Waters 2998 Operator's Manual
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Waters 2998 Operator's Manual

Photodiode array detector
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2998 Photodiode Array
Detector
Operator's Guide
71500121902/Revision C
Copyright © Waters Corporation 2010
All rights reserved

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  • Page 1 2998 Photodiode Array Detector Operator’s Guide 71500121902/Revision C Copyright © Waters Corporation 2010 All rights reserved...

  • Page 2: Copyright Notice

    Corporation assumes no responsibility for any errors that may appear in this document. This document is believed to be complete and accurate at the time of publication. In no event shall Waters Corporation be liable for incidental or consequential damages in connection with, or arising from, its use.

  • Page 3: Customer Comments

    Contacting Waters ® Contact Waters with enhancement requests or technical questions regarding the use, transportation, removal, or disposal of any Waters product. You can reach us via the Internet, telephone, or conventional mail. Waters contact information Contacting medium Information...

  • Page 4: Safety Advisories

    Confirms that a manufactured product complies with all applicable United States and Canadian safety requirements Consult instructions for use Audience and purpose This guide is intended for personnel who install, operate, and maintain the Waters 2998 Photodiode Array (PDA) detector.

  • Page 5: Intended Use Of The 2998 Photodiode Array Detector

    Intended use of the 2998 Photodiode Array detector Waters designed the 2998 Photodiode Array detector to analyze and monitor various types of compounds. The 2998 PDA detector is for research use only. Calibrating To calibrate LC systems, follow acceptable calibration methods using at least five standards to generate a standard curve.

  • Page 6: Ec Authorized Representative

    EC authorized representative Waters Corporation (Micromass UK Ltd.) Floats Road Wythenshawe Manchester M23 9LZ United Kingdom Telephone: +44-161-946-2400 Fax: +44-161-946-2480 Contact: Quality manager...

  • Page 7: Table Of Contents

    Safety advisories ....................iv Operating this instrument ................. iv Applicable symbols ..................... iv Audience and purpose..................iv Intended use of the 2998 Photodiode Array detector......... v Calibrating ......................v Quality-control ..................... v ISM classification ....................v ISM Classification: ISM Group 1 Class B ............v EC authorized representative ................
  • Page 8 Monitoring detector LEDs ................2-20 Shutting down the detector ................2-21 Using a cuvette ....................2-21 3 Maintaining the Detector ..............3-1 Contacting Waters technical service ............3-2 Maintenance considerations ................3-2 Safety and handling..................3-2 Spare parts ....................... 3-3 Routine Maintenance ..................
  • Page 9 Replacing the flow cell ..................3-5 Replacing the lamp ................... 3-7 Replacing the fuses ..................3-10 4 Diagnostic Tests and Troubleshooting ..........4-1 Diagnostic tests ....................4-2 Verifying detector calibration ................. 4-2 Reading lamp energy ..................4-3 Performing the erbium calibration ..............4-3 Reading the calibration constants ..............
  • Page 10 Caution symbol ....................A-5 Warnings that apply to all Waters instruments ......... A-6 Electrical and handling symbols ..............A-12 Electrical symbols ..................A-12 Handling symbols ..................A-13 B Specifications ..................B-1 C Solvent Considerations ............... C-1 Introduction ...................... C-2 Clean solvents ....................C-2 Solvent quality ....................
  • Page 11 Wavelength selection for chromophore detection......... C-13 Mobile phase absorbance................C-14 Index ..................... Index-1 Table of Contents...
  • Page 12 Table of Contents...

  • Page 13: 2998 Pda Detector Optics Principles

    2998 PDA Detector Optics Principles To use the 2998 PDA detector effectively, you should understand the principles that underlie operation of the detector’s optics and electronics. Contents: Topic Page Detector optics Flow cell operating principles Resolving spectral data Measuring light at the photodiode array...

  • Page 14: Detector Optics

    Lamp Filter Flow cell TP02819 flag/shutter The following table describes the optics assembly components. Optics assembly components Component Function Lamp Deuterium source lamp. M1 mirror Focuses light from the deuterium source lamp. 2998 PDA Detector Optics Principles...

  • Page 15: Calculating Absorbance

    Optics assembly components (Continued) Component Function Window Used to help minimize air infiltration into the lamp housing. Filter flag/shutter Flag positions for measuring open (sample) and blocked (dark) beam energies and a third for wavelength verification. Flow cell Houses the segment of the flow path (containing eluent and sample) through which the polychromatic light beam passes.

  • Page 16: Flow Cell Operating Principles

    Concentration Flow cell operating principles The Waters TaperSlit™ flow cell used in the 2998 detector renders the detector baseline essentially insensitive to changes in mobile phase refractive index (RI). RI changes occur during gradient separations or result from temperature or pump-induced pressure fluctuations.

