Finnigan SpectraSYSTEM RI-150

Transcription

1 Finnigan SpectraSYSTEM RI-150 Refractive Index Detector Reference Manual A Revision A May 2004

2 Finnigan, PushLoop, and SpectraSYSTEM are trademarks of Thermo Electron Corporation. This manual and the instruments to which it applies have satisfied the requirements for CSA, FCC, the EMC, and Low Voltage Directives. Use of non-approved components and repair methods may reduce or invalidate the built-in protection that is required to meet the above certifications. Technical information contained in this publication is for reference purposes only and is subject to change without notice. Every effort has been made to supply complete and accurate information; however, Thermo Electron Corporation assumes no responsibility and will not be liable for any errors, omissions, damage, or loss that might result from any use of this manual or the information contained therein (even if this information is properly followed and problems still arise). This publication is not part of the Agreement of Sale between Thermo Electron Corporation and the purchaser of an LC system. In the event of any conflict between the provisions of this document and those contained in Thermo Electron Corporation s Terms and Conditions, the provisions of the Terms and Conditions shall govern. System Configurations and Specifications supersede all previous information and are subject to change without notice. Printing History: Revision A printed in May 2004 The products of Thermo Electron San Jose are produced under ISO 9001 accredited quality management systems. Australia: P.O. Box 239 Rydalmere Unit 14, South Street Rydalmere, N.S.W [61] (02) Austria: Wehlistrasse 27b A-1200 Wien [43] (01) Belgium: Technologiestraat 47 B-1082 Brussels [32] (02) Canada: 5716 Coopers Avenue, Unit 1 Mississauga, Ontario L4Z 2E8 [1] (905) France: 16 Avenue du Québec Silic 765 Z.A. de Courtaboeuf F Les Ulis Cédex [33] (01) Germany: Im Steingrund 4-6 D Dreieich [49] (06103) Italy: Strada Rivoltana I Rodano (Milano) [39] (02) Japan: C-2F 3-9, Moriya-cho, Kanagawa-ku Yokohama, Kanagawa [81] (45) Japan: Esaka Grand Building Esaka-cho, Suita City Osaka [81] (06) Netherlands: Takkebijsters 1 NL-4817 BL Breda [31] (076) P.R. China: Room 901, Ping-an Mansion No. 23, Jin Rong Street Xi Cheng District Beijing [86] (010) Spain: Sepulveda 7 A ES Alcobendas (Madrid) [34] (091) Spain: Acer Edificio Sertram Planta 2, Modulo 3 ES Barcelona [34] (093) Sweden: Pyramidbacken 3 S Kungens Kurva (Stockholm) [46] (08) United Kingdom: Stafford House 1 Boundary Park Boundary Way Hemel Hempstead Hertfordshire HP2 7GE [44] (01442) U.S.A.: 355 River Oaks Parkway San Jose, CA [1] (408) Notes: The country code is enclosed in square brackets [ ]. The city code or area code is enclosed in parenthesis ( ). For countries other than the U.S.A., when you are dialing from within the specified country, dial the 0 of the city code. For countries other than Italy, when you are dialing from outside the country, do not dial the 0 of the city code. Published by Technical Publications, Thermo Electron Corporation, San Jose, California. Copyright 2004 Thermo Electron Corporation. All rights reserved. Printed in the United States of America.

3 Customer Registration... Register now and receive all the privileges associated with being a Thermo Electron, Finnigan product user, including application reports and technical reports. Name Title Company Address City/State/Postal Code Country Telephone Ext. SpectraSYSTEM RI-150 Detector Serial # Date Purchased Tell us more... Let us know more about how you use this product: My Organization Is: (Check one only) My Primary Application Is: (Check one only) Commercial (for profit) lab Analytical Government lab Biomedical Hospital / Clinic Clinical / Toxicology Research Institute Energy University / College Food / Agriculture Veterinary Forensic / Toxicology Other Pharmaceutical Research / Education Job Function: (Check one only) Other Administration Lab Management Operator Other Reader Survey... Help us to improve the quality of our documentation by answering a few questions: Finnigan SpectraSYSTEM RI-150 Refractive Index Detector Reference Manual Strongly Agree Agree Disagree Revision A A Strongly Disagree The manual is well organized The manual is clearly written The manual contains all of the information I need The instructions are easy to follow The instructions are complete The technical information is easy to understand The figures are helpful I was able to operate the system by using this manual. (If not, please comment below.) Additional Comments: (Attach additional sheets if necessary.) Tear this sheet from the manual, fold it closed, stamp it, and drop it in the mail.

4 From Place Stamp Here fold EDITOR, TECHNICAL PUBLICATIONS THERMO ELECTRON SAN JOSE 355 RIVER OAKS PARKWAY SAN JOSE, CA UNITED STATES OF AMERICA fold

5 Contents Customer Support... iii Startup Checklist... vii Getting Started Features Front Panel Back Panel Side Panel Bottom Panel LCD Display (User Screens) Error Messages Installation Unpacking Standard Accessories Installing the Detector Operation Before You Begin Operating Instructions (General) Suggestions Validation Test QuickStart Theory of Operation Optical System Purging the Flowcell Electrical System Troubleshooting Common Problems Troubleshooting Table Service and Preventive Maintenance Service Preventive Maintenance Specifications... A-1 Accessories and Replacement Parts...B-1 05/2004 Thermo Electron i

