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(12) Patent Application Publication (10) Pub. No.: US 2005/0078308A1 Gilby (43) Pub

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(12) Patent Application Publication (10) Pub. No.: US 2005/0078308A1 Gilby (43) Pub US 2005.0078308A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2005/0078308A1 Gilby (43) Pub. Date: Apr. 14, 2005 (54) OPAQUE ADDITIVE TO BLOCK STRAY (30) Foreign Application Priority Data LIGHT IN TEFLONOOab AF LIGHT GUIDING FLOWCELLS Feb. 25, 2003 (WO)........................... PCT/US03/05811 (75) Inventor: Anthony C. Gilby, Foxborough, MA Publication Classification US (US) (51) Int. Cl. .................................. G01N 21/05 Correspondence Address: (52) U.S. Cl. .............................................................. 356/246 Anthony J. Janiuk Waters Technologies Corporation cio Waters Investments Limited (57) ABSTRACT 34 Maple Street Milford MA 01757 (US 9 (US) The present invention is directed to the use of a light (73) Assignee: WATERS INVESTMENTS LIMITED, absorbing wall material to eliminate Stray light paths in New Castle, DE light-guiding Structures, Such as those used for HPLC absor bance detection. More Specifically, the present invention (21) Appl. No.: 10/911,126 relates to the use of carbon-doped Teflon(R) AF, or “black Teflon(RAF, for all or part of the walls of a light-guiding (22) Filed: Aug. 4, 2004 flowcell adapted for use in HPLC absorbance detection. 52 Patent Application Publication Apr. 14, 2005 Sheet 3 of 5 US 2005/0078308A1 NQ ss s : 3 C S. 3 O s 3 cus Z O C 9 d Z Patent Application Publication Apr. 14, 2005 Sheet 4 of 5 US 2005/0078308A1 ent Application Publication Apr. 14, 2005 Sheet 5 of 5 SN 50 Y 'N Y ro FIG. 5 US 2005/0078308A1 Apr. 14, 2005 OPAQUE ADDITIVE TO BLOCK STRAY LIGHT IN TEFLONOOAB AF LIGHT-GUIDING FLOWCELLS AC AA (1c)1 CROSS REFERENCE TO RELATED APPLICATIONS 0007 Ac represents the smallest analyte concentration 0001. This application claims priority from a PCT Inter that can be detected and AA the corresponding Smallest national Patent Application No. PCT/US03/05811, filed Feb. change in absorbance that can be measured. This represents 25, 2003 and U.S. Provisional Patent Application No. the noise at the absorbance baseline, the output of the 60/359,354, filed Feb. 25, 2002. The contents of these absorbance detector. applications are incorporated herein by reference. 0008 As illustrated by equation (1b), low absorbance noise requires a high light signal 10 and low noise in the BACKGROUND OF THE INVENTION measurement of I, i.e. a high signal-to-noise (S/N) ratio in the raw transmittance measurement. In a well-designed 0002 The present invention relates generally to the use of detector, shot noise, which is proportional to the Square root a light absorbing wall material to eliminate Stray light paths of the light Signal, dominates, So high SIN requires highlight in light-guiding applications, Such as High Performance throughput. Liquid Chromatography (HPLC), and Capillary Zone Elec trophoresis (CZE) spectroscopic analysis. 0009 Light-guiding flowcells enable low volume cells to be constructed with high light throughput and long path 0003) Systems for light absorption detection generally length. The liquid Sample is contained in a tube of material comprise four basic components, a light Source, a means for having a lower refractive index (RI) than the mobile phase. Selecting wavelengths to be used, a light-guiding vessel, Light is introduced into one end of the tube and propagates typically in the form of a hollow tube or capillary through down the axis of the tube making multiple internal reflec which a sample to be analyzed and light are passed (a tions before emerging at the other end. The liquid is analo flowcell), and a light detector which measures the amount of gous to the core of an optical fiber and the material of the light transmitted through the flowcell. Large optical tube is analogous to the cladding. The condition for light throughput can be achieved when the light is guided along guiding is that the rays incident on the liquid/wall boundary the capillary Similar to the way light is guided along an do So at an angle of incidence greater than the critical angle optical fiber. 0004. A flowcell must be constructed from materials that are resistant to the Solutions encountered in liquid chroma 12 (2a) tography or CE. To achieve high Sensitivity to Small con centrations of analyte, the cell must have a high optical throughput and a long pathlength. If the quantity of analyte 0010 where n is the RI of the liquid, and n is the RI of is Small and capillary Separation techniques are used, the the wall of the flowcell. volume of the cell must also be Small, otherwise band Spreading and loSS of chromatographic resolution occurs. 