  • Page 17: Resolving Spectral Data

    The Waters TaperSlit flow cell, so-called because of the shape of the flow cell exit face, matches the shape of the spectrograph slit. Compared to a conventional flow cell with a cylindrical shape, the 2998 detector achieves higher light throughput for a given spectral resolution with the TaperSlit cell design.

  • Page 18: Measuring Light At The Photodiode Array

    The array consists of a row of 512 photodiodes. Each photodiode acts as a capacitor initially holding a fixed amount of charge. Light striking a photodiode discharges the diode. The magnitude of the discharge depends on the amount of light striking the photodiode. 2998 PDA Detector Optics Principles...

  • Page 19: Optimizing The Signal-To-Noise Ratio

    Photodiodes discharged by light Sample in flow cell Deuterium lamp absorbs at specific wavelengths. Grating Flow cell Light from grating dispersed onto diodes. Slit Mirror The detector measures the amount of charge required to recharge each photodiode. The charge is proportional to the amount of light transmitted through the flow cell over the interval specified by the diode exposure time.
  • Page 20 25, you should specify a higher sampling rate in the instrument method. If the value is greater than 50, you should specify a lower sampling rate in the instrument method. Example of how baseline noise increases with higher sampling rates 5 Hz 10 Hz 20 Hz 40 Hz 2998 PDA Detector Optics Principles...

  • Page 21: Computing Absorbance Data Points

    Computing absorbance data points The absorbance data points are calculated by the detector and then transmitted to the database (MassLynx or Empower). The absorbance calculation is – λ λ Absorbance ----------------------- t λ , – t λ , λ where S = sample energy D = dark energy R = reference energy...
  • Page 22 A resolution of 3.6 nm is effective for many analytes. Tip: 1-10 2998 PDA Detector Optics Principles...
  • Page 23 Resolution comparison for anthracene 0.55 0.50 0.45 0.40 0.35 0.30 0.25 0.20 0.15 0.10 0.05 0.00 3.20 3.30 3.40 3.50 3.60 3.70 3.80 3.90 4.00 4.10 4.20 252 nm at 1.2 nm, height 0.53 252 nm at 6.0 nm, height 0.45 252 nm at 12.0 nm, height 0.35 Averaging chromatographic sampling rate Sample rate is the number of data points acquired per second.

  • Page 24: Filtering Data

    No filter, 0.050 to 0.050 0.500 0.100 0.050 0.025 No filter, 0.025 to 0.025 0.250 0.050 0.025 0.0125 No filter, 0.0125 to 0.0125 0.100 For best chromatography resolution, choose Fast to avoid peak Tip: broadening. 1-12 2998 PDA Detector Optics Principles...

  • Page 25: Reference Wavelength Compensation

    Reference wavelength compensation A compensated reference wavelength collects wide-band absorbance data in a region of the spectra where no known analytes exist. It is used to reduce detector drift and wander, which can affect integration quality. The detector calculates the compensation value by averaging the absorbance values within a range of wavelengths that you select.
  • Page 26 1-14 2998 PDA Detector Optics Principles...

  • Page 27: Setting Up The Detector

    Setting Up the Detector The chapter provides the information necessary to set up the detector. Contents: Topic Page Before you begin Unpacking and inspecting Laboratory site selection Connecting to the electricity source Making signal connections Plumbing the detector 2-16 Starting up and shutting down the detector 2-18 Using a cuvette 2-21...

  • Page 28: Before You Begin

    Before you begin To install the 2998 PDA detector, you should know how, in Requirement: general, to set up and operate laboratory instruments and computer-controlled devices and also how to handle solvents. Before installing the detector, ensure that • the required components are present.

  • Page 29: Laboratory Site Selection

    Laboratory site selection To ensure the reliable operation of your detector: • do not situate it under a heating or cooling vent • connect it to a power supply that is grounded AC, 100 to 240 VAC • provide clearance of at least 15 cm (6 inches) on the back side for ventilation To avoid damaging the detector, the weight of items stacked on Caution:...

  • Page 30: Detector Dimensions

    (7.64 inches) 34.3 cm (13.5 inches) TP02753 Connecting to the electricity source The 2998 PDA detector requires a separate, grounded electricity source. The ground connection in the electrical outlet must be common and connected near the system. Avoid electrical shock: Warning: •...

  • Page 31: Making Signal Connections

    Do not turn the detector on at this time. Important: Making signal connections Waters Ethernet Instrument Getting Started Guide. See also: The following figure shows the rear panel location of the connectors used to operate the detector with external devices.

  • Page 32: Connecting The Ethernet Cable

    Ethernet cable to connect the network switch and the acquisition computer. You must install the Waters instrument control software in the acquisition computer so that the computer can control the Waters instrument. See the software installation instructions that accompany the software instrument driver disk.

