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					     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                           Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892

                                                        18
                                 List of SOPs for [ f]Fallypride for Injection


                                            Name of SOP                                      SOP #
      1             Preparation of Stock Solution of Kryptofix 2.2.2* and            SOP#GP101
                    Potassium Carbonate
       2            Cleaning Procedure for Radiochemistry Glassware, Reactor         SOP # GP102
                    Vials and Magnetic Bars
       3            Fallypride HPLC Mobile Phase Preparation                         SOP # GP103
       4            Micropore Filter Testing and Drug Product /Filter                SOP # GP104
                    Compatibility (Bubble Point Test).
       5            Preparation of Stock Acetate Buffer                              SOP # GP105
       6            BOXCLEAN Program (GE Tracerlab FXFN)                             SOP # MP201
       7            GE Tracerlab FXFN Module. Vacuum Pressure Check                  SOP # MP202
                                      18
       8            Production of [ F]Fallypride for Injection; Part 1: Pre-         SOP # MP203
                    Synthesis Procedures
                                      18
       9            Production of [ F]Fallypride for Injection; Part 2: Synthesis    SOP # MP204
                    and Purification
                                      18
      10            Production of [ F]Fallypride for Injection; Part 3:              SOP # MP205
                    Formulation
      11            Standard Curve of Reference Fallypride                           SOP # QA301
      12            Analysis of Organic Residues by Gas Chromatography               SOP # QA304
                       18
                    in [ F]Fallypride for Injection
      13            Analytical HPLC QC-Method                                        SOP # QA305
      14            Sterility Test                                                   SOP # QA306
      15        P Pyrogen Test (LAL Bacterial Endotoxin Test).                       SOP # QA307
                Si Simplified Procedure
      16            Analysis of The Purity of Fallypride and Tosyl-Fallypride by     SOP # QA308
               HP HPLC




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                       Page 1 of 34
       [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                          PROCEDURES
PET Radiopharmaceutical Sciences Section,                                              Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


                                                    SOP # GP101
            Preparation of Stock Solution of Kryptofix 2.2.2* and Potassium Carbonate


Approved by: _____________________________Initials________ Date:___________
                 Victor W Pike, Ph.D., Chief, PET Radiopharmaceutical Sciences, NIMH

           Purpose: to prepare a stock solution of Kryptofix 2.2.2* and potassium carbonate

1. Procedure:
       1. Using an analytical balance and a plastic screw-cap vial (20 mL size), weight out 25 mg of
           potassium carbonate (Aldrich, anhydrous, 99.99%). Add deionized water (0.5 mL) and mix
           to dissolve.
       2. Using weighing paper, weigh 250 mg of Kryptofix 2.2.2 (anhydrous; 98%; Aldrich) and
           transfer to the same vial. Add dry acetonitrile (4.5 mL) and mix to dissolve.
       3. Place appropriate label showing preparation and expiry date (to be determined) and keep
           the solution in the refrigerator until use (do not store in glass vial).
       4. Record the data of Kryptofix 2.2.2-potassium carbonate complex in Table 1.

2. Record: Lot #__________________________
                                        18
           Table 1. Reagents : [ F]fluoride-Kryptofix 2.2.2-potassium carbonate complex
Item           Reagent              Specifications            Manufacturer     Lot #    Expiry date Quantity
  1        Kryptofix 2.2.2        Anhydrous                Aldrich Chem.
           (250 mg)               98%                      Co.


  2        Potassium              Anhydrous                Aldrich Chem.
           carbonate              99.99%                   Co.
           (25 mg)
  3        Acetonitrile           Anhydrous 99.8%          Aldrich Chem.
           (4.5 mL)                                        Co.
   4       Water                  De-ionized               M; Millipore, In
           (0.5 mL)                                        house
Note: 100 µL of solution contains 0.5 mg potassium carbonate and 5 mg Kryptofix 2.2.2


Chemist_____________________________Initials_________________Date________________



Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                          Page 2 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                      Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


                                                    SOP # GP102
     Cleaning Procedure for Radiochemistry Glassware, Reactor Vials and Magnetic Bars


Approved by: _____________________________Initials________ Date:_____________
                 Victor W Pike, Ph.D., Chief, PET Radiopharmaceutical Sciences, NIMH,


         Purpose: to clean glassware, reactor, vials and magnetic bars that are used in the
                        18
         production of [ F]Fallypride for Injection.



     Procedure:
     1. Rinse with acetone (HPLC grade) to remove organic residues.
     2. Rinse with hexane (HPLC grade) to remove residual silicon oil.
     3. Bathe the glassware in an aqueous 2% solution of Liqui-Nox (Valconox, made from dilution
          of a 10% stock solution).
          a) Bring solution to vigorous boiling for 5–10 min.
          b) Swab material being washed with wood, cotton-tipped applicators (6 in).
     4. Rinse glassware 3 times with Millipore water.
          a)   Carefully spot-check each item.
          b) Give final rinses with de-ionized water bottle (3 times).
          c)   Leave the clean material to drip, over paper towel.
     5. Place small V-vials, magnetic bars and reaction vials in beakers (Pyrex) and cover with
          aluminum foil. Place material in the oven at 170−180 ºC and allow to dry overnight.
     6. Keep all the material in the oven until use.


Notes:
          a) Except for the aqueous Valconox that can be disposed in the sink, all other washing
solvents must be placed in their respective waste container.
          b) In the Oven Log enter date and temperature IN and date and temperature OUT.


Chemist_____________________________Initials_________________Date________________




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                  Page 3 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                        Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


                                                    SOP # GP103
                                 Fallypride HPLC Mobile Phase Preparation


Approved by: _____________________________Initials________ Date:_____________
                 Victor W Pike, Ph.D., Chief, PET Radiopharmaceutical Sciences, NIMH,

                                                                           18
       Purpose: to prepare mobile phase for the HPLC purification of [ F]fallypride and QC
                      18
       analysis of [ F]Fallypride for Injection.