6 Figures Figure 1-1 Front panel Figure 1-2 Back panel Figure 1-3 Side panel Figure 1-4 Bottom panel Figure 2-1 Loosening locking screws Figure 2-2 Integrator connections Figure 2-3 Recorder connections Figure 2-4 Terminal block connections Figure 4-1 Deflection type refractive index detector Figure 4-2 Optical system Figure 4-3 Movement of the image on the photodiode Figure 4-4 Solvent path schematic Figure 4-5 Electrical system Figure 6-1 Fuses Tables Table 1-1 Front Panel Table 1-2 Back panel Table 1-3 Side panel Table 1-4 Parameters for SpectraSYSTEM RI-150 refractive index detector Table 2-1 Standard accessories Table 3-1 Chromatography conditions for Quick Start Table 4-1 Deflection type refractive index detector Table 4-2 Optical system Table 4-3 Movement of the image on the photodiode Table 4-4 Solvent Path Table 4-5 Electrical System Table B-1 Accessories and replacement parts...b-1 ii Thermo Electron 05/2004

7 Customer Support Thermo Electron San Jose products are supported by Thermo Electron San Jose Customer Service Engineers with customer support available in North America, in Europe, and in Australasia and Asia. IN NORTH AMERICA In North America, Thermo Electron San Jose Customer Service Engineers are available from the following offices: Northeastern Region Phone [1] (732) Fax [1] (732) Southern Region Phone [1] (770) Fax [1] (770) Central Region Phone [1] (847) Fax [1] (847) Western Region Phone [1] (408) Fax [1] (408) Canada Phone [1] (905) Fax [1] (905) /2004 Thermo Electron iii

8 REPLACEABLE PARTS Contact Customer Service Operations to order replaceable parts. The location and telephone and fax numbers for North America are as follows: North America Customer Service Operations 1400 Northpoint Parkway, Suite 10 West Palm Beach, FL Phone [1] (800) Fax [1] (561) TECHNICAL SUPPORT You can contact Technical Support at the following location, telephone and fax numbers, and address: North America Technical Support Operations 1400 Northpoint Parkway, Suite 10 West Palm Beach, FL Phone [1] (800) Fax [1] (561) iv Thermo Electron 05/2004

9 IN EUROPE In Europe, customer support, replaceable parts, and technical support are available from each of the following offices. Technical support is also available from North America Technical Support Operations at the following phone number and address: Phone [1] (561) Wien (Vienna), Austria Phone [43] (01) Fax [43] (01) Brussels, Belgium Phone [32] (02) Fax [32] (02) Les Ulis Cédex, France Phone [33] (01) Fax [33] (01) Dreieich, Germany Phone [49] (06103) Fax [49] (06103) Milano, Italy Phone [39] (02) Fax [39] (02) Breda, Netherlands Phone [31] (076) Fax [31] (076) Madrid, Spain Phone [34] (091) Fax [34] (091) Barcelona, Spain Phone [34] (093) Fax [34] (093) Stockholm, Sweden Phone [46] (08) Fax [46] (08) Hemel Hempstead, United Kingdom Phone [44] (01442) Fax [44] (01442) For all other countries, contact your local Thermo Electron San Jose products dealer. 05/2004 Thermo Electron v

10 IN AUSTRALASIA AND ASIA In Australasia and Asia, customer support, replaceable parts, and technical support are available from each of the following offices. Technical support is also available form North America Technical Support Operations at the following phone number and address: Phone [1] (561) Rydalmere, N.S.W., Australia Phone [61] (02) Fax [61] (02) Yokohama, Japan Phone [81] (45) Fax [81] (06) Osaka, Japan Phone [81] (06) Fax [81] (06) Beijing, P.R. China Phone [86] (010) Fax [86] (010) For all other countries, contact your local Thermo Electron San Jose products dealer. vi Thermo Electron 05/2004

11 Startup Checklist This is a brief summary of the steps for installation of your SpectraSYSTEM RI-150 Refractive Index Detector. Complete installation information can be found in Chapter 2. Unpack and inspect your instrument. Position the detector appropriately. Loosen the locking screws on the bottom of the detector. Ensure that you are using the appropriate power cord for your power source. Then, connect the power cord. Make rear panel connections. Connect the flowcell. Turn on the instrument. Check initial response to power-on. Verify operation with the Quick Start procedure that begins on page 3-4. This detector was installed by: (Name) (Date) 05/2004 Thermo Electron vii

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13 1 Getting Started Introduction This chapter provides general information about your Thermo Electron, Finnigan TM SpectraSYSTEM TM RI-150 Detector. Features The RI-150 is a highly sensitive refractive index detector with the following features: The refractive index operating range is 1.00 to 1.75 RIU. A broad selection of mobile phases can be used, making the detector suitable for a variety of applications. An automatic intensity adjustment compensates for deterioration of the sensitivity due to flowcell contamination or reduced light levels. Microprocessor controlled high-resolution automatic zeroing, offset adjustment for output signal, and push-button reference liquid purge functions provide easy and prompt operation. Electrical design improvements provide low noise and high operational stability. A low volume (8 µl) flowcell and modified inlet tubing significantly reduce band broadening. The solenoid valve is operated at half of its rated voltage and the tungsten lamp is operated at reduced voltage to maximize component life. A discharge channel (drain port) on the side of the instrument directs solvent out of the detector to keep electronic and optical components dry in the unlikely event of leakage within the detector. A variety of input and output terminals, which are located on the back panel of the detector, makes it easy to automate a liquid chromatography system. 05/2004 Thermo Electron 1-1

14 Front Panel [3] [4] [5] [6] [7] [8] [9] LEAK [2] PURGE A/Z CL [10] ESC MARKER ENT SpECTRASYSTEM RI-150 [1] POWER Figure 1-1. Front panel Table 1-1. Front Panel Item Description Function No. 1 POWER key Press the POWER key once to turn the unit On or Off. Tungsten lamp lights automatically when switch is in the ON position 2 LCD Display Displays the following screens: Status, Parameters, Validation Information, and Validation Execute. Displays the refractive index value in RIU units in the upper portion of the Status screen. Display range is from -999 to 999. No figures are displayed when the refractive index value goes out of range. 3 ESC key Pressing the ESC key returns you to the Status screen. 4 PURGE key Press the PURGE key to purge air out of the flowcell or to flush the reference compartment of the flowcell with fresh solvent. 1-2 Thermo Electron 05/2004

