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Caution:

The user shall be made aware that if the equipment is used in a manner not

specified by the manufacturer, the protection provided by the equipment may be impaired.

Attention :

L’utilisateur doit être informé que si le matériel est utilisé d’une façon non

spécifiée par le fabricant, la protection assurée par le matériel risque d’être défectueuses.

Vorsicht:

Der Benutzer wird darauf aufmerksam gemacht, dass bei unsachgemäßer

Verwenddung des Gerätes unter Umständen nicht ordnungsgemäß funktionieren.

Precauzione:

l’utente deve essere al corrente del fatto che, se l’apparecchiatura viene

usta in un modo specificato dal produttore, la protezione fornita dall’apparecchiatura
potrà essere invalidata.

Advertencia:

El usuario deberá saber que si el equipo se utiliza de forma distinta a la

especificada por el fabricante, las medidas de protección del equipo podrían ser
insuficientes.

Caution:

To protect against fire hazard, replace fuses with those of the same type and

rating.

Attention :

Remplacez toujours les fusibles par d’autres du même type et de la même

puissance afin d’éviter tout risque d’incendie.

Vorsicht:

Zum Schutz gegen Feuergefahr die Sicherungen nur mit Sicherungen des

gleichen Typs und Nennwertes ersetzen.

Precauzione:

per una buona protezione contro i rischi di incendio, sostituire i fusibili con

altri dello stesso tipo e amperaggio.

Precaución:

sustituya los fusibles por otros del mismo tipo y características para evitar

el riesgo de incendio.

Содержание 2996

Страница 1: ...Waters 2996 PDA Detector Operator s Guide 34 Maple Street Milford MA 01757 71500023202 Revision C...

Страница 2: ...or consequential damages in connection with or arising from the use of this document 2001 WATERS CORPORATION PRINTED IN THE UNITED STATES OF AMERICA ALL RIGHTS RESERVED THIS DOCUMENT OR PARTS THEREOF...

Страница 3: ...oid the user s authority to operate the equipment Important Toute modification sur cette unit n ayant pas t express ment approuv e par l autorit responsable de la conformit la r glementation peut annu...

Страница 4: ...trouvant proximit N utilisez pas de tuyau de Tefzel fortement ab m ou d form N utilisez pas de tuyau de Tefzel avec de l acide sulfurique ou nitrique ou du t trahydrofurane THT Sachez que le chlorure...

Страница 5: ...dimetilsolfossido provocano rigonfiamento nei tubi Tefzel che riducono notevolmente il limite di pressione di rottura dei tubi stessi Advertencia manipular con precauci n los tubos de polimero bajo pr...

Страница 6: ...la protezione fornita dall apparecchiatura potr essere invalidata Advertencia El usuario deber saber que si el equipo se utiliza de forma distinta a la especificada por el fabricante las medidas de p...

Страница 7: ...d effectuer la maintenance de l instrument Vorsicht Zur Vermeidung von Stromschl gen sollte das Ger t vor der Wartung abgeschaltet und vom Netz getrennt werden Precauzione per evitare il rischio di sc...

Страница 8: ...terminal Borne du cadre ou du ch ssis Rahmen oder Chassisanschluss Terminale di struttura o telaio Borne de la estructura o del chasis Caution or refer to manual Attention ou reportez vous au guide V...

Страница 9: ...ica alta tensi n Caution risk of needle stick puncture Attention risques de perforation de la taille d une aiguille Vorsicht Gefahr einer Spritzenpunktierung Precauzione rischio di puntura con ago Pre...

Страница 10: ...ling CDC NIH Manual 1984 Calibration Follow acceptable methods of calibration with pure standards to calibrate methods Use a minimum of five standards to generate a standard curve The concentration ra...

Страница 11: ...Connecting Analog Output Cables 6 1 4 2 Connecting Event Cables 7 1 5 Fluidic Connections 9 1 6 Starting Up and Shutting Down the Detector 11 Chapter 2 Diagnostics and Calibration 15 2 1 Diagnostics 1...

Страница 12: ...ta Points 35 4 4 1 Calculating Absorbance 35 4 4 2 Resolution 38 4 4 3 Filtering Data 38 Chapter 5 Spectral Contrast Theory 41 5 1 Comparing Absorbance Spectra 41 5 2 Representing Spectra as Vectors 4...

Страница 13: ...Table of Contents xiii Appendix A Detector Specifications 51 Appendix B Spare Parts 53 Appendix C Mobile Phase Absorbance 55 Index 59...

Страница 14: ...xiv Table of Contents...