Procedure:
     1. Preparation of HPLC solvent for the preparative column
               a) Fill up a flask (1 L size) for HPLC solvent with acetonitrile (HPLC grade) containing
                 0.6% TEA (6 mL of TEA /L).
               b) In a similar flask, prepare 1 L of HPLC water only.


     2. Preparation of HPLC solvent for analytical column.
               a) To prepare stock ammonium formate solution (100 mM), weigh out ammonium
               formate (6.013 g) on weighing paper and then dissolve in HPLC water (1 L), mix well
               and store at room temperature.
               b) Put stock ammonium formate solution (50 mL; 100 mM) into a volumetric cylinder (1
               L size), then dilute with HPLC water (450 mL; HPLC Grade)
               c) Put acetonitrile (HPLC grade; 500 mL) into another volumetric cylinder. Mix
               solutions from (a) and (b).


Chemist_____________________________Initials_________________Date________________




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                    Page 4 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                           Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


                                                    SOP # GP104
                   Micropore Filter Testing and Drug Product /Filter Compatibility
                                                (Bubble Point Test).


Approved by: _____________________________Initials________ Date:____________
                 Victor W Pike, Ph.D., Chief, PET Radiopharmaceutical Sciences, NIMH


                              Purpose: Testing of Micropore filter compatibility.


         Filter-Integrity test. The integrity of the Millex-MP sterile filter that was used for sterile
                         18
          filtration of [ F]Fallypride for Injection is tested by the following method. The knob on the
          pressure regulator that is hooked to the house compressed air gas supply is turned counter
          clockwise so as to minimize outlet pressure. The filter assembly is removed from the
          sterile dose vial and placed on the male luer-lock fitting of the pressure-regulated 1/8”
          Teflon line. A disposable needle (1.5 inch 22 gauge) is placed on the male luer fitting of
          the Millex-MP filter. A portion of this needle is submerged in a test tube (12 x 50 mm)
          containing about 3 mL of HPLC grade water. The knob of the pressure regulator is slowly
          turned clockwise and the pressure gauge is monitored visually. The pressure is brought to
          45 p.s.i. If there is an absence of a steady stream of bubbles in the test tube when the
          pressure gauge reads 45 p.s.i., then the filter passes the test. The result of the filter test is
          recorded on the batch record and in the summary section of the quality control test form.




Chemist_____________________________Initials_________________Date________________




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                       Page 5 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                         Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892




                                                    SOP # GP105
                                        Preparation of Stock Acetate Buffer


Approved by: _____________________________Initials________ Date:______________
                  Victor W Pike, Ph.D., Chief, PET Radiopharmaceutical Sciences. NIMH


                                                                                   18
          Purpose: to prepare stock acetate buffer (250 mM; pH 5.5) for use in [ F]Fallypride
          for Injection prouction



A. Reagents:
             Name of reagent             Manufacturer                            Quantity
             Sodium acetate              Aldrich Chem. Co.
                                         Cat. # 241245, ACS reagent
             Acetic acid                 Aldrich Chem. Co.
                                         Cat. # A6283, 500 mL
             Water                       EMD Chemicals Inc.
                                         Cat. # WX0004-1


B. Procedure:
     1.     On weighing paper, weigh out sodium acetate (14.45 g) and then dissolve this in HPLC
            water (500 mL) in a bottle (500 mL size).
     2.     Take acetic acid (2.15 mL) and put it into above solution.
     3.     Mix well.
     4.     Measured the pH of the buffer solution; the pH of this buffer should be 5.3−5.7.


Chemist_____________________________Initials_________________Date________________




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                       Page 6 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                         Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892




                                                  SOP # MP201
                              BOXCLEAN program (GE TRACERlab FXFN)


Approved by: _____________________________Initials________ Date: _____________
                 Victor W Pike, Ph.D., Chief, NIMH, PET Radiopharmaceutical Sciences

       Purpose: to clean the GE TracerLab FXFN module, check its functions and prepare it
       for production.



1. Procedure:
     1. If present, remove used Sep-Pak and re-connect line to its holder-connector.
     2. Disconnect the following three lines placing the output into the waste reservoir:
               a) The one coming from the Sep-Pak holder-connector into waste.
               b) The one coming from the Sep-Pak holder-connector, going into the collection V-
                    vial (10 mL size)
               c) The one going from the HPLC loop waste into a V-vial (5 mL size) connected to
                    an external empty syringe (20 mL size), to pull diluted reaction mixture into
                    injector loop.
               d) A fourth line, from the injector to a Blue Max (50 mL size) or to a vial (20 mL
                    size) (waste) remains the same.
     3. Check that nitrogen and air compressed air valves are open. Check nitrogen cylinder is
          open. Be sure there is a reactor vessel connected.
     4. Place dry-ice in the vacuum trap.
     5. Start the TracerLab automated software program.
          Select Synthesis: "Boxclean", ENTER.
          Follow instructions.
          Add acetone as follows:
               a) 1−2 mL acetone to target vial
               b) 1–2 mL acetone to support vials V1 to V9 (fill small ones and half fill large ones)
               c) 25 mL acetone to collection bulb vessel
               d) 0.5 mL acetone to V-vial (10 mL size) (leave the spinal needle way up)




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                     Page 7 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                      Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


     6. In Preparation CHECK LIST, check each item in the list, going down with repeated
          ENTER. Finally, enter operator initials.
     7. The screen will show the list of timed events for BOXCLEAN and the process will start
                                                                           o
          with the vacuum pup ON and the temperature increasing to 100 C.
                                                            o
          Check that the temperature reaches 100 C and the pressure in reactor goes down.
     8. At the end of the Program, the system will be flushed with nitrogen, go under vacuum to
          remove most of the acetone vapors and then stop.
     9. Remove the acetone that may be trapped in the vacuum trap. Dispose of it in the
          appropriate container.