  • Page 33: Network Installation Guidelines

    Network installation guidelines Configurations for multiple Waters instruments use a dedicated LAN, which requires a design based on the following guidelines: • Ethernet cable • A maximum distance of 100 meters (328 feet) You must use a network switch for multiple Ethernet Requirement: instruments.

  • Page 34: Connecting To Other Instruments

    Detector inject start connections Inject start input connection (on the Inject start output source 2998 PDA detector, connector A) Waters Alliance Separations Module Inject Start + / – Waters 717 Autosampler Inject Start + / – Waters manual injector, or Inject Start + / –...
  • Page 35 To connect the detector to other instruments, use the two analog-out/event-in (I/O) connectors and their mating connectors on the rear panel. 2998 rear panel analog-out/event-in connectors Connector B (outputs) Connector A (inputs)
  • Page 36 The following table describes the detector I/O connections. Detector analog-out/event-in connections Signal Description connections Inject start Activates timed events by triggering the run-time clock to start. Lamp off When the input is enabled, the lamps is turned off. The lamp can only be turned on by sending a new method to the detector, using the lamp button, or rebooting the detector.
  • Page 37 The firmware defaults to auto zero on inject. Tip: Detector connections to an Alliance separations module Alliance separations module 2998 PDA detector (connector A) (connector B) Pin 1 inject start (red) Pin 1 inject start + (red) Pin 2 inject start − (black)
  • Page 38 Detector connections to an Alliance separations module Alliance separations module 2998 PDA detector (connector B) (connector B) Pin 4 stop flow + (red) Pin 6 switch 1 + (red) Pin 5 stop flow –...
  • Page 39 To send an analog output signal from the detector to a chart recorder, make the connections shown in the following table and figure. Analog output connections to a chart recorder Chart recorder connectors 2998 PDA detector (connector B) Pen 1 + Pin 1 signal out + (red) Pen 1 –...
  • Page 40 Empower or MassLynx system (through a two-channel SAT/IN Module), make the connections shown in the following table and figure. Detector connections to the eSAT/IN module SAT/IN module connector 2998 PDA detector (connector B) Pin 1 signal out + (white) CHANNEL 1 Pin 2 signal out – (black)
  • Page 41 The firmware defaults to auto zero on inject. Tip: Inject start connections to an injector (pulse duration 0 to 10 seconds) 2998 PDA detector (connector A) Injector connector Pin 1, inject start + (red) Two spade lug (or similar) terminal connectors.

  • Page 42: Plumbing The Detector

    Plumbing the detector “Minimum tubing bend radius recommendations” on page C-6. See also: Warning: • To avoid chemical hazards, always observe Good Laboratory Practices when operating your system. Refer to the Material Safety Data Sheets shipped with solvents for handling information. •...
  • Page 43 Use a Waters 1/16-inch stainless steel tubing cutter or a file with a cutting edge to scribe the circumference of the tubing at the desired break point. Grasp the tubing on both sides of the scribed mark with cloth- or plastic-covered pliers (to prevent marring the surface), then gently work the tubing back and forth until it separates.

  • Page 44: Making The Gas Supply Connection

    ® Slide the 3/8-inch OD Tygon tubing, supplied in the startup kit, over the barbed drain fitting located on the drip tray and route it to a suitable waste container. Making the gas supply connection The detector can be connected to a nitrogen source to improve operation at lower wavelengths.

  • Page 45: Starting Up The Detector

    Starting up the detector To start up the detector: In your instrument method, set the solvent delivery system or pump to deliver 10 mL of HPLC-grade water. For details, refer to the Empower or MassLynx online Help. Guidelines: • Use only thoroughly degassed HPLC-grade solvents. Gas in the mobile phase may form bubbles in the flow cell and cause the detector to fail the reference energy diagnostic test.

  • Page 46: Monitoring Detector Leds

    Wait 1 hour for the detector to stabilize before acquiring data. If the detector fails to stabilize, see Chapter Detector indicator lights On/off switch Power Lamp indicator indicator TP02757 Monitoring detector LEDs Light-emitting diodes on the detector indicate its state of functioning. Power LED The power LED, on the detector’s front panel, indicates when the instrument is powered-on or powered-off.