15 Table 1-1. Front Panel, continued Item Description Function No. 5 LEAK LED The LEAK LED Illuminates when a solvent leak is detected. 6 A/Z key Pressing the A/Z key automatically zeroes the detector. Zero the detector every time you change the REC. RANGE value. As the detector is autozeroing, the LED above the A / Z key is illuminated and the message AUTO ZERO appears on the lower line of the Status screen. 7 MARKER key Outputs a pulse ( 5% full scale) to the MARKER OUT terminals on the back panel. This marker is most commonly used to indicate the start of a run. 8 Arrow keys The arrows keys allow you to move from screen to screen. They also allow you to edit parameter settings. 9 CL key Press the CL (Cancel) key to cancel your entries. 10 ENT key Press the ENT (Enter) key to confirm entries. 05/2004 Thermo Electron 1-3

16 Back Panel [13] [12] [11] [14] [15] [16] [17]! WARNING DANGEROUS VOLTAGES EXPOSED WHEN COVER IS REMOVED UNPLUG POWER CORD! WARNING FOR CONTINUED PROTECTION REPLACE ONLY WITH SAME FUSE TYPE AND RATING PURGE IN ZERO IN MARKER IN READY OUT LEAK OUT ERROR OUT COM INTEG OUT FG REC OUT + _ + _ [18] [19] [20] SERIAL NO. MODEL NO. 054/27768 RI150 [21] ~LINE This Equipment complies with the requirements in Part 15 of FCC Rules for a Class A computing device. Operation of this equipment in a residential area may cause unacceptable interference to radio and TV reception requiring the operator to take whatever steps are necessary to correct the interference.! SEE MANUAL (250V) ~LINE POWER MAX. FREQUENCY T3, 15A V 150VA 50-60Hz [22] Figure 1-2. Back panel Table 1-2. Back Panel of RI-150 refractive index detector Item Function Description No. 11 MARKER IN terminal External Signal In (Event Marker) 12 ZERO IN External Signal In (Auto Zero) 13 PURGE IN External Signal In (Purge On) 14 READY OUT Signal Out Ready 15 LEAK OUT Signal Out (Solvent leak) 16 ERROR OUT Signal Out (Error). 17 INTEG OUT Integrator out 18 FG Ground for signal cable 19 REC. OUT Recorder out 20 COM RS-232 port 21 Ground terminal Refer to page Power inlet Connect to line power 1-4 Thermo Electron 05/2004

17 Side Panel OUT IN [23] [24] [25] Figure 1-3. Side panel Table 1-3. Side panel Item Description Function No. 23 Outlet Port Solvent exits the flowcell through the OUT port. 24 Inlet Port Solvent enters the flowcell through the IN port. 25 Drain Port In the case of an internal solvent leak, solvent is expelled through the drain port on the side of the detector. 05/2004 Thermo Electron 1-5

18 Bottom Panel There are two locking screws on the bottom panel of the detector. These screws lock the position of the optical bench in order to prevent damage to the instrument during shipment. During installation of the detector, you must loosen these screws. Refer to Locking Screws on page 2-3 for information on loosening these screws. Drain Port Locking Screws Figure 1-4. Bottom panel of RI-150 detector 1-6 Thermo Electron 05/2004

19 LCD Display (User Screens) STARTUP SCREENS The RI-150 refractive index detector has an LCD display on the front panel as shown in Figure 1-1. The display consists of following four screens: Status, Parameter Setting, Validation Information, and Validation Execute. After you press the Power key, the following start-up screens appear. RI 150 Ver. RA06E01 The first screen that appears displays the model name on the first line and the version number on the second line. INITIALIZING The second screen that appears displays the message INITIALIZING as the detector initializes itself µriu 35 C TEMP. UNSTABILIZED After the detector completes its initialization process, the message TEMP. UNSTABILIZED is displayed in the second line of the display until the detector reaches its default temperature setting of 35 C µriu 35 C RD When the unit reaches the default temperature setting, the TEMP. UNSTABILIZED message disappears and the RD message appears at the end of the first line. FOUR SCREENS The display consists of the following four screens: Status, Parameter Setting, Validation Information, and Validation Execute. Pressing the right and left arrows allows you to move from screen to screen as shown below. Status Screen -0.0 µriu 35 C RD PARAMETER SETTING VALIDATION INFORMATION VALIDATION EXECUTE 05/2004 Thermo Electron 1-7