Copy of the BOXCLEAN Program see Attached file: Document 9: Validation Runs, attached
BOX Clean Program on the batch: FAY-032404.


Chemist_____________________________Initials_________________Date________________




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                  Page 8 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                          Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


                                                    SOP # MP202
                       GE TRACERlab FXFN Module. Vacuum Pressure check


Approved by: _____________________________Initials________ Date: _____________
                 Victor W Pike, Ph.D., Chief, NIMH, PET Radiopharmaceutical Sciences

            Purpose: to check the vacuum efficiency and the leak of GE TRACERlab FXFN
            module to ensure the success of production synthesis.


Procedure:
     1. Open nitrogen and compressed air valves located outside hot-cell
     2. Start Dell Computer-Windows 98. Enter Name and Password. After Virus Scan
          Computer will open. Click twice on TRACERLab icon. Enter "Admin" and "Tracerlab"
     3. Once on the Instrument SCHEME, look at the square area in the right side of the screen
          that shows pressure of compressed air and nitrogen.
     4. Turn on the vacuum on by click “ON” of the "Power”, which represents the vacuum on
          the Instrument SCHEME. (Note: this can be turned OFF in the same way).
     5. Compressed air pressure should be around _____kPa and nitrogen around______ kPa.
     6. Click the mouse in V24 and V25 should open with space bar (1 = open, 0 = close), this
          will set full vacuum on reactor vessel, reaching around ≤ 3 kPa. Record the lowest
          pressure. _______kPa
     7. Close V24 (space bar), pressure should hold (change should be ≤ 1 kPa/min).
     8. If pressure is not holding find and correct the source of leak; e.g. tighten stoppers in
          support vials, check valves 2 to 6, 13, 14, 20, 24; tight reactor vial, check O-ring.
     9. With mouse, go to V20 (Nitrogen), open V20 with space bar (can also close with space
          bar). Pressure should go up. Close with space bar and open V24. Check the pressure
          again. It should go below 2 kPa again.
     10. If pressure holds, go ahead with the next step.
Note. When both vacuum and pressure are open during the procedure, pressure should hold
between 21.5 to 22 kPa (V20 and V24 open).


Chemist_____________________________Initials_________________Date________________




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                      Page 9 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                         Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


                                                  SOP # MP203
                               18
           Production of [ F]Fallypride for Injection; Part 1: Pre-synthesis Procedures



Approved by: _____________________________Initials________ Date: _____________
              Victor W Pike, Ph.D., Chief, NIMH, PET Radiopharmaceutical Sciences


                                                            18
         Purpose: to prepare for the synthesis of [ F]Fallypride for injection on the GE
         TRACERlab FXFN module in hot-cell 5.


Procedures:
     1. Check that nitrogen and compressed air valves are open.
     2. Check nitrogen cylinder is open.
     3. Turn on the computer with TRACERlab FXFN software and with Beckman HPLC
          software.
     4. Open the TRACERlab FXFN software and check the pressure of compressed air and
          nitrogen pressure. The pressure of compressed air should be 400 p.s.i and nitrogen
          pressure > 100 p.s.i.
     5. Clean GE TRACERlab FXFN Module and hot-cell 5.
               4.1    Follow the program "CLEAN BOX" (SOP # MP201). Clean and dry the GE
               TRACERlab FXFN module on the day of preparation or the day before the preparation.
               All the reagent vessels and transfer lines of the module to be used in the preparation
               should be washed with acetone and dried under nitrogen flush before using.

               4.2 Inspect hot-cell compounding/dispensing area for cleanliness. Remove extraneous
               materials and labels. Spray ethanol to clean this area if necessary.

               5.3 Inspect and set if necessary charcoal trap in the module
                      18
     6. Prepare [ F]fluoride ion-Kryptofix 2.2.2-potassium carbonate complex solution
               5.1 Add stock Kryptofix 2.2.2-potassium carbonate (70−100 µL) into V-Vial (1 mL
                     size).
                                                                                         18
               5.2 Take V-Vial 1 to Cyclotron Facility (CC, NIH) to collect aqueous [ F]fluoride ion;
                                      18
                     the activity of [ F]fluoride ion should be between 150−250 mCi water ( ≤ 500 µL).




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                      Page 10 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                              Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


     6. Verify integrity of glassy reaction vessel assembly (containing small magnetic bar stirrer) in
          TRACERlab FXFN Module. Follow SOP # MP202 and record the data in the sheet of
                                      18
          Summary Records of [ F]Fallypride for Injection run:
               6.1 Vacuum pressure check (gauge, kPa).
               6.2 The compress air pressure (kPa)
               6.3 Nitrogen pressure (kPa)
     7.        Prepare the preparative HPLC system
               7.1 Start the HPLC computer software, enter the necessary information and select the
                    right method “ Fally_Production”
               7.2 Verify that mobile phase bottles in the right position on the Beckman System Gold
                    Delivery module.
                              7.2.1        The solvent bottle of 0.6% TEA connects to B1 line (marked red)
                              7.2.2        The solvent bottle of HPLC water connects to A1 line (marked red)
                                           Above connection should match the setup in the method
                                           “Fally_Production”.
                              7.2.3        Set Acetonitrile w 0.6%TEA /HPLC water (30: 70 v/v) as initial
                                           conditions. Equilibrate the Luna column (Phenomenex) with
                                           fallypride preparative mobile phase (~ 200 mL)
               7.3 Note the pump pressure while increasing the flow rate to 2 mL/min.
               7.4 Pump the HPLC mobile phase through the preparative column at a low flow (1−2
                    mL/min) and maintain this flow until purification time.
     8. Prepare in the clean bench a sterile filtration/collection Unit, using aseptic technique:
               8.1. Prepare the final sterile product vial. The final sterile empty vial, product needle,
                    vent needle and two filters (Millex 0.22 µm) are assembled in a certified laminar
                                                                           18
                    flow sterile cabinet. Attach a prepared label for [ F]fallypride with the current batch
                    number and date to the product vial.
               8.2. Move a prepared product vial unit from the laminar flow sterile cabinet and attach it
                    to the product line of TracerLab FN Module.
     9. Open the Tracerlab Fx Computer Controlling System ("Admin" and "Tracerlab") and Select
          Synthesis method: "Fallypride". Click “Synthesis-Start” to start preparation of the
          synthesis.
     10. Enter all the required information (including date, radiotracer, batch #, human preparation
          #) into the corresponding method of the HPLC computer software program.


Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                         Page 11 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                                Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


     11. Check that you have the correct UV absorbance (254 nm) and the correct settings for the
          gamma 12. Place the solvents and precursor solution in the corresponding module support
          vials:
           ________Support vial 1:                acetonitrile (0.5 mL).
                                                                                           18
           ________Support vial 2                  [Used as a port (V-vial; 1 mL size)]: [ F]fluoride-K 2.2.2-
                                                  potassium carbonate solution; 100 µL).
           ________Support vial 4:                dry acetonitrile (1 mL) plus tosyl fallypride (2.0 ± 0.5 mg).
           ________Support vial 5:                dry acetonitrile (0.5 mL)
           ________Support vial 6:                dry acetonitrile (1.5 mL)
           ________Support vial 7:                sterile saline solution (9–14 mL)
           ________Support vial 8:                Ethyl alcohol for Injection (1 mL)
           ________Support vial 9                 sterile water (8 mL)
           _______ Large (10 mL size)             V-vial: 0.5 mL sterile water with 0.6%TEA.
            _______ Large bulb vessel:            HPLC water (90 mL) and sodium acetate (250 mM; pH
                                                  5.5; 10 mL; see SOP # GP104).
     12. Check that „complex solution‟ delivery line goes through valve V2 and into reactor vessel
          (glassy carbon).
     13. Place acetonitrile (0.5 mL) in a syringe (3 mL) connected to an external line going into the
                                                   18
          "complex" vial for recovery of [ F]fluoride-K 2.2.2-potassium carbonate solution left in
          transportation vial and its addition to the reaction vessel.
     14. Activate one C-18 plus Sep-Pak with ethanol (10 mL) sterile water (10 mL) and place in its
          corresponding connection site in the module.
     15. Place the small double-neck round bottom flask to receive the ethanolic eluate of the Sep-
          Pak in its corresponding location in the Box.
     16. In a lead pot, lace a sterile empty vial (30 mL size) fitted with a vent needle/filter and a
          second needle and Millev GV filter for sterilization of the final product.
     17. Place enough dry-ice/acetone mixture in the vacuum trap.
     18. Check that the nitrogen and compressed air valves close to the hot cell are open (also
          check the nitrogen cylinder).
     19. Assure that syringes and needles have been changed. Use depyrogenated reaction vessel
          (with magnetic stirrer), HPLC collection bulb (10 mL size) V-vial, two neck round-bottom
          flask, sterile vial etc.
     20. Keep the HPLC column flow at 2 mL/min.


Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                           Page 12 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                  Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


Synthesis comments:
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________




The approved procedure has been followed without any deviation


Chemist_____________________________Initials_________________Date________________




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures             Page 13 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                             Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892




                                                       SOP # MP204
                               18
           Production of [ F]Fallypride for Injection; Part 2: synthesis and purification


Approved by: _____________________________Initials________ Date: _____________
                 Victor W Pike, Ph.D., Chief, NIMH, PET Radiopharmaceutical Sciences

                                                                 18
                       Purpose: to synthesize and purify [ F]fallypride


                 18
Procedures: [ F]Fallypride synthesis and purification
                               18                                                                    18
     1. Preparation of [ F]fluoride ion/Kryptofix 2.2.2-potassium carbonate complex ([ O]water-
                                            18
          acetonitrile-Kryptofix 2.2.2-[ F]fluoride ion-potassium carbonate).
               1.1. Using the dose calibrator (Biodex AtomLab 300) set at F-18, determine the amount
                      of radioactivity that will be transferred to the clean glassy reaction vessel (20 mL
                                                                                18            18
                      size).   Record the data: EOB, starting activity of [ F]fluoride-[ O]water and
                                    18            18
                      Volume of [ F]fluoride-[ O]water.
               1.2. Connect one vent needle, one needle and line from external syringe containing
                      acetonitrile (0.5 mL) and one spinal needle and line going from the radioactive
                      solution through V2 to reactor vial. Make sure that spinal needle is all the way
                      down to the bottom of the radioactive solution.
     2. Starting the preparation and transferring the F-18 complex.
               2.1 Start the TRACERLab automated software program. Select Synthesis: "Fallypride"
                      in method box, click “Synthesis-Start” in main menu bar, follows process
                      instructions. Organize "Instrument screen", "Program" and "Graphics" in the
                      computer screen.
               2.2 The program will have already started setting the vacuum pump ON and
                      transferring the F-18 complex from the V-vial (1 mL size) into the reaction vessel,
                      then MESSAGE BOX comes out. At this moment add acetonitrile (0.5 mL) from the
                      outside syringe into the transport F-18 vial (pushing with air until the line is empty
                      and the vial is full). Press “Enter” or click “OK” when done.
          18
     3. [ F]Fallypride synthesis step 1: remove water from the F-18 complex.