  • Page 47: Shutting Down The Detector

    Indicates a detector failure that prevents further operation. Power-off the detector, and then power-on. If the LED is still steady red, contact your Waters service representative. Shutting down the detector To shut down the detector: If the mobile phase contains buffers, set the solvent delivery system or pump to deliver 10 mL of HPLC-grade water.
  • Page 48 2998 PDA Detector cuvette holder with the cuvette inserted Frosted sides of cuvette facing up and down Because the measurement is actually a composite of both the Restriction: contents of the cuvette and the flow cell, you need to perform cuvette measurements under identical flow cell conditions.
  • Page 49 Cuvette measuring procedure To begin a cuvette measurement: Remove the detector front left panel cover. To remove the cuvette holder, slide it toward you. Cuvette holder With the spring guide facing you, gently insert the cuvette (containing eluent) up and under the guide, with the cap facing upward (into the holder) and a frosted side of the cuvette facing up.
  • Page 50 Reinstall the detector front left panel cover: • To prevent invalid subsequent chromatographic results, remove the cuvette from the detector, and replace the empty holder after running your cuvette measurements. • To maintain optimum system performance, replace the front left panel cover before resuming normal operation of the detector.
  • Page 51 Maintaining the Detector This chapter describes the routine maintenance procedures you should perform on the detector. Contents: Topic Page Contacting Waters technical service Maintenance considerations Routine Maintenance Maintaining the flow cell Replacing the lamp Replacing the fuses 3-10...

  • Page 52: Maintaining The Detector

    Waters Technical Service (800 252-4752). Otherwise, phone the Waters corporate headquarters in Milford, Massachusetts (USA) or contact your local Waters subsidiary. Our Web site includes phone numbers and e-mail addresses for Waters locations worldwide. Go to www.waters.com, and click About Waters >...

  • Page 53: Spare Parts

    Spare parts Waters recommends that you replace only parts mentioned in this document. For spare parts details, see the Waters Quality Parts Locator on the Waters web site’s Services/Support page or the 2998 PDA Detector spare parts list (part number 71500121906).

  • Page 54: Maintaining The Flow Cell

    Test the lamp energy by performing the Read energy diagnostic test (see page 4-3). If the lamp diagnostic test fails and the lamp has not been used more than 2000 hours or 1 year from date of purchase (whichever comes first), call Waters Technical Service (see page 3-2). Maintaining the Detector...

  • Page 55: Replacing The Flow Cell

    Stop the solvent flow. Lift up the detector front cover and pull the front cover from the detector chassis. Disconnect the detector’s inlet tubing from the column outlet connection. 2998 PDA detector analytical flow cell Lamp Flow cell handle Thumbscrews...
  • Page 56 • Use a 1/4-inch flat-blade screwdriver to loosen the three thumbscrews on the flow cell assembly’s front plate. • Grasp the handle and gently pull it toward you. Unpack and inspect the new flow cell. Square the flow cell assembly in front of the opening, and then insert it into the optics bench.

  • Page 57: Replacing The Lamp

    Change the lamp when it repeatedly fails to ignite or when the detector fails to calibrate. The 2998 detector source lamp is warranted to light and pass startup diagnostic tests for 2000 hours or 1 year from the date of purchase, whichever comes first.
  • Page 58 Lamp gas is under slight negative pressure. To prevent Warning: shattering the glass, use care when disposing of the lamp. Waters suggests that you adequately cushion an old lamp by containing it in the packaging of its replacement before you dispose of it.
  • Page 59 Inspect the new lamp and lamp housing. Position the lamp so that the cut-out on the lamp base plate is at the 1 o’clock position, in line with the alignment pin on the lamp housing, and then gently push the lamp forward until it bottoms into position. Ensure that it is flush to the optics bench.

  • Page 60: Replacing The Fuses

    Type the serial number for the new lamp (see the label attached to the lamp connector wire), and then click OK. Replacing the fuses To avoid electric shock, power-off and unplug the 2998 Warning: PDA detector before examining the fuses. For continued protection against fire, replace fuses with those of the same type and rating only.
  • Page 61 Removing the fuseholder Fuses Power entry module Fuse holder TP02523 Remove and discard the fuses. Make sure that the new fuses are properly rated for your requirements, and then insert them into the holder and the holder into the power entry module, gently pushing until the assembly locks into position.
  • Page 62 3-12 Maintaining the Detector...
  • Page 63 Diagnostic Tests and Troubleshooting Consult this chapter when troubleshooting problems with the 2998 PDA detector. However, bear in mind that the detector measures only the bulk properties of a system. Therefore, the cause of an apparent detector problem may actually originate with the chromatography or other system instruments.

  • Page 64: Diagnostic Tests And Troubleshooting

    Diagnostic tests The 2998 PDA detector automatically runs a series of internal diagnostic tests upon start up. The indicator LEDs on the front of the detector and messages at the Empower workstation show the results of the start up internal diagnostic tests.

  • Page 65: Reading Lamp Energy

    Ensure that the flow cell is clean before you read lamp energy. To read the lamp energy: In the console, select 2998 PDA Detector from the system tree. In the PDA detector information window, click Maintain > Read energy > Read. The Read Energy dialog box appears.

  • Page 66: Reading The Calibration Constants

    Displaying PDA detector rear panel interface connections You can use the console to determine the status of input/output signal connections or contact closures on the rear panel of the 2998 PDA detector. This display gives you a real-time status of the instrument’s signal connections.