20 PARAMETER SETTING SCREENS PARAMETER SETTING Table 1-4. From the Status screen, press the Right arrow key to access the Parameter Setting screen. From the Parameter Setting screen, you can set the values for the parameters listed in Table 1-4. Parameters for SpectraSYSTEM RI-150 refractive index detector Parameter Default Available Range Recorder Range 512 µriu 512, 256, 128, 64, 32, 16, 8, 4, 2, 1, 0.5, or 0.25 µriu Integrator Range 500 µriu 500 or 125 µriu Temperature 35 C 30 C to 50 C (in increments of 1 C ) or OFF Time Constant 3 seconds 6, 3, 2, 1.5, 1, 0.5, 0.25, or 0.1 seconds Polarity positive (+) positive (+) or negative ( ) Baseline Shift 0 µriu 0 to 50 (in increments of 1) To move from parameter to parameter, do the following: 1. Press the Up arrow or the Down arrow. 2. To access the available settings in a parameter screen, press the ENT key. A flashing cursor appears indicating the parameter setting is selectable. 3. Select a setting by using the Up and Down arrow keys. 4. Then, press the ENT key to confirm the selection. From the Parameter Setting screen, press the Down arrow screen to access the next screen. PARAMETER REC. RANGE 512 µriu The RECORDER RANGE screen allows you to set the range of your recording device. Press the ENT key. A flashing cursor appears indicating that the Recorder Range setting is selectable. Select 512, 256, 128, 64, 32, 16, 8, 4, 2, 1, 0.5, or 0.25 by using the Up and Down arrow keys. Then, press the ENT key to confirm the selection. Press the Down arrow key to access the next screen. PARAMETER INTEG. RANGE 500 The INTEGRATOR RANGE screen allows you to set the range of your integrator. Press the ENT key. A flashing cursor appears indicating that the Integrator Range setting is selectable. Select 500 or 125 by using the Up and Down arrow keys. Then, press the ENT key to confirm the selection. Press the Down arrow key to access the next screen. PARAMETER TEMPERATUE 35 C The TEMPERATURE screen allows you to set the operating temperature of the detector. Press the ENT key. A flashing cursor appears indicating that the Temperature setting is selectable. Select a 1-8 Thermo Electron 05/2004

21 temperature setting from 30 C to 50 C or select OFF by using the Up and Down arrow keys. Then, press the ENT key to confirm the selection. Press the Down arrow key to access the next screen. PARAMETER TIME CONSTANT 1.5SEC The TIME CONSTANT screen allows you to select the time constant of the detector s response. Press the ENT key. A flashing cursor appears indicating that the Time Constant setting is selectable. Select 6, 3, 2, 1.5, 1, 0.5, 0.25, or 0.1 seconds by using the Up and Down arrow keys. Then, press the ENT key to confirm the selection. Press the Down arrow key to access the next screen. PARAMETER POLARITY + The POLARITY screen allows you to select the polarity of your analysis. Press the ENT key. A flashing cursor appears indicating that the Polarity setting is selectable. Select + or - by using the Up and Down arrow keys. Then, press the ENT key to confirm the selection. Press the Down arrow key to access the next screen. PARAMETER BASELINE SHIFT 0 The BASELINE SHIFT screen allows you to select the offset of your baseline. Press the ENT key. A flashing cursor appears indicating that the Baseline Shift setting is selectable. Select a value from 0 to 50 by using the Up and Down arrow keys. Then, press the ENT key to confirm the selection. Press the Down arrow key to access the next screen. PARAMETER SELECT DEFAULT ERASE CURRENT SETTING? NO The SELECT DEFAULT screen allows you to reset the parameter settings to their defaults. If you want to reset all of the parameter settings to their defaults, press the ENT key. A flashing cursor appears. Select YES by using the Up and Down arrow keys. Then, press the ENT key to confirm the selection. Select NO if you do not want to erase your settings. After pressing the ENT key to confirm that you want to reset the detector to its default settings, the message Erase Current Setting? appears. Leave the setting at NO if you do not want to erase your parameter settings. Select YES if you want to erase your entries. If you select Yes, the parameter settings are returned to their defaults, the detector beeps, and LCD display goes back to the Status Screen. If you selected NO above, press the Down arrow to access the next parameter setting screen. PARAMETER LCD CONTRAST 4 The LCD CONTRAST screen allows you to adjust the contrast of the detector s LCD screen. Press the ENT key. A cursor appears to indicating that you can change the contrast of the LCD. Select a 05/2004 Thermo Electron 1-9

22 contrast between 1 and 7 by using the Up and Down arrow keys. Then, press the ENT key to confirm the selection. VALIDATION INFORMATION SCREENS VALIDATION INFORMATION The Validation Information screens allow you to access information about the performance of the RI-150 refractive index detector. To access the Validation Information screen from the Status screen, press the Right arrow key. Press the Down arrow key to access the next screen. LAST VALIDATION DATE.. The LAST VALIDATION DATE screen displays the date of the most recent validation of the RI-150 refractive index detector. This screen does not display a date until you execute your first validation and enter a date in the INPUT DATE screen. Press the Down arrow key to access the next screen. LAMP VOLTAGE 3.75 V The LAMP VOLTAGE screen displays the lamp voltage detected during the last validation of the detector. If the lamp voltage exceeds 4.5 V, purge the flowcell with fresh solvent. The applied lamp voltage increases automatically as the light intensity from the flowcell drops. The reduction in light intensity can be due to either a dirty flowcell or an aging light source. Refer to page 6-2 for instructions on cleaning the flowcell. The typical lifespan of the tungsten lamp is 40,000 hours. Call service for lamp replacement. SPAN 512µRIU Press the Down arrow key to access the next screen. The SPAN screen displays the difference in the refractive index between the reference solvent (typically deionized water) and a standard solution (typically a solution of sucrose) from the last validation of the detector. The span function is a validation of the detector s response to a defined standardized solution. VALIDATION EXECUTE SCREENS VALIDATION EXECUTE ENT The Validation Execute feature allows you to execute a validation routine to verify the accuracy of your RI-150 detector. From the VALIDATION INFORMATION screen, press the Right arrow to access the VALIDATION EXECUTE screen. To execute the Validation routine, press the ENT key once Thermo Electron 05/2004