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                           Page 14 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                           Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


          The reactor temperature will be increased first to 65 ºC and later to 88 ºC (T1 + 3 min) and
          then back to 60 ºC. At this point, the contents of V5 are added and the temperature cycle
          starts again: heat at 65ºC, then at 88ºC and finally at 60 ºC (drying process).
          18                                             18
     4. [ F]Fallypride synthesis step 2:                   F-Fallypride reaction. Check V4 and reaction
     temperature.
          4.1 Once drying is complete, V4 will open and the tosylate precursor in acetonitrile (1 mL)
               will be added to the reaction vessel and the mixture heated at 100 ºC for 30 min. At
               the end of the reaction, the reaction vessel will be cooled at 35 ºC and the first part of
               the Program will finish.
          4.2 A message will shown saying "End of synthesis, Hit OK to continue to next Program".
               Hit CONTINUE.
     5. Prepare Beckman HPLC system for purification.
          5.1 Verify that the HPLC is in Preparative and in LOAD and that the detectors are in the
               right settings.
          5.2 Check that method setting of Beckman HPLC instruments is right
          5.3 Set the flow rate to 9 mL/min. Wait for pressure to go up.
          5.4 Click “Single run” in Beckman HPLC software to start acquisition and wait the
               acquisition Box appears.
          5.5 Check that the injector turn to inject position
        18
     6. [ F]Fallypride purification step 1: transferring the initial reaction mixture.
          6.1 Second program continues purification process. The reaction mixture will be
               transfered to the V- vial (10 ml size) containing sterile water (0.5 mL) for pre-dilution
               before HPLC injection.
          6.2 Check that the injector transfer needle in V- vial (10 mL size) is far up from the level of
               the solution.
          6.3 Check that valve 6 will open and the acetonitrile (1.5 mL) is added to the reaction
               vessel.
         18
     7. [ F]Fallypride purification step 2: loading and injection
          7.1 After transferring, a message box will appear "System now ready to load Loop. Hit OK
               when completed". System will load the reaction mixture solution from T-vial (10 mL
               size) into HPLC loop automatically.
          7.2 At this moment, new message box will appear “System now ready to inject. Hit OK
               when the loading is completed.”


Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                      Page 15 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                         Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


          7.3 Watch the 10 mL T-vial closely. While the solution loading completed, hit OK.
          7.4 At same time hit “OK to start the HPLC run in Beckman software
         18
     8. [ F]Fallypride step 3: HPLC separation and product collection.
          8.1 In GRAPHICS observe the UV and and radioactivity traces on the data acquisition
               systems.
          8.2 When the desired peak start coming out click on the collect arrow. Record the Rt
               =____________min.
          8.3 The mobile phase containing the product will be added to the bulb vessel containing
               sterile water (90 mL) for dilution and of sodium acetate (10 mL) for dilution and
               neutralization.
          8.4 At the end of collection click on the end collection arrow to stop fraction collection (the
               mobile phase will go into the waste reservoir).




Synthesis comments:
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________


The approved procedure has been followed without any deviation


Chemist_____________________________Initials_________________Date________________




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                    Page 16 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                        Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892




                                                    SOP # MP205
                                         18
                      Production of [ F]Fallypride for Injection, Part 3: formulation


Approved by: _____________________________Initials________ Date: _____________
                 Victor W Pike, Ph.D., Chief, NIMH, PET Radiopharmaceutical Sciences


                                                         18
                      S   Purpose: Formulation of [ F]Fallypride for Injection


                 18
Procedures: [ F]Fallypride formulation
    1. The program will continue Fallypride Production Part 2 and the diluted fraction will be
         passed through the solid phase cartridge (SPE; Waters C-18 Sep-Pak) for concentration.
         This process is programmed for 6.5 min. After passing through Sep-Pak, valve V9 will open
         and Sep-Pak will be washed with sterilized water.
    2. After the water wash, V8 will open and the product will be eluted from the Sep-Pak with
         ethanol (1mL) into the two-neck round bottom flask.
    3. Program will stop to allow sampling of ethanolic solution if necessary. The message "Take
         QC sample and hit OK to proceed with filtration" will appear. Take 40-100 µL of sample for
         QC from the two-neck round bottom flask if needed.
    4. The program can be continued hitting OK. Valve 7 will open and saline solution will be
                                                                       18
         added to the neck round bottom flask containing [ F]fallypride product.
    5. The program can be continued hitting OK. The saline solution will be then transferred to
         the sterile and non-pyrogenic vial (10 mL size, Flip-top- Vial – Glass) through the Millex
         filter Unit (0.22 µm) for filtration and sterilization.
                                                                                           18
    6. After filtering product, remove sterile filter, measure the activity of the final [ F]fallypride
         product, record the number on the master batch Sheet and fill the number in the labels.
                             18
          Activity of final [ F]Fallypride for Injection for _____________mCi at____________;
         The volume:_____mL and the concentration:___________ mCi/mL.
         The final Collection Vial and the lead shielded pot will be labeled as follows (21 CFR 361.1):




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                   Page 17 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                                                        Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