  • Page 67: Changing The Rear Panel Interface Connections

    Opening or closing output connections can be useful when you need to start or stop an injection or troubleshoot system connectivity. To change PDA detector rear panel interface connections: In the console, select 2998 PDA Detector from the system tree. In the PDA detector information window, click Troubleshoot > Rear Panel.

  • Page 68: General Troubleshooting

    Most detector problems are relatively easy to correct. If, after running the diagnostic functions applicable to your problem and troubleshooting the detector, you cannot correct an error condition, contact Waters’ Technical Service department. Power surges Power surges, line spikes, and transient energy sources can adversely affect detector operations.

  • Page 69: Clearing Bubbles From The Flow Cell

    Flashing red Indicates an error stopped the detector. Information regarding the error that caused the failure can be found in the console log. 2998 PDA Detector online Help See also: Constant red Indicates a detector failure that prevents further operation. Power-off the detector, and then power-on.
  • Page 70 PDA detector troubleshooting (Continued) Symptom Possible cause Corrective action Change in reference Mobile phase contains Prepare fresh mobile spectrum gas or is contaminated phase and degas thoroughly. Air bubbles trapped in Reseat and check flow cell alignment of flow cell. Flush the flow cell (see page 3-4), or apply...
  • Page 71 PDA detector troubleshooting (Continued) Symptom Possible cause Corrective action Lamp light flashes red Failed startup Reseat and check and power light diagnostic tests alignment of flow cell constant green (see page 3-5). Flush the flow cell (see page 3-4). Dirty flow cell causing Flush the flow cell (see shutter diagnostic test page...
  • Page 72 PDA detector troubleshooting (Continued) Symptom Possible cause Corrective action Solvent in drain line Leak from flow cell Replace the flow cell gasket (see page 3-5). Leak from flow cell Check fittings for inlet and outlet fittings overtightening or undertightening, and replace fittings if necessary.
  • Page 73 Spectral Contrast Theory The spectral contrast algorithm compares the UV/Vis absorbance spectra of samples the detector collects. This chapter describes the theory on which the algorithm is based, explaining how it exploits differences in the shapes of the absorbance spectra. It also explains how spectral contrast represents those spectra as vectors, determining whether differences among them arise from the presence of multiple compounds in the same peaks (coelution) or from nonideal conditions...

  • Page 74: Spectral Contrast Theory

    Comparing absorbance spectra When measured at specific solvent and pH conditions, the shape of a compound’s absorbance spectrum characterizes the compound. The varying extent of UV/Vis absorbance occurring at different wavelengths produces a unique spectral shape. The following figure shows the absorbance spectra for two compounds, A and B.

  • Page 75: Vectors Derived From Two Wavelengths

    • Length – Proportional to analyte concentration. • Direction – Determined by the relative absorbance of the analyte at all wavelengths (its absorbance spectrum). Direction is independent of concentration for peaks that are less than 1.0 absorbance units (AU) across the collected wavelength range. Vector direction contributes to the identification of a compound, because the direction is a function of the absorbance compound’s spectrum.

  • Page 76: Vectors Derived From Multiple Wavelengths

    Compound A), at the two wavelengths represented by each axis. The remaining vector is similarly derived from the spectrum of Compound B. Compound B’s vector points in a direction different from Compound A’s. θ Expressed by the spectral contrast angle ( ), this difference reflects the difference between the two compounds’...
  • Page 77 To illustrate the relationship between the spectral contrast angle and spectral shape differences, consider the pairs of spectra shown in the next three figures. Spectra with different shapes In the following figure, the absorbance spectra of two compounds, A and B, are distinctly different.
  • Page 78 Spectra with similar shapes In the following figure, the absorbance spectra of two compounds, A and B, are similar. They therefore produce a small spectral contrast angle (3.0°). Spectra with a small spectral contrast angle Spectral contrast angle: 3.0° Compound A Compound B Wavelength (nm) Differences between spectra of the same compound...

  • Page 79: Undesirable Effects

    Normalized absorbance spectra of a compound at two concentrations Normalized spectra of a compound at different concentrations Spectral contrast angle: 3.4° Region of little or no analyte absorption Wavelength (nm) Undesirable effects Shape differences between absorbance spectra can be caused by one or more of the following undesirable effects: •...

  • Page 80: Detector Noise

    Detector noise Statistical and thermal variations add electronic noise to the detector’s absorbance measurements. The noise, which manifests itself as fluctuations in the baseline, is known as baseline noise. The magnitude of any absorbance differences caused by statistical and thermal variations can be predicted from the instrument noise in the baseline region of a chromatogram.
  • Page 81 Comparison of a spectral contrast angle to its threshold angle can help determine whether the shape difference between spectra is significant. In general, a spectral contrast angle less than its threshold angle indicates that shape differences are attributable to nonideal phenomena alone and that no evidence exists for significant differences between the spectra.
  • Page 82 5-10 Spectral Contrast Theory...
  • Page 83 Safety Advisories Waters instruments display hazard symbols designed to alert you to the hidden dangers of operating and maintaining the instruments. Their corresponding user guides also include the hazard symbols, with accompanying text statements describing the hazards and telling you how to avoid them.

