figure 5.
The column tube 29 is closely received in one end of the
union 12 and the juncture is sealed by a ferrule 36 and a
. nut 37 threaded to the union. The column 14 contains a
packing 39. The lower end of the column receives a porous
plug column packing retainer 40 that permits liquid passing
through the column packing to enter the flow cell and exit
through the exit tube 32. The exit tube 32 is spaced from
the plug 40 by the cylindrical quartz window 18. Two Teflon*
gaskets 42, 43 provide seals on opposite sides or ends of
the window 18.
The exit tube 32 tightly fits within the central
passage 35 of the union and provides a central passageway
45 downstream from the flow cell portion 28. Optionally, a
porous stainless steel plug or the like can be provided at
the bottom of the flow cell portion 28 to serve as a boundary
to the central passageway 45 of the exit tube 32. A reduced
diameter cylindrical boss 46 extends from the upper end of
the exit tube to tightly be received within one end of the
cylindrical window 18. The axial distance between the upper
end of the boss 46 and the lower end of the plug 40 defines
the flow cell portion 28 within the cylindrical window 18.
Two diametrically opposite aligned slots 48, 49 are formed
in the boss 46, and receive a plate 51, which extends
axially the distance between the boss 46 and the plug 40.
*Trademark
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A central slot 52 in the plate 51 extends axially the
distance between the boss 46 and plug 40, providing a path
between opposite sides of the plate, through which light
rays can pass when directed through the cylindrical window
18 on one side of the plate Sl. The plane of the plate 51
is aligned with two web portions 54, 55 ~Fig. 5) of the union
12, that are formed as a result of the cut out portions 16,
17. It will be apparent that these web portions essentially
divide the cylindrical window 18 into two cylindrical
window segments, one for light input and the other for light
output. Other means of forming optical stops will of course
be apparent to those skilled in the art, the important
feature being the provision of a narrow gap with a wide
acceptance angle (assured in the disclosed embodiment by the
use of a thin plate 51) ln the center of the pathway through
the cylindrical flow cell portion 28. A ferrule 56 and nut
57 seal and retain the exit tube in the union.
A general detector arrangement is shown in Figure 6
of .the drawings and includes the lamp 20, in this case an
ultraviolet lamp elongated in the direction shown with
respect to the orientation of the flow cell assembly 10, and
which typically can emit radiation of a prescribed wavelength,
such as 254 nm. A lens 60 is positioned to receive diverging
rays that pass through the flow cell 28 to focus them upon
the photocell 22. The locations of the lamp 20 and lens 60
are selected relative to the union 12 so that converging rays
from the lamp pass through the cut out slot 16 or 17 and
emerge through the opposite cut out slot. A W filter 62
is provided in front of the photocell 22 to limit the rays
detected to a prescribed wavelength, and the photocell 22
produces an output signal proportional to the intensity of
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the rays. The light acceptance angle of the flow cell is
controlled by the plate 51, although in some constructions
the slots 16, 17 could be a limiting factor. For good
results, the acceptance angle should be at least 45 and
preferably 90. In the embodiment shown, the acceptance
angle is approximately 90.
As best shown diagra~matically in Figure 7, a wide
acceptance angle ~ is provided as long as the slot width g
is not small with respect to the thickness t of the pllate
51. This assures that a large quantity of light, i.e.,
high light intensity, can be received through the relatively
narrow slot 52. With n the acceptance angle ~, all rays
passing through the cylindrical window 18 and the slot 52 are
essentially of equal length through the flow cell. By way
of example, with reference to Figure 7, if the flow cell
diameter d is 2.0 mm, and the plate thickness t and slot
width g are both equal to 0.5 mm, the minimum light path
length L is 1.937 mm or only 3.2% shorter than the maximum
length (i.e. diameter d) of 2.0 mm. By keeping the width g
small, the difference is minimized, and by keeping the
thickness t small, the acceptance angle is maximized. In
addition, as shown by the schematic diagram of Figure 6,
light converging from the entire length of the ultraviolet
lamp 20 will pass through the slot 52, thereby averaging
any variations in light that might occur throughout the
area of the lamp.
A schematic optical diagram of an alternative embodi-
ment of the detector is shown in Figure 8, in which the
optical stops of a flow assembly 10' are omitted and
convergence of the light rays is achieved solely through
a lens system. In Figure 8, a focusing lens 70 is provided
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to receive light rays from a spectrophotometer 71 and to
direct the rays through a flow cell portion 28', which is
identical to the flow cell 28 except for the absence of
optical stops. Light emitted through the flow cell 28'
is received by a focusing lens 72 and directed to a photo-
sensor 74. As in the previously described embodiment, all
rays passing through the flow cell 28' are normal to the
cylindrical window and pass through the center of the flow
cell so that all are of equal length.
The dual flow cell 25 embodying the present invention
is shown in detail in Figure 9 of the drawings. A union
82 is provided into which two liquid chromatographic
columns 84, 85 terminate. Each column enters the union
from an opposite direction and terminates with a porous
stainless steel plug 88, 89 respectively. Two pairs of
cut out zones gO, 91 are provided axially~spaced along the
length of the union,~ each corresponding basiCally with the
cut out porticns 16, 17 of the union 12. Two cylindrical
windows 92, 93 are received within a central passage 94 of
the union, axially spaced bv porous plugs 96, 97 and a
spacer ring 98 and central exit zone 99. A perpendicular
passage 100 extends through the wall of the union 82, at
right angles to the central passage 94, from the exit zone
99. Each zone defined by the two cylindrical windows 92,
93 and the porous plugs 88, 96 and 89, 97 comprises an
individual flow cell. Each of these cells receives liquid
from one of the columns 84, 85. After passage through the
cells, the liquid flows to the common exit zone 99 and
thence through the outlet passage 100. Separate light
beams are directed through the cut out zones 90, 91 and
are separately detected. The separate beams are directed
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from a ~single source. A sample is passed in a solvent
through one of the flow cells, and the solvent alone is
passed through the other flow cell. The light outputs from
both flow cells are detected, and the effects of the light
source and solvent are factored out by suitable comparisons.
While the operations of these different embodiments
have been described in connection with the structures
thereof, it will be apparent in summary that in all embodi-
ments the flow cell receives only convergent light rays
directed normal to the cylindrical window of the flow cell,
so that the light rays pass through the center of the flow
cell. This assures that all light rays passing through
the flow cell are essentially equal in length, are not
refracted and are gathered from a wide angle that provides
high intensity and effectively averages any variations in
light output throughout the area of the light source.
While preferred embodiments of the present invention-
have been described with particularity, it will be apparent
that various modifications and alterations may be made
therein without departing from the spirit and scope of the f
invention set forth in the appended claims.
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