                                       [18F]Fallypride for Injection
                        Sterile, apyrogenic solution for intravenous administration
                        Caution: New drug limited by Federal law to investigational use 0nly
                       NIMH MIB       Store at room temperature                  Expires 4 h after calibration
                       Activity:______________ mCi                                Time: _________________
                       Volume: ___________           mL.                          Concentration: ______________
                       Batch #: FAY-________________                              Date:_____________
         In addition, one of the labels will be attached to the Radiopharmacy Form as record.
     7. Sampling for QC: The 1 cc sterile polypropylene syringe may be used to withdraw a 1 mL
                                                       18
          sample of the formulated [ F]Fallypride for Injection in a certified laminar flow sterile
                                                                   18
          cabinet. The 1 mL sample of [ F]Fallypride for Injection is transferred to sterilized
                                                          18
          depyrogenated tube labeled “[ F]Fallypride for Injection” and the vial is dated and stored in
          a lead pig.         This aliquot will be used for the following QC tests: radioconcentration,
          radiochemical purity, specific radioactivity, pH, residual solvents, and bacterial endotoxins
          (LAL test).        Standard operating procedures for these tests can be found in the quality
          control and radiopharmacy forms.
     8. Post-Synthesis Procedures:
          8.1 HPLC and columns clean up: set the flow to 6 mL/min. and move the B pumps line into
               the solvent bottle, which contains acetonitrile only. Keep the acetonitrile as 70%. Run
               50-60 min and then stop running. Close the Beckman HPLC software.

          8.2 Machine turn off: Turn off nitrogen and compressed air valves located outside hot-cell.
              On the TRACERLab click synthesis – Reset in main menu to set all valves and vacuum
              on default status, then close the software. It is ready for next Box Clean procedure
              (SOP # MP201).


Synthesis comments:

______________________________________________________________________________
____________________________________________________________________



The approved procedure has been followed without any deviation


Chemist_____________________________Initials_________________Date________________




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                                                   Page 18 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                  Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892




Diagram 1. Layout of GE TRACERlab FXFN Module




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures             Page 19 of 34
         [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                            PROCEDURES
    PET Radiopharmaceutical Sciences Section,                                       Date of review: 05/17/04
    Molecular Imaging Branch,
    National Institute of Mental Health,
    National Institutes of Health,
    Bldg. 10, Rm. B3 C338,
    Bethesda, MD 20892




                                                          SOP # QA301
                                       Standard Curve of Reference Fallypride




    Approved by: _____________________________Initials________ Date: _____________
                       Victor W Pike, Ph.D., Chief, NIMH, PET Radiopharmaceutical Sciences



 S Purpose: To set a standard curve for authentic fallypride for calculation of the mass of fallypride
in a in an analysis sample.


         A) Stock solution of reference fallypride.
              Weigh an accurate mass of fallypride (1−2 mg). Disolve in HPLC mobile phase (10−25
              mL) in a volumetric flask for QC. Mix thoroughly for several minutes. When not in use,
                             ○
              store at −20 C in the freezer.
                                  Concentration of solution = ~ 0.1 µg/µL


         B) "Daily" dilution.
              From the stock solution make a dilution 1: 100:
                       Example: take 100 µL of Stock solution.diluted with solvent (acetonitrile: HPLC
                        water 65: 35 v/v) in a volumetric flask (10 mL size)
                                  Concentration: ~ 1−2 ng/1.0 µL


                        nmoles of fallypride (M.W. = 364.45) in this dilution:
                                                     -3
                        1 ng/364.45 = 2.74 x 10 nmoles in 1.0 µL


              Mix well to ensure homogeneity. When not in use, store at 4 ºC under refrigeration.


         C) Procedure for obtaining standard curve
                   1. Open the Beckman 32 karat software and select the instrument ”Analysis System”
                        to start HPLC system.




    Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                  Page 20 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                              Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


               2. Set the analytical HPLC System (Beckman Coulter) in working conditions and
                    keep flow for 30 min for column equilibration.

                     HPLC System conditions:
                     Column: Prodigy ODS (3) (100 μ; 4.6 x 250 mm; Phenomenex)
                     Mobile Phase: Acetonitrile-ammonium formate (5 mM) (50: 50 v/v)
                     Flow Rate: 1.0 mL/min.
                     UV Wavelength: 305 nm

               3. Select the method ”FallyprideQ.C” in method box and check the instrument setup
                    on the “Instrument Setup” box, then close box.
               4. Using the "daily" dilution make injections into the HPLC system varying the
                    injected volume, measuring the areas and ensuring that plate count and peak
                    shape is consistent.
               5. Suggested volumes of "Daily" dilution: 10, 20, 60, 80, 100 µL.
               6. Make two to four injections of each volume.
               7. Perform Regression Analysis on the data, plotting area units versus µg or µmoles.
               8. Using the external or internal standards, calculate the slope of calibration line (a),
                    Y-axis intercept of calibration line (b) and correlation coefficient (r).
               9. Following is linear calibration fit of external or internal standards:
                    The equation for calculating the uncorrected amount is:
                                                                  Y = ax+b




Chemist_____________________________Initials_________________Date________________




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                         Page 21 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                          Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


                                                    SOP # QA 304
                         Analysis of Organic Residues by Gas Chromatography
                                              18
                                          in [ F]Fallypride for Injection


Approved by: _____________________________Initials________ Date: _____________
                          Victor W Pike, Ph.D., Chief, NIMH, PET Radiopharmaceutical Sciences


                                                                           18
               Purpose: to analyze for volatile solvent residues in [ F]Fallypride for Injection


     1. Responsibility -
          1.1 It is the responsibility of the Chief of PET Radiopharmaceutical Sciences to ensure
               that personnel are trained in this procedure.