23 INPUT DATE.. The INPUT DATE screen allows you to enter a date for the validation. Press the ENT key. A cursor appears indicating that you can edit the date. Use the Up and Down arrow keys to edit the date. Then, press the ENT key to confirm your entry. Press the Down arrow key to access the next screen. ZERO MEASUREMENT PUSH ENTER 0.0 SAMPLE MEASUREMENT PUSH ENTER The ZERO MEASUREMENT screen allows you to re-zero the detector s baseline. Ensure that the detector s temperature is stabilized and that you are pumping the appropriate mobile phase through both the reference and sample compartments of the detector s flowcell. Then, press the A/Z key to autozero the baseline. When 0.0 appears, press the ENT key. The SAMPLE MEASUREMENT screen allows you to validate the accuracy of your detector. With the purge valve closed, pump an appropriate standard solution through the system to verify the accuracy of the detector s response. Press the ENT key to confirm the value that appears. This value will be reported in the SPAN screen. Error Messages To alert you about a condition that might damage your instrument or compromise your analysis, the RI-150 refractive index detector displays the following error messages in the second line of the Status screen: Home Position Error, Leakage Error, Overheat Error, Optical Balance Error, and Parameter Error µriu 35 C HOME POSITION ERROR µriu 35 C OPT. BALANCE ERROR µriu 35 C LEAKAGE ERROR The HOME POSITION ERROR message indicates that the null glass does not come back to its home position. If this message appears when you attempt to autozero the detector from an external control device, try purging the flowcell. If this message appears during instrument startup, power off the detector and contact your local Thermo Electron representative. The OPT. BALANCE ERROR message indicates that Auto-Zero was not successful. Insufficient solvent exchange of the reference flow path might cause this. Try purging the flowcell to fill the reference compartment with fresh solvent. If purging the flowcell does not correct this problem, power off the detector and contact your local Thermo Electron representative. The LEAKAGE ERROR message indicates a solvent leak inside the chassis of the detector. CAUTION! If the LEAKAGE ERROR message appears, power off the detector at once. 05/2004 Thermo Electron 1-11

24 600.0 µriu 35 C PARAMETER ERROR The PARAMETER ERROR message indicates that the detector has failed to store data µriu 35 C OVERHEAT ERROR The OVERHEAT ERROR message indicates overheating of the detector s optical block. If this message appears, power off the detector and contact your local Thermo Electron representative. MISCELLANEOUS MESSAGES In addition to the error messages listed above, there are several other messages that might be displayed as follows: Intensity, Temp. Unstabilized, Purge, and Autozero. 0.0 µriu 35 C INTENSITY The INTENSITY message indicates inadequate light intensity. There are several possible causes for this condition: 1. The reference side of the flowcell might contain an inappropriate solvent. Set the LC pump to deliver an appropriate solvent. Then, press the PURGE key to fill the reference side of the flowcell with fresh solvent. Purge the flowcell for 5 minutes at a flow rate of 1 ml/min to refill the reference compartment with fresh solvent. Then, press the PURGE key a second time to return to normal operation. CAUTION! Leaving the purge valve in the On position for excessive periods of time will shorten its lifespan. At a flow rate of 1 ml/min, 5 minutes is typically sufficient to flush the flowcell. 2. There are air bubbles in the flowcell. Press the PURGE key on and off a few times to remove air bubbles from both the reference and sample sides of the flowcell. 3. The optical axis is off center. Press the A/Z key to autozero the detector. 0.0 µriu 35 C TEMP. UNSTABILIZED 0.0 µriu 35 C PURGE 0.0 µriu 35 C AUTOZERO The UNSTABILIZED TEMPERATURE message indicates that the temperature of the optical system has not met the preset temperature. The PURGE message indicates that the purge valve is open. When the purge valve is open, solvent flows through both the reference and the sample sides of the detector s flowcell. See Figure 4-4 on page 4-5. The AUTOZERO message indicates that the detector is zeroing its intensity readout Thermo Electron 05/2004

25 2 Installation Introduction This chapter covers the initial installation of your RI-150 detector, including connection to other chromatographic instrumentation. As you go through unpacking and installation, you may want to use the Start-up Checklist located at the beginning of this manual. The checklist is an abbreviated version of this chapter and is supplied as a quick reference of how to conduct a successful installation. Unpacking Carefully remove the detector from the shipping container and inspect both the detector and packing for any signs of damage. If you find any damage, immediately contact the shipping company. CAUTION! The RI-150 refractive index detector weighs 13 kgs (27 lbs). Use proper lifting techniques to avoid potential injuries. Standard Accessories The shipping container should contain the detector and the standard accessory kit. Carefully check to make sure you have received all the items listed in Table 2-1. If any items are missing, contact your Thermo Electron representative immediately. Place the detector on a vibration-free bench top as close as possible to the chromatographic column outlet (thus minimizing the length of tubing necessary for connection to the flowcell inlet). Refractive index detectors are very sensitive to changes in temperature and airflow, which can cause a drifting baseline. Therefore, be sure to place the detector in a draft-free location away from an open window, air conditioner vents, or other circulating air source. Allow at least 13 cm (5 inches) of clear space between the rear panel of the detector and any wall or obstructions to provide access to the rear-panel connectors. 05/2004 Thermo Electron 2-1

26 Table 2-1. Standard accessories Item Item No. 1 Power cord for 120 V line power source and power cord appropriate for shipping destination, if different Quantity 1 or 2 2 Fuse, 3.15 A 2 3 Signal cable, double core, shielded, 2 meter length 4 Inlet tubing, stainless steel, preflanged, with nut and ferrule 5 Outlet tubing, stainless steel, with nut, ferrule and Teflon tubing 6 5 mm Allen wrench 1 7 Reference manual Thermo Electron 05/2004