  • Page 84: A Safety Advisories

    Heed all warnings when you install, repair, and operate Waters instruments. Waters assumes no liability for the failure of those who install, repair, or operate its instruments to comply with any safety precaution.

  • Page 85: Specific Warnings

    The following warnings can appear in the user manuals of particular instruments and on labels affixed to them or their component parts. Burst warning This warning applies to Waters instruments fitted with nonmetallic tubing. Pressurized nonmetallic, or polymer, tubing can burst. Warning: Observe these precautions when working around such tubing: •...
  • Page 86 Also ensure a gas-fail connection is connected to the LC system so that the LC solvent flow stops if the nitrogen supply fails. Mass spectrometer shock hazard This warning applies to all Waters mass spectrometers. To avoid electric shock, do not remove the mass spectrometer’s Warning: protective panels.

  • Page 87: Caution Symbol

    Biohazard warning This warning applies to Waters instruments that can be used to process material that might contain biohazards: substances that contain biological agents capable of producing harmful effects in humans. Waters instruments and software can be used to analyze or...

  • Page 88: Warnings That Apply To All Waters Instruments

    Warnings that apply to all Waters instruments When operating this device, follow standard quality control procedures and the equipment guidelines in this section. Attention: Changes or modifications to this unit not expressly approved by the party responsible for compliance could void the user’s authority to operate the equipment.
  • Page 89 • Keine Schläuche verwenden, die stark geknickt oder überbeansprucht sind. • Nichtmetallische Schläuche nicht für Tetrahydrofuran (THF) oder konzentrierte Salpeter- oder Schwefelsäure verwenden. • Durch Methylenchlorid und Dimethylsulfoxid können nichtmetallische Schläuche quellen; dadurch wird der Berstdruck des Schlauches erheblich reduziert. Warnings that apply to all Waters instruments...
  • Page 90 Attenzione: fare attenzione quando si utilizzano tubi in materiale polimerico sotto pressione: • Indossare sempre occhiali da lavoro protettivi nei pressi di tubi di polimero pressurizzati. • Spegnere tutte le fiamme vive nell'ambiente circostante. • Non utilizzare tubi eccessivamente logorati o piegati. •...
  • Page 91 농축 질산 또는 황산과 함께 사용하지 마십시오. • 염화 메틸렌(Methylene chloride) 및 디메틸술폭시드(Dimethyl sulfoxide)는 비금속 튜브를 부풀려 튜브의 파열 압력을 크게 감소시킬 수 있으므로 유의하십시오. 警告：圧力のかかったポリマーチューブを扱うときは、注意してください。 • 加圧されたポリマーチューブの付近では、必ず保護メガネを着用してください。 • 近くにある火を消してください。 • 著しく変形した、または折れ曲がったチューブは使用しないでください。 • 非金属チューブには、テトラヒドロフラン(THF)や高濃度の硝酸または硫酸などを流 さないでください。 • 塩化メチレンやジメチルスルホキシドは、非金属チューブの膨張を引き起こす場合が あり、その場合、チューブは極めて低い圧力で破裂します。 Warnings that apply to all Waters instruments...
  • Page 92 Warning: 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.
  • Page 93 警告 : 為了避免火災， 更換保險絲時， 請使用與儀器保險絲蓋旁面板上所印刷之相同類 型與規格的保險絲。 警告 : 为了避免火灾，应更换与仪器保险丝盖旁边面板上印刷的类型和规格相同的 保险丝。 경고: 화재의 위험을 막으려면 기기 퓨즈 커버에 가까운 패널에 인쇄된 것과 동일한 타입 및 정격의 제품으로 퓨즈를 교체하십시오. 警告 : 火災予防のために、ヒューズ交換では機器ヒューズカバー脇のパネルに記 載されているタイプおよび定格のヒューズをご使用ください。 Warnings that apply to all Waters instruments A-11...

  • Page 94: Electrical And Handling Symbols

    Electrical and handling symbols Electrical symbols These can appear in instrument user manuals and on the instrument’s front or rear panels. Electrical power on Electrical power off Standby Direct current Alternating current Protective conductor terminal Frame, or chassis, terminal Fuse Recycle symbol: Do not dispose in municipal waste.

  • Page 95: Handling Symbols

    Handling symbols These handling symbols and their associated text can appear on labels affixed to the outer packaging of Waters instrument and component shipments. Keep upright! Keep dry! Fragile! Use no hooks! Electrical and handling symbols A-13...
  • Page 96 A-14 Safety Advisories...