          1.2 It is the responsibility of personnel to adhere to this approved SOP.

          1.3 Procedure adheres to GMP/GLP guidelines.

     2. Scope - Quality control testing of volatile organic solvents in NIMH produced PET
          radiopharmaceuticals.
     3. Reference Document(s) –
          3.1 6850A GC User Information
          3.2 Installation, operation and maintenance manual for hydrogen
              generator (Model H2-90; Parker Balston). (Bulletin TI-H2-90C)
     4. Safety Precautions

         4.1 Radiation Safety – ALARA (As Low as Reasonably Acceptable)

        4.2 Chemical laboratory safety

     5. Materials and equipment

          5.1 Agilent 6850 GC with flame ionization detector (FID)

          5.2 Agilent 6850 series autosampler

          5.3 J & W DBWAX column, (30 m (l) x 0.25 mm (id) x 0.25 μm (film thickness) (Alltech,
               part # 122-7032)




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                     Page 22 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                           Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


          5.4 Acquisition and data processing software: GC Chem Station (version: Rev. A.09.03
               [1417] )

          5.5 Inlet liner: split inlet glass liner with glass wool packing (Agilent part number, 5183-
               469119251-60540)

          5.6 Parker Balston H2-90 Hydrogen Generator

          5.7 High purity grade (99.995 %) compressed helium compressed (Roberts Oxygen,
               cat.no. R 102 F3)

          5.8 In -house air purified by Parker Balston Zero Air Generator, Model 75-83NA

          5.9 In -house deionized water (18 mega ohm) purified by Millipore Milli-Q j.) Autosampler
               glass vial ( Agilent part no. 5182-0864) k.) Autosamper conical glass insert (Agilent
               part no. 5183-2085)

          5.10 Pipetman 200 L pipet (NIH stock # 6640-02-032-1955)

          5.11 386 ppm internal standard aqueous solution of propionitrile prepared via 1 in 10
               dilution of a 3860 ppm solution (0.5 mL propionitrile diluted to 100 mL mark with
               water).

     6. GC system configuration.

          6.1 Injection port: split sample injection split ratio of 20:1, 250 ˚C

          6.2 Carrier gas: Helium; 2 mL/min

          6.3 Column temperature gradient: column temperature is initially operated at 50˚C and
               held at this temperature for 1 min, and then increased to 150 ˚C at a rate of 20˚C /min.
               The temperature is held at 150˚C for 0.5 min and then increased to 220˚C at a rate of
               50˚C/min.      After 3 min at 220˚C the column temperature is returned to starting
               temperature of 50˚C.

          6.4 Detector: FID with hydrogen at 40 mL/min and air at 450 mL/min.            Helium make up
               flow: 45 mL/min. Detector at 250 ˚C.

          6.5 Autosampler: 10 L syringe. Sample injection volume: 1 L.

          6.6 Needle/Syringe wash:            four times prior to injection of sample and two times after
               injection.


Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                      Page 23 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                              Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


     7. Procedure before data acquisition. Ensure that the water level in the H2-90 hydrogen
          generator is above the lower limit.              Otherwise, pour in 18 mega ohm water into the
          reservoir until the level is just below the upper limit of water reservoir. Check the hydrogen
          pressure is about 28 p.s.i. Check helium pressure is ca. 60 p.s.i. on main cylinder gauge.
          Make sure that the solvents A and B (in the autosampler tray) for rinsing injection syringe
          needle are filled with deionized (18 mega ohm water). Via a 200 L Pipetman pipettor,
          pipette 50 L of [ F]Fallypride for Injection (test sample) into a glass insert housed in a GC
                               18


          autosampler vial. Discard the radioactive pipette tip in radioactive waste. Add 50 L of the
          internal standard solution of propionitrile (386 ppm) to the test sample.                   Cap the
          autosampler vial with septum.             Tap the bottom of the autosampler vial to remove air
          bubbles. Place autosampler vial in autosampler rack and note rack position. On the GC
          Control panel, use up or down arrow buttons to verify that FID is lit and the background
          signal is ca. 5.
                                    18
     8. Data acquisition of [ F]FALLYPRIDE test sample spiked with internal standard. Double
          click “Instrument 1 online” on the desktop.               Select Method and Run Control from drop
          down men bar below the file menu bar. Select ISPRN.M from drop down menu box on the
          right side of Method and Run Control Box. Go to sequence on the main menu and select
          sequence parameter. Type in subdirectory for radiopharmaceutical that will be tested (i.e.
          Fallypride). Select Sequence Table from Sequence drop down menu. Enter 1 for the
          number of injections per sample and location of the injection. Enter the sample name, file
          name, and injection volume (1.0 L). When the system shows a ready sign (above start
          button), hit the start button. To view on line signal, go to view on the main menu bar and
          select on line signal/signal window 1. After 3.5 min, the GC chromatogram and report can
          be generated; this is performed by going to the desktop and opening “Instrument 1 offline”.
          On the left side of the screen, scroll the drop down menu to “data analysis”. Under the file
          menu, scroll to “load signal. “ Find the file name in the appropriate subdirectory on the C
          drive (e.g. C:\. . . \data\Fallpride). After loading the signal, scroll down to “print preview . . .
          report” that is found in the file menu. The chromatogram and report should appear on the
          computer screen and this can be printed out be hitting the print button on the bottom of the
          report page. Attach the chromatogram and report to the quality control form.               Note that
          18
          [ F]Fallypride fro Injection may be released before completion of the residual solvent test
          (see section



Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                         Page 24 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                      Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


     9. Post-run method.           Remove all samples and label radioactive samples.   Download the
          method Default.M that is used to maintain the oven temperature at 150 ˚C when GC is idle.