27 Installing the Detector POWER REQUIREMENTS LOCKING SCREWS The RI-150 refractive index detector requires a 50 or 60 Hz single-phase power source capable of providing 100 to 240 Vac. CAUTION! The RI-150 refractive index detector operates with singlephase power only. If your facility provides only three-phase power, consult your local Thermo Electron representative. To prevent damage that might occur during shipment, the optical block of the detector is fixed by two locking screws (5 mm Allen bolts). After you place the detector on your laboratory bench top, loosen these screws as follows: 1. Move the RI-150 detector to the edge of the laboratory bench top until the locking screws on the bottom of the detector are accessible. Do not place the detector on its side to access the locking screws. Be careful to support the detector during this procedure to prevent it from falling. 2. Use a 5 mm hexagonal Allen wrench to loosen the screws. Do not remove the screws. 3. After you have loosened the locking screws, move the detector back to a secure position on the bench top. NOTE: The detector will not stabilize if the locking screws are not loosened. Be careful not to tip the detector off the bench top as you loosen the locking screws. 5 mm Allen wrench Figure 2-1. View of the RI-150 detector overhanging the edge of the bench top 05/2004 Thermo Electron 2-3

28 ELECTRICAL CONNECTIONS Connect the power cord to the power receptacle on the rear panel of the detector. CAUTION! Ensure that you are using an appropriate power cord for your power source. The RI-150 refractive index detector is shipped with a 3-prong parallel blade plug that is appropriate for a 120 V, 50 to 60 Hz line power source and a power cord that is appropriate for the shipping destination. CAUTION! Ground the instrument properly to protect the operator from electrical shock. Proper grounding also protects the instrument from power line noise. Use the ground terminals that are located on the lower right side of the rear panel to ground your instrument if you are not connecting it to a power outlet with a protective earth contact (ground). See Figure 2-2.! SEE MANUAL (250V) ~LINE POWER MAX. FREQUENCY T3, 15A V 150VA 50-60Hz Figure 2-2. Grounding terminals Make the appropriate signal cable connections using the accessory signal cable. Be sure to connect the shielded terminal of the cable to the FG terminal. See the integrator connections shown in Figure 2-3 and the recorder connections shown in Figure 2-4. Red Lead INTEG OUT FG REC OUT + _ + _ White Lead Grounding Lead Figure 2-3. Integrator connections 2-4 Thermo Electron 05/2004

29 INTEG OUT FG REC OUT + _ + _ Grounding Lead White Lead Red Lead Figure 2-4. Recorder connections Use a duplex wire to connect the following contact closure terminals to external devices as appropriate: Input signal terminals: Purge In, Zero In, and Marker Output signal terminals: Ready Out, Leak Out, Error Out To connect the contact closure terminals to external devices, do the following: 1. Strip about 7 to 8 mm of insulating sleeve off the end of a duplex wire. 2. As you depress the terminal button by using a small flat-head screwdriver, insert the end of the wire into the terminal. Then, release the terminal button to lock the wire into the terminal. See Figure 2-5. PURGE IN READY OUT ZERO IN LEAK OUT MARKER IN ERROR OUT Figure 2-5. Terminal block connections 05/2004 Thermo Electron 2-5

30 PLUMBING CONNECTIONS Make the following connections after ensuring that the liquid chromatograph and the column are thoroughly purged with the solvent to be used and that the solvent is thoroughly filtered and degassed. 1. Remove the plugs from the IN and OUT ports on the side panel. Do not discard the plugs. 2. Connect the accessory stainless steel inlet tubing (Table 2.1, item 4) to the IN port. 3. Connect the accessory stainless steel/teflon outlet tubing (Table 2.1, item 5) to the OUT port. CAUTION! The RI-150 refractive index detector must be the last component in your LC system. Do not connect another detector, a backpressure regulator, or narrow-bore tubing to the OUT port. 2-6 Thermo Electron 05/2004

31 3 Operation Introduction This chapter provides step-by-step instructions for general detector operation as well as suggestions for ensuring the accuracy and reliability of the detector. Before You Begin Before you begin this chapter, your detector should be installed in a chromatographic system (see Chapter 2), and you should have completed the Startup Checklist located at the front of this manual. We also recommend that you review Chapter 1: Getting Started to familiarize yourself with the layout of the detector controls. Operating Instructions (General) 1. Turn on the detector by pressing the POWER key on its front panel. 2. Set the Temperature parameter to an appropriate setting and then allow the temperature to stabilize for at least 15 minutes. For maximum sensitivity, allow the temperature to stablize for 24 hours. Refer to page 1-8 for instructions on setting the detector s temperature. A 76 C thermal fuse prevents the unit from overheating by shutting down power to the heating element if necessary. NOTE: To avoid long warm-up periods, you can leave the detector in continuous operation. The typical lifespan of the tungsten lamp is 4.5 years during continuous operation. NOTE: If the sample or eluent cannot be heated to 35 C, set the temperature parameter to OFF and use the detector in a room with minimum temperature fluctuations. 2. Pump eluent at a flow rate of 1 ml/min through the detector for about 10 minutes. 05/2004 Thermo Electron 3-1

32 3. Press the PURGE key to the ON position to thoroughly flush both the sample and reference compartments of the flowcell. Leave the PURGE key in the On position and pump mobile phase through both compartments of the flowcell for approximately 5 minutes. As you purge the detector, verify that no eluent leaks from the flowcell. If you suspect that the flowcell contains adhering gas bubbles, remove the bubbles by pressing the PURGE key On and Off every 10 seconds until all the bubbles are removed. 4. Press the PURGE key Off. 5. When a recorder is used, set the desired RANGE and turn on the recorder. 6. Wait until the baseline has stabilized. Then, press the A/Z key to adjust the zero position. 7. Use the Baseline Shift parameter setting to shift the baseline as required. 8. Perform a validation test (refer to page 3-3), if required. Then, start your analysis. Suggestions Observe the following precautions. The flowcell has a pressure rating of 75 psi (5.2 bar). Do not apply backpressure to the outlet of the detector. When the detector is used in series with other detectors, place it at the end of the series. When using mobile phases containing salts, be sure to thoroughly wash the flowcell with water after use to prevent precipitation, which will damage the detector. Some salts will precipitate if exposed to high concentrations of organic solvents. When a high concentration of sample is analyzed, pump a sufficient amount of the eluent through the detector to remove the entire sample. Always use a thoroughly degassed eluent. Place the waste liquid receptacle a little higher than the detector so as not to cause negative pressure to develop at the eluent outlet. Be aware of potential miscibility problems when changing solvents in the detector. If the solvents are insoluble, flush the detector completely with an intermediate solvent such as acetone or ethanol. If a leak occurs, turn off the detector immediately and contact your local Thermo Electron representative. Do not use any chemical substances that can corrode stainless steel, such as hydrochloric acid. 3-2 Thermo Electron 05/2004