  • Page 97: B Specifications

    Specifications This appendix contains product specifications for the 2998 PDA detector. Physical specifications Attribute Specification Height 19.4 cm (7.64 inches) Depth 61 cm (24.0 inches) Width 34.3 cm (13.5 inches) Weight 12.6 kg (27.75 pounds) Environmental specifications Attribute Specification Operating temperature 4 to 40 °C (39.2 to 104 °F)
  • Page 98 Electrical specifications (Continued) Attribute Specification Normal (IPXO) Moisture protection Grounded AC Line voltages, nominal Voltage range 100 to 240 VAC nominal Frequency 50 to 60 Hz Fuse Two fuses, 100 to 240 VAC, 50 to 60-Hz, F 3.15-A, 250-V (fast-blow), 5 ×...
  • Page 99 Performance specifications (Continued) Attribute Specification Noise – UV (dry) 10 µAU peak to peak Filter = 1 second, 30-second segments Wavelength = 254 nm Bandwidth = 3.6 nm (3-pixel bunch) Flow cell = analytical, 10 mm Data rate = 2 Hz 10 µAU peak to peak Noise –...
  • Page 100 Waters 2998 PDA flow cell specifications Tubing internal Pressure diameter Pathlength Description Volume (μL) rating (inches) (mm) (bar/psi) Inlet Outlet Analytical 0.010 0.010 69/1000 Auto-purification • analytical 12.4 0.010 0.040 138/2000 • prep 0.040 Microbore 0.005 0.005 69/1000 Semi-preparative 18.3 0.020...

  • Page 101: C Solvent Considerations

    Solvent Considerations This appendix contains certain solvent considerations you must take into account when operating or maintaining the 2998 PDA Detector. Contents: Topic Page Introduction Solvent miscibility Buffered solvents Head height Minimum tubing bend radius recommendations Solvent viscosity Mobile phase solvent degassing...

  • Page 102: Introduction

    Introduction Clean solvents Clean solvents provide reproducible results and permit you to operate with minimal instrument maintenance. A dirty solvent can cause baseline noise and drift. It can also block the solvent filters with particulate matter. Solvent quality Use HPLC-grade solvents for the best possible results. Filter solvents through 0.45-µm filters before their use.

  • Page 103: Tetrahydrofuran

    Tetrahydrofuran When using unstabilized tetrahydrofuran, ensure that your solvent is fresh. Previously opened bottles of tetrahydrofuran contain peroxide contaminants, which cause baseline drift. Tetrahydrofuran contaminants (peroxides) are potentially Warning: explosive if concentrated or taken to dryness. Solvent miscibility Before you change solvents, refer to the table below to determine the miscibility of the solvents to be used.
  • Page 104 Solvent miscibility (Continued) Boiling Miscibility λ Cutoff Polarity Viscosity Point °C Solvent Number CP, 20 °C Index (nm) (1 atm) Isopropyl ether 0.33 68.3 –– Toluene 0.59 100.6 P-xylene 0.70 138.0 Benzene 0.65 80.1 Benzyl ether 5.33 288.3 –– –– Methylene chloride 0.44 39.8...

  • Page 105: Solvent Miscibility

    Solvent miscibility (Continued) Boiling Miscibility λ Cutoff Polarity Viscosity Point °C Solvent Number CP, 20 °C Index (nm) (1 atm) Dimethylsulfoxide 2.24 189.0 –– Methanol 0.60 64.7 Formamide 3.76 210.5 –– Water 1.00 100.0 –– –– How to use miscibility numbers Use miscibility numbers (M-numbers) to predict the miscibility of a liquid with a standard solvent (see page...

  • Page 106: Buffered Solvents

    Buffered solvents When using a buffer, use a good quality reagent and filter it through a 0.45-µm filter. Do not leave the buffer stored in the system after use. Flush all fluid line pathways with HPLC-quality water before shutting the system down and leave distilled water in the system (flush with 90% HPLC-quality water: 10% methanol for shutdowns scheduled to be more than one day).

  • Page 107: Solvent Viscosity

    Minimum bend radius for stainless steel tubing (Continued) Tubing size (OD) Minimum bend radius 1/8-inch tubing 1/2-inch Solvent viscosity Generally, viscosity is not important when you are operating with a single solvent or under low pressure. However, when you are running a gradient, the viscosity changes that occur as the solvents are mixed in different proportions can result in pressure changes during the run.

  • Page 108: Solvent Degassing Methods

    • the chemical affinity of the gas for the liquid. • the temperature of the liquid. • the pressure applied to the liquid. Changes in the composition, temperature, or pressure of the mobile phase can all lead to outgassing. Effects of intermolecular forces Nonpolar gases (N , CO , He) are more soluble in nonpolar solvents than...