0011 The numerical aperture (NA) of the guided beam is The transmittance, T, of light through Such a System filled given by: with a light absorbing Sample is determined in accordance with Beer's law: 0012. Where p is the largest angle, between a ray entering the cell from air and the cell axis, which meets the guiding (1a) condition. The guiding mechanism is termed total internal reflection (TIR) (1b) 0013 Recently, flowcells having an inner surface of an amorphous fluoropolymer material that has an index of refraction lower than that of common chromatography Sol vents, e.g. water, have enabled light-guiding flowcells to be 0005 where I is the light exiting the flowcell when constructed. it is filled with clear mobile phase and I is the light 0014 Light introduced along the axis of the tube is power exiting the flowcell when analyte is present. b guided in the fluid by total internal reflection at the fluid wall is the path length of the flowcell conventionally boundary. The amorphous fluoropolymer material Teflon(R) expressed in centimeters, c is the analyte concentra AF 1600 and 2400 are preferred tube materials because they tion in M or moles/liter and e is the molar absorp are transparent throughout the visible and ultraViolet Spec tivity expressed in units of cm (moles/liter). A is trum, they have an unusually low refractive index (1.31 and the absorbance, a dimensionless number expressed 1.29 respectively) and are chemically inert. As a compari in absorbance units (au). son, the RI of water at the same wavelength is 1.333. All common Solvents (as methanol/water mixtures and acetoni 0006 The requirement for highlight throughput and long trile) have a higher RI than water and therefore also Teflon(R) pathlength is illustrated by differentiating equation (1b). AF. Only pure methanol has an index slightly below water, US 2005/0078308A1 Apr. 14, 2005 but still above that of Teflon(E) AF. Even at different wave inlet and outlet ports, whereby the walls and fluid effect lengths, the fluoropolymerS retain the RI advantage. guiding of the light by attenuated total reflection, the walls having minimal absorptive effect on the guided light. The 0015. However, it is difficult to construct a cell with amorphous fluoropolymer walls without Some light entering walls have an absorption coefficient in the range of 0.1 to the end croSS-Section of the wall, or Some light 18 being 100 mm at a wavelength within the wavelength range of scattered into the wall from the liquid, or some light 18 UV, visible and near IR. A preferred material for the walls entering the fluid from the walls after bypassing part or all is Teflon(R) AF fluoropolymer doped with a black dopant of the Sample fluid. These aberrant light paths result in a Such as carbon black. A concentration of carbon black Stray light background and inaccuracy in the readings, between 0.01% and 1% by weight of the fluoropolymer is limiting the linearity and dynamic range of a detector that is sufficient to absorb stray light. Supposed to receive only light that has passed through the 0021 Another embodiment is an apparatus for housing a liquid. liquid Sample and for exposing the liquid Sample to light. 0016 One Strategy to control Stray light positions opaque The apparatus comprises a conduit having a wall formed of masks between the walls and the light, but the Small diam an amorphous fluoropolymer having a refractive indeX leSS eter tubes of HPLC and CZE equipment makes the align than the refractive index of water and having an absorption ment of Such masks difficult and time consuming. A Second coefficient of a magnitude Such that when the conduit is Strategy to control Stray light Supplies entering light through filled with water, visible and ultra-violet light can be trans mitted, Substantially without loss, along the axis of the an optical fiber having an OD that fits between the walls, but conduit by attenuated total reflection but visible and ultra the amount of light coupled into the liquid is reduced Violet light are Substantially completely absorbed in passage geometrically by the reduced area through the walls of the conduit. 0.017. The difficulty of controlling stray light becomes greater as fluid croSS-Sections are made Smaller. For capil 0022. A method of performing photometric analysis of a lary HPLC or CZE detection, a fluid channel ID of 100 um liquid Sample with improved linearity of detection com or leSS is needed to create a Small Volume flowcell, with prises introducing the liquid Sample into a conduit having a Sufficient pathlength to preserve analytical Sensitivity. A wall formed of an amorphous fluoropolymer having a refrac better way is needed to fabricate light guiding flowcells to tive indeX less than a refractive index of water and having an absorption coefficient Sufficient to Substantially attenuate avoid the difficulties of controlling stray light outlined above light propagation through the Wall, Shining light axially onto the conduit filled with Sample liquid, receiving light trans SUMMARY OF THE INVENTION ferred through the liquid Sample at a detector; and deter 0.018.
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