33 Validation Test Thermo Electron recommends that you validate the accuracy of the detector on a regulary scheduled basis. Validation data is stored in the VALIDATION INFORMATION screens. SUCROSE SOLUTION Prepare a fresh solution of sucrose as follows: 1. Weigh out 350 mg of sucrose and transfer quantitatively to a 100 ml volumetric flask. 2. Add approximately 50 ml of deionized, filtered, degassed water to the flask. Swirl the flask to dissolve the sucrose. Then, fill the flask to volume with water. SPAN VALIDATION EXECUTE INPUT DATE ZERO MEASUREMENT PUSH ENTER 0.0 SAMPLE MEASUREMENT PUSH ENTER 0.0 Determine the span of the detector as follows: 1. With the PURGE key ON, equilibrate the detector by pumping deionized water through both the reference and sample compartments of the flowcell at a flow rate of 1 ml/min. 2. Ensure that the baseline is stabilized. The baseline drift should be less than or equal to 500 nriu/hour. 3. Go to the VALIDATION EXECUTE screen. Then, press the ENT key to access the INPUT DATE screen. 4. Enter a date for the validation. Then, press the ENT key to access the ZERO MEASUREMENT screen. 5. Press the A/Z key to autozero the refractive index readout. Then, press the ENT key to access the SAMPLE MEASUREMENT screen. 6. Disconnect the tubing from the IN port of the detector. Then, attach an adapter to the IN port of the detector. 7. Press the PURGE key to turn the purge valve off. This shuts off the flow of solvent through the reference compartment of the flowcell. Ensure that the PURGE LED is not lit. 8. Fill a syringe with the freshly prepared sucrose solution. Then, gently inject approximately 2mL of the solution into the IN port of the detector. CAUTION! The flowcell cannot withstand a backpressure greater than 700 kpa (100 psi). Therefore, you can easily break the flowcell by forcing fluid through it. 05/2004 Thermo Electron 3-3

34 SAMPLE MEASUREMENT PUSH ENTER SPAN LAMP VOLTAGE Check the refractive index reading in the SAMPLE MEASUREMENT screen. The value should be within 487 to 537 µriu (512 µriu ±5%). The full scale reading for the detector is 1024 µriu (1.0 V). 10. Press the ENT key once to approve the result. The confirmation screen appears, showing the span and lamp intensity readings for the detector. Press the ENT key a second time to store the sample measurement and return to the VALIDATION EXECUTE screen. 11. After you complete the validation accuracy test, reattach the tubing to the IN port of the detector. 12. Set up the LC system to pump deionized water at a flow rate of 1 ml/min. Then, press the PURGE key to flush the sucrose solution out of the detector s flowcell. After a period of approximately 5 to 10 minutes, press the PURGE key a second time to return to normal operation. QUICK START LC SYSTEM SETUP AND PREPARATION CONFIGURING THE LC SYSTEM 1. Prepare a sample solution of sucrose (2.5 ppm) dissolved in 100% water. This example uses a 100% water mobile phase at a flow rate of 1 ml/min and a 10 cm C8 column. You can substitute other samples, solvents, and columns as needed for specific applications, but you will have to make appropriate changes in the method. 2. Configure your LC system as you normally would prior to analysis. Use the chromatography conditions listed below for this analysis. Table 3-1. Chromatographic conditions Parameter Mobile phase Flow rate Run time Column Injection type Injection volume Flush solvent Setting 100% water 1 ml/min 10 minutes Octyl B, C-8, cm, 5 µm particle size PushLoop 20 µl (50 ng of sucrose) 100% water 3-4 Thermo Electron 05/2004

35 PURGING THE DETECTOR S FLOWCELL DETECTOR FUNCTIONAL TESTS 3. When using any refractive index detector, the elimination of air bubbles, which can be accomplished by flushing the flowcell (sample and reference compartments), is critical for optimal performance. To remove air bubbles, do the following: a) Press the POWER key to turn on the detector. b) To begin flushing the flowcell, press the PURGE key. The green LED above the PURGE key should light indicating that the purge function is activated. c) Pump 100% water at a flow rate of 1 ml/min through the flowcell for 10 minutes. Purging is complete when the purge valve can be switched from ON to OFF with minimal baseline disturbance. 4. Before attempting to use the RI-150 detector, do the following: a) Before the initial use of the detector, ensure that the locking screw on the bottom of the detector has been loosened. Refer to the installation procedure on page 2-3. b) Perform an accuracy validation test. Refer to the validation procedure on page 3-3. c) Ensure that the temperature of the detector has stabilized. The TEMP UNSTABILIZED message is displayed on the second line of the Status screen until the temperature of the detector stabilizes. d) Ensure that no error messages are displayed in the Status screen. Refer to pages 1-11 to 1-12 for a list of possible error messages. e) Press the A/Z key and verify that the refractive index display in the Status screen reads zero. For assistance, refer to Chapter 5: Troubleshooting if any of the above checks fail. 05/2004 Thermo Electron 3-5