  • Page 109: Sparging

    Sparging Sparging removes gases from solution by displacing dissolved gases in the solvent with a less soluble gas, usually helium. Well-sparged solvent improves pump performance. Helium sparging brings the solvent to a state of equilibrium, which may be maintained by slow sparging or by keeping a blanket of helium over the solvent.

  • Page 110: Wavelength Selection

    To increase the length of membrane, you can connect two or more vacuum chambers in series. The in-line degasser is available as an option or factory-installed in the ® Waters 2695 Separations Module, XE model. Wavelength selection This section includes UV cutoff ranges for •...
  • Page 111 UV cutoff wavelengths for common chromatographic solvents (Continued) UV Cutoff UV Cutoff Solvent Solvent (nm) (nm) Amyl alcohol Isopropyl ether Amyl chloride Methanol Benzene Methyl acetate Carbon disulfide Methyl ethyl ketone Carbon tetrachloride 265 Methyl isobutyl ketone Chloroform Methylene chloride Cyclohexane n-Pentane Cyclopentane...

  • Page 112: Mixed Mobile Phases

    Triton-X™ 100, 0.1% Hydrochloric acid, 0.1% ® Waters PIC Reagent A, 1 vial/liter MES, 10 mM, pH 6.0 Waters PIC Reagent B-6, 1 vial/liter Potassium phosphate, Waters PIC Reagent B-6, monobasic, 10 mM low UV, 1 vial/liter dibasic, 10 mM...
  • Page 113 Wavelength selection for chromophore detection Certain functional groups found in most compounds absorb light selectively. These groups, known as chromophores, and their behavior can be used to categorize the detection of sample molecules. The table below lists some common chromophores, and their detection wavelengths ( ), as well as the molar absorptivity (ε...
  • Page 114 Electronic absorption bands of representative chromophores (Continued)* λ ∈ λ ∈ Chemical Chromophore Configuration (nm) (L/m/cm) (nm) (L/m/cm) Nitro —NO 2 strong Nitrile —ONO 220-230 1000-2000 300-400 10 —N=N— 285-400 3-25 Nitroso —N=O Nitrate —ONO 2 (shoulde Allene —(C=C) 2 — 210-230 21,000 (acyclic)
  • Page 115 absorbance is due only to the sample. Absorbance by the mobile phase also reduces the linear dynamic range of the detector by the amount of absorbance the autozero function cancels, or “autozeroes,” out. Wavelength, pH, and concentration of the mobile phase affects its absorbance. Examples of several mobile phases are given in the table below.
  • Page 116 Mobile phase absorbance measured against air or water (Continued) Absorbance at specified wavelength (nm) Acids and bases Acetic acid, 2.61 2.63 2.61 2.43 2.17 0.87 0.14 0.01 <0.01 — Hydro- 0.11 0.02 <0.01 — — — — — — — chloric acid, 0.1% Phosphoric...
  • Page 117 Mobile phase absorbance measured against air or water (Continued) Absorbance at specified wavelength (nm) Potassium 0.03 <0.01 — — — — — — — — phosphate, monobasic 10 mM Potassium 0.53 0.16 0.05 0.01 <0.01 — — — — — phosphate, dibasic, 10 mM...
  • Page 118 Mobile phase absorbance measured against air or water (Continued) Absorbance at specified wavelength (nm) ® ® Waters reagents PIC A, 0.67 0.29 0.13 0.05 0.03 0.02 0.02 0.02 0.02 <0.01 1 vial/L PIC B6, 1 2.46 2.50 2.42 2.25 1.83 0.63...
  • Page 119 Index external analog data collection device 2-13 absorbance inject start calculating injection trigger signals 2-15 maximum input and output photometric error manual injector spectra, comparing MassLynx data system 2-14 absorbance data points, computing signal cables acids C-16 cooling time, lamp analog signals 2-13 cuvette...
  • Page 120 unpacking solubility – detergents C-18 generating dimensions inject start 2-10 stop flow 2-12 EC Authorized Representative electrical handling symbols A-13 specifications hardware, preparing electrical symbols A-12 electricity source, connecting inject start Empower data system, connecting 2-14 connection environmental specifications generating 2-10 equipment guidelines signal...
  • Page 121 changing displaying maintaining, flow cell reference maintenance spectrum considerations wavelength 1-13 routine removing mass spectrometer shock hazard cuvette 2-24 MassLynx data system, connecting flow cell 2-14 lamp match angle, photometric error effects replacing cuvette holder 2-24 maximum absorbance flow cell miscibility of solvents –...
  • Page 122 UV cutoff C-10 – C-12 viscosity considerations TaperSlit flow cell solvent angle, photometric error effects threshold angle transient energy solvent changes troubleshooting spare parts contacting Waters sparging detector specifications diagnostic functions electrical tubing environmental bend radius, minimum flow cell cutting 2-16...

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