Страница 15: ...ling the Flow Cell 23 3 4 Lamp Power Connector and Mounting Screws 26 3 5 Fuse Block 27 4 1 Optics Assembly Light Path 30 4 2 Benzene Spectrum at 1 2 nm Resolution 32 4 3 Photodiodes Discharged by Lig...

Страница 16: ...xvi List of Figures...

Страница 17: ...1 3 Event Out Terminal Specifications on Contact Closure 9 1 4 LED Indicator Sequences During Startup 12 2 1 2996 Detector Troubleshooting 15 4 1 Optics Assembly Components 30 A 1 2996 Detector Speci...

Страница 18: ...xviii List of Tables...

Страница 19: ...l and set up the 2996 Detector Chapter 2 describes how to troubleshoot the 2996 Detector Chapter 3 describes how to replace the flow cell the lamp and the fuse Chapter 4 explains the principles involv...

Страница 20: ...nts as part of the Millennium 32 system Also covers the installation configuration and use of acquisition servers such as the LAC E32 module the busLAC E card and interface cards used to communicate w...

Страница 21: ...at more of the same type of item can optionally follow For example You can store filename1 filename2 in each folder A right arrow between menu options indicates you should choose each option in sequen...

Страница 22: ...xxii...

Страница 23: ...t are not compatible with Millennium32 or MassLynx software control 1 1 Installation Site Requirements Install the 2996 PDA Detector at a site that meets the specifications in Table 1 1 and Figure 1 1...

Страница 24: ...requires no voltage adjustment The electrical power requirements for the Waters 2996 PDA Detector are Voltage range 100 to 240 Vac Total power 100 VA Frequency range 50 to 60 Hz Fuses The 2996 PDA De...

Страница 25: ...er end to a power outlet Figure 1 2 Detector Rear Panel 1 3 Millennium32 Workstation Connections The 2996 Detector requires signal connections to the Millennium 32 Workstation over the IEEE 488 bus Al...

Страница 26: ...rd on the workstation is not important For example you can connect the 2690 Separations Module before or after the 2996 Detector 2 Use another IEEE 488 cable to connect to the stackable connector on t...

Страница 27: ...g the IEEE 488 Address To set the IEEE 488 address for the 2996 Detector Use a small screwdriver or similar device to set the IEEE 488 switches on the detector rear panel Figure 1 4 to the IEEE 488 ad...

Страница 28: ...the Millennium 32 Help Required Materials One small flat blade screwdriver One electrical insulation stripping tool Analog signal cables included with the Waters 2996 Detector Startup Kit Procedure To...

Страница 29: ...enerally used for inject start Two output programmable event table signal terminals If an inject start signal is not available over the IEEE 488 bus you must provide a signal at Event In 1 terminal on...

Страница 30: ...eps 1 Insert the bare wires at one end of the event signal cable into the positive and negative slots of the appropriate event input or output terminal Figure 1 6 Tighten the two screws to secure the...

Страница 31: ...Low trigger 1 8 V High trigger 3 0 V Protected to 30 V Minimum pulse width 30 msec this may limit compatibility with valves that require a quick pulse Maximum current 5 mA Table 1 3 Event Out Terminal...

Страница 32: ...east 1 to 2 feet 30 to 60 cm to prevent air bubbles from forming in the flow cell 2 Cut the two lengths of tubing as follows a Use a Waters 1 16 inch stainless steel tubing cutter or a file with a cut...

Страница 33: ...in the waste container 1 6 Starting Up and Shutting Down the Detector The entire startup procedure takes less than 1 minute Once completed you should allow the 2996 Detector to warm up for at least 1...

Страница 34: ...anel of the detector Figure 1 8 If both lights remain illuminated the detector passed the internal diagnostics If either indicator light blinks or is off refer to the troubleshooting tables in Chapter...

Страница 35: ...of the detector to the 0 Off position OFF FLASHING The 2996 is running power on self tests FLASHING FLASHING 2996 has failed one of the self tests Contact Waters Technical Service OFF FLASHING The 299...

Страница 36: ...14 Installation 1...

Страница 37: ...w accessed from Run Samples in the Millennium 32 software If you encounter a problem that you cannot troubleshoot Section 2 1 Diagnostics contact Waters Technical Service at 800 252 4752 U S and Canad...

Страница 38: ...ctor waste outlet Weak lamp Replace lamp Section 3 1 2 Removing the Flow Cell Shutter failure message Shutter failed Run the Shutter diagnostic For details see Millennium 32 Help Detector not respondi...

Страница 39: ...cs from Run Samples in the Millennium 32 software For more information on Run Samples and PDA diagnostics refer to the Millennium 32 Help Change in reference spectrum Mobile phase contains gas or is c...