36 SAMPLE ANALYSIS 5. From the TIME CONSTANT screen, set the detector s response to 1.5 seconds. 6. Optimize the offset and the range settings if you are using a chart recorder. Refer to pages 1-7 to 1-9 for instructions on entering your analysis parameters. 7. Inject 20 µl of the sample. If you are using a chart recorder, you might want to press the MARKER key to indicate the start of the injection. The sucrose peak should elute at approximately 3 minutes. If the peak is negative, switch the polarity. The peak will be negative if the refractive index of the sample is less than the refractive index of the solvent used to flush the reference side of the flowcell. EXPECTED RESULTS Table 3-2 shows the typical retention times and peak areas for a 2.5 ppm solution of sucrose in water analyzed on the RI-150. Table 3-2. Sucrose analysis data Compound Injection Amount (ng) Retention Time (min) RI-150 Display (µriu) Sucrose Note: The RI-150 display is RIU. The actual reading for this example is RIU. 3-6 Thermo Electron 05/2004

37 SOLVENT SELECTION NOTE: Chromatographic solvents should be thoroughly degassed before and during use. This is especially important for obtaining the best performance with the refractive index detector. Detector response can be enhanced or degraded by the choice and preparation of the mobile phase. In addition to providing proper separation, the solvent must also be compatible with the detector and provide a sufficient refractive index difference so that the sample can be measured. Any refractive index detector is sensitive to small changes in solvent composition, and the majority of problems associated with refractive index detectors can be traced to this cause. Changes in composition can occur from incompletely mixed binary or ternary solvents, column bleed, leaching of prior samples or solvents, decomposition of the solvent, or by changing the amount of dissolved gas in the solvent. A specific example of the refractive index detector s sensitivity to small changes in solvent composition is the baseline drift caused by absorption of air in a previously degassed solvent. If a degassed solvent is left exposed to air, air will absorb in the top layer of solvent and will gradually disperse through the solvent by natural convection and molecular diffusion. As the solvent reservoir is drawn down, the withdrawn solvent will increase in amount of dissolved air, and the refractive index detector will drift towards lower refractive index. It is better to use continuously degassed solvents or maintain a helium blanket over a vacuum degassed solvent so that this drift effect is eliminated. Vacuum membrane degassing (using the Thermo Electron, Finnigan Vacuum Membrane Degasser) and helium sparging are two acceptable methods of continuously degassing the solvent. Binary and ternary solvents must be thoroughly mixed and remain mixed to prevent baseline offsets. This detector is typically sensitive to changes in composition in the low parts per million range. Pre-mixed solvents in a stirred reservoir provide the most stable operation. Stream mixing of solvents from separate reservoirs can result in baseline upsets. Solvent delivery systems used in HPLC ordinarily have compositional mixing precision of about 1% and not parts per million. Solvents mixed and delivered by the pump will show compositional changes detectable by the refractive index detector even after passage through the chromatographic system. This effect can be minimized but not eliminated by adding additional mixing before the injector. 05/2004 Thermo Electron 3-7

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39 4 Theory of Operation Introduction This brief theory of operation describes the functions of the components that make up your RI-150 refractive index detector. For more information, contact your Thermo Electron representative. Optical System A beam of light is directed through a flowcell with separate compartments containing a reference solvent and the sample to be analyzed. As the light passes through the reference and sample compartments of the flowcell, it is deflected. This deflection is proportional to the difference in the refractive indices of the reference and sample solvents. The deflection is detected by a photodiode and the result is displayed as the RI. This process is shown in Figure 4-1. The numbered components shown in Figure 4-1 are described in Table /2004 Thermo Electron 4-1

40 1 3 2 n s n r Figure 4-1. Deflection type refractive index detector Table 4-1. Deflection type refractive index detector Item No. 1 Light Description 2 Sample compartment 3 Reference compartment 4 Light beam at the time of n s > n r 5 Light beam at the time of n s = n r 6 Displacement 7 Photodiode n s n r Refractive index of liquid in sample compartment Refractive index of liquid in reference compartment 4-2 Thermo Electron 05/2004

41 The optical system is shown in Figure 4-2. The numbered components shown in Figure 4-2 are described in Table Figure 4-2. Optical system Table 4-2. Optical system Item Description No. 1 Tungsten lamp 2 Condensing lens 3 Slit No. 1 4 Collimator lens 5 Slit No. 2 6 Flow cell 7 Mirror 8 Null glass 9 Photodiode The condensing lens (item 2) and slit no.1 (item 3) form an image of the tungsten filament (item 1). This image passes through the collimator lens (item 4) and slit no.2 (item 5), which project the image of slit no. 2 through the flowcell (item 6) and onto the photodiode (item 9). When there is a difference in the refractive indices of the sample and reference solvents in the flowcell, the image on the photodiode moves horizontally as illustrated in Figure 4-3. The components shown in Figure 4-3 are described in Table /2004 Thermo Electron 4-3

42 a) No difference in the refractive index b) Some difference in the refractive index Figure 4-3. Movement of the image on the photodiode Table 4-3. Movement of the image on the photodiode Item Description No. 1 Diode A 2 Diode B 3 Image The electrical signals emitted respectively from the diodes A and B change in proportion to the movement of the image. A signal corresponding to the difference in refractive index can be obtained from the difference between the two signal outputs. Purging the Flowcell The flowcell is designed to enable replacement of a reference liquid by pushing the Purge key. Figure 4-4 shows a block diagram of the flowcell. The components shown in Figure 4-4 are described in Table 4-4. When the PURGE key is in the ON position, NC is open and NO is closed, allowing flow out of the sample compartment to the waste liquid receptacle by way of the reference compartment. When the PURGE key is in the OFF position, NC is closed and NO is open, allowing flow from the sample compartment directly to the waste liquid receptacle without going through the reference compartment. 4-4 Thermo Electron 05/2004