Страница 40: ...rate to set the 486 nm peak at the proper wavelength Ensure precise data for library matching Note The system administrator can restrict access to the PDA Calibration window by disabling access to Run...

Страница 41: ...ow fails and the lamp status light is on Table 2 1 The 2996 Detector causes high backpressure Note Conditions other than a dirty flow cell may cause decreased lamp intensity For more information refer...

Страница 42: ...n then run the pump for 10 minutes 3 Test the lamp energy by performing the Lamp diagnostic test For details refer to the Millennium 32 Help If the lamp diagnostic fails and the lamp has not been used...

Страница 43: ...he Flow Cell Assembly 6 Use the Phillips screwdriver to loosen the three thumbscrews that hold the flow cell assembly to the optics bench and the thumbscrew that secures the bracket holding the fluidi...

Страница 44: ...surface finish and the alignment of the lenses are critical to the performance of the 2996 Detector Be careful not to touch or damage the lenses and the lens holders STOP Attention To prevent lens co...

Страница 45: ...he Flow Cell 2 Use the small flat blade screwdriver to gently pry the lens assembly from the flow cell body at the slots 3 Use a lens tissue or a nonparticulating swab to wipe the lens with methanol S...

Страница 46: ...Tighten until the screws meet the flow cell disk then tighten each screw 1 4 turn If a torque screwdriver is available tighten the screws to 16 in oz 0 113 N m 6 Check for leaks If you find any leaks...

Страница 47: ...mobile phase also affects the apparent lamp intensity For example acetonitrile is more transparent than methanol at wavelengths less than 220 nm Caution To avoid electrical hazards and exposure to UV...

Страница 48: ...ay from the chassis 3 Open lamp secure panel Figure 3 4 Lamp Power Connector and Mounting Screws Caution To avoid electrical hazards when you perform the following procedure power off the 2996 Detecto...

Страница 49: ...Reconnect the lamp power connector Figure 3 4 10 Secure the lamp access panel 11 Install the front panel cover 12 Reconnect the power cord and power on the 2996 Detector 3 3 Replacing the Fuses Replac...

Страница 50: ...ock while you pull out the block 4 Remove the fuses from the block then install the new fuses 5 Position the fuse block with the small tab pointing down then push in the block until the side clips eng...

Страница 51: ...of operation of the optics and electronics of the Waters 2996 PDA Detector 4 1 2996 Detector Optics The 2996 Detector is an ultraviolet visible light UV Vis spectrophotometer with 512 photodiodes Opt...

Страница 52: ...back to a reference diode which measures the intensity of the light emitted by the lamp The detector uses this measurement to keep the lamp output constant Flow cell assembly Houses the segment of the...

Страница 53: ...mirror and mask The mirror focuses light transmitted through the flow cell onto the aperture at the entrance to the spectrographic portion of the optics The mirror mask defines the beam of light focus...

Страница 54: ...t of light striking the photodiode array to determine the absorbance of the sample in the flow cell The array consists of 512 photodiodes arranged in a row Each photodiode acts as a capacitor by holdi...

Страница 55: ...s of an individual diode is the exposure time The 2996 Detector requires less than 10 msec to sequentially read all of the diodes in the array and process the data The minimum exposure time is 10 msec...

Страница 56: ...terium light source from 190 to 800 nm To minimize detector noise Auto Exposure adjusts the exposure time from 50 to 90 of full scale The Auto Exposure time setting ensures that the photodiodes are No...

Страница 57: ...hs of interest and over which the mobile phase absorbs minimally Appendix C Mobile Phase Absorbance Setting the bandwidth to higher values can improve signal to noise ratios 4 4 Computing Absorbance D...

Страница 58: ...eer s Law applies only to well equilibrated dilute solutions It assumes that the refractive index of the sample remains constant that the light is monochromatic and that no stray light reaches the det...

Страница 59: ...ely after the dark current measurement and before any components are eluted the 2996 Detector records a reference spectrum The reference spectrum is a measure of lamp intensity and mobile phase absorb...

Страница 60: ...ple Rate Sample rate is the number of data points per second reported to the Millennium 32 database The number of times the photodiodes are read during the sample rate interval is dependent on the exp...

Страница 61: ...a good signal to noise ratio for most chromatographic separations Note the following with regard to filtering data The noise filter is a digital low pass filter The filter calculates a data point tha...

Страница 62: ...40 Principles of the 2996 PDA Detector Optics 4...

Страница 63: ...f multiple compounds in the same peaks coelution or due to nonideal conditions such as noise photometric error or solvent effects 5 1 Comparing Absorbance Spectra The shape of an absorbance spectrum i...

Страница 64: ...centration for peaks that are less than 1 0 AU across the collected wavelength range Vector direction contributes to the identification of a compound since the direction is a function of the absorbanc...

Страница 65: ...the spectrum of Compound B The vector for Compound B points in a direction different from that of the vector for Compound A The difference in direction which reflects the difference in the absorbance...

Страница 66: ...at have different shapes have vectors that point in different directions The angle between the two vectors of any two spectra the Spectral Contrast angle quantifies the magnitude of the shape differen...

Страница 67: ...ctral Contrast Angle Spectra with Similar Shapes In Figure 5 4 the absorbance spectra of two compounds A and B are similar and therefore have a small Spectral Contrast angle 3 0 Normalized Absorbance...

Страница 68: ...may exhibit slight differences because of detector noise photometric error high sample concentration or variations in solvent conditions The spectra in Figure 5 5 for example show how instrument nois...

Страница 69: ...ogeneity You can assess the significance of spectral inhomogeneity by comparing a Spectral Contrast angle to a Threshold angle Section 5 4 4 5 4 1 Detector Noise Statistical and thermal variations add...

Страница 70: ...d composition do not change isocratic operation the background absorbance if any by the solvent remains constant A change however in solvent pH or composition such as occurs in gradient operation can...

Страница 71: ...Effects of pH and Solvent Concentration on the Absorbance Spectrum of p Aminobenzoic Acid Wavelength nm Absorbance Absorbance Wavelength nm pH 6 9 pH 5 1 pH 3 1 Effect of Concentration Effect of pH No...

Страница 72: ...50 Spectral Contrast Theory 5...

Страница 73: ...to 800 nm Wavelength accuracy 1 nm Linearity range Per ASTM 685 79 5 at 2 0 AU propylparaben at 256 nm Spectral resolution 1 2 nm Baseline noise 1 5 10 5 AU peak to peak dry at 254 nm Drift 1 10 3 AU...

Страница 74: ...52 Detector Specifications A...

Страница 75: ...7919 Flow cell semi preparative WAT057463 Flow cell microbore WAT057462 Flow cell inert WAT057461 Flow cell Autopurification 289000612 Variable pathlength flow cell WAT057664 Gasket flow cell 2 WAT057...

Страница 76: ...54 Spare Parts B...

Страница 77: ...that is autozeroed out Wavelength pH and concentration of the mobile phase will affect its absorbance Examples of several mobile phases are provided in Table C 1 Table C 1 Mobile Phase Absorbance Meas...

Страница 78: ...2 42 2 50 2 45 2 37 1 96 0 50 0 12 0 04 0 01 Buffers and Salts Ammonium acetate 10 mM 1 88 0 94 0 53 0 29 0 15 0 02 0 01 Ammonium bicarbonate 10 mM 0 41 0 10 0 01 0 01 EDTA disodium 1 mM 0 11 0 07 0 0...

Страница 79: ...citrate 10 mM 2 48 2 84 2 31 2 02 1 49 0 54 0 12 0 03 0 02 0 01 Sodium formate 10 mM 1 00 0 73 0 53 0 33 0 20 0 03 0 01 Sodium phosphate 100 mM pH 6 8 1 99 0 75 0 19 0 06 0 02 0 01 0 01 0 01 0 01 0 01...

Страница 80: ...02 0 02 0 02 0 02 0 01 Detergents BRIJ 35 1 0 06 0 03 0 02 0 02 0 02 0 01 0 01 CHAPS 0 1 2 40 2 32 1 48 0 80 0 40 0 08 0 04 0 02 0 02 0 01 SDS 0 1 0 02 0 01 0 01 Triton X 100 0 1 2 48 2 50 2 43 2 42...

Страница 81: ...n IEEE 488 6 rear panel 3 terminal strip 8 Contact closures 6 Contacting Waters Technical Service 15 17 Conventions documentation xx D Dark current 37 Data acquisition Auto Exposure parameter 34 Expos...

Страница 82: ...lengths 55 N Noise effects 47 Nonidealities 47 Non IEEE 488 connections 6 O Outputs 6 8 9 P Parts spare 53 Photodiode array 32 Photometric error 48 Power connections 2 Purity Angle photometric error e...

Страница 83: ...multiple wavelengths 44 derived from two wavelengths 43 spectra representing 42 spectral contrast 42 W Waters 2996 absorbance calculations 35 37 aperture width 31 dark current 37 detector optics over...

Страница 84: ...62 Index I N D E X...

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