Chemist_____________________________Initials_________________Date________________




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                 Page 25 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                        Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


                                                          SOP #QA305

                                           Analytical HPLC QC Method
Approved by: _____________________________Initials________ Date: _____________
              Victor W Pike, Ph.D., Chief, NIMH, PET Radiopharmaceutical Sciences

                                                                    18
           Purpose: to perform analytical HPLC QC on [ F]Fallypride for Injection


Procedure
     1. Set the analytical HPLC system (Beckman Coulter) in working conditions and maintain flow
          for 40 min for column equilibration.
                     HPLC system conditions:
                     Column: Prodigy ODS (3) (100 μm; 4.6 x 250 mm; Cat. # 00G-4244-E0;
                     Phenomenex)
                     Mobile phase: Acetonitrile-5 mM ammonium formate (50: 50 v/v)
                     Flow rate: 1.0 mL/min.
                     UV Wavelength: 305 nm
     2. Download the analytical method “Fally_analysis_secondSys” or “Fally_analysis_firstSys”
          from method box in Beckman software.
     3. Validation of HPLC system
                   Inject standard fallypride (20−40 µg), which is already prepared according to SOP
                    # QA301, and record the exactly mass (ng) injected and retention time of fallypride
                    standard.
                   Calculate the mass of fallypride corresponding to the peak of fallypride in HPLC
                    chromatography, based on the standard curve of fallypride.
                   Compare the mass of the fallypride injected with the mass calculated, the
                    difference should be equal or less then 10%. (Recovery ≤ 90%)
     4. HPLC system cleaning
                   Inject solvent (100 µL) once or twice to clean the system in 10-15 min after
                    standard fallypride running. Check that only the solvent front peak should shows in
                    the chromatography.
                   Inject 10% ethanol in saline (100 µL). Check that only the solvent front and saline
                    peaks show in the chromatography.



Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                   Page 26 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                          Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892

                                   18
     5. QC of final product: [ F]Fallypride for Injection
               
                                                                             18
                    Measure the radioactivity of sample of final product of [ F]Fallypride for Injection
                    (50−150 L) before and after injection onto the HPLC system in hot-cell 5, and
                    record the number in QC record sheet. Calculate the activity injected (decay
                    correlation)
                   Inject the sample of [ F]Fallypride for Injection (50−150 L depending on the
                                                 18


                    radioactivity concentration of formula).
               
                                                                                        18
                    Check the retention time (Rt) of the standard and the [ F]fallypride for

                    radiochemical identity (the difference of Rt < I min)

               
                                                                              18
                    Calculate the mass corresponding the UV area of the [ F]fallypride peak in the
                    chromatograph, based on the standard curve of fallypride.
                   Calculate the specific radioactivity according to the radioactivity injected and the
                    mass calculated at HPLC injection time.
                   Calculate the specific activity at the time of HPLC injection, EOS and EOB (decay
                    correction).
                   Calculate the mass (g) of whole formula based on the radioactivity of final
                    formula, which recorded on master batch Sheet Page 4, and the specific activity at
                    EOS.
                   Record the chemical purity, radiochemical purity, the concentration of the final
                    formula and the expired time.
     6. Post-analysis HPLC system and column cleaning: to run the HPLC system for 40−50 min
          with the solvent (acetonitrile-HPLC water; 70: 30 v/v) for cleaning the HPLC system and
          the column. Turn off the HPLC system and software.


Chemist_____________________________Initials_________________Date________________




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                     Page 27 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                             Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


                                                    SOP # QA306
                                                     Sterility Test
Approved by: _____________________________Initials________ Date: _____________
              Victor W Pike, Ph.D., Chief, NIMH, PET Radiopharmaceutical Sciences


                                                             18
                     Purpose: To test the sterility of [ F]Fallypride for Injection



     Notes.


     1. Routine sterility tests will be run by the Clinical Microbiology laboratory of the Department of
          Laboratory Medicine at NIH, through a FDA approved procedure.


     2. The radiochemistry lab will provide a sample of each formulation (in its original filtration
          vial), 24 h after its preparation or when no traces of radioactivity are detected anymore (ten
          or more half life periods).


     3.   Before their transportation to the testing laboratory, the samples will be stored in the
          refrigerator until decay.


     4. A log book will be kept in the radiochemistry lab indicating the date the samples were
          transported, their lot number and the date and results of the test (readings at 7 and 14 d
          should be indicated).


Chemist_____________________________Initials_________________Date________________




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                        Page 28 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                              Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892




                                                    SOP # QA307


               Pyrogen Test (LAL Bacterial Endotoxin Test). Simplified Procedure


Approved by: _____________________________Initials________ Date: _____________
              Victor W Pike, Ph.D., Chief, NIMH, PET Radiopharmaceutical Sciences

                                                             18
                      Purpose: to test the sterility of [ F]Fallypride for Injection


Materials
     LAL test kit vials (e.g., Endosafe Inc., Charleston, SC or Cape Cod Associates, Falmouth MA)
          Two vials 0.125 EU/mL(one for test sample, one for negative test)
          One positive vial (5 EU/mL)
          Sterile saline solution (from the same vial used for the final formulation)
          Pyrogen-free test tubes (if not provided with kit)
          Micropippets and tips (50 µL, 100 µL, 500 µL)
          Tuberculin syringes (0.5 mL, 1 mL)
          Incubator, 37°C
LAL Test Procedure: See “RADIOPHARMACY TEST RESULTS” form

Simplified Test Procedure ( it may not be used as a procedure of LAL test.)

          a) From the final formulation in saline, take 50 µLand add to the test sample tube. To the
               same LAL tube add 150 µLof saline (for a total of 200 µL).

          b) To the negative test tube add 200 µL saline only.

          c) To the positive test tube add also 200 µL saline.

          d) Incubate all vials at 37 °C for 1 h. Remove the test vials at 20 min and in one smooth
               careful motion, invert each tube (without totally disturbing the gel) and note gel
               formation.

          e) Continue incubation of the vials for the full 60 min. Again in one smooth motion, invert
               each vial and note gel formation.




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                         Page 29 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                            Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


          f)   The test sensitivity equals the concentration of the test vial multiplied by the dilution
               factor. If the test sample does not gel, report the result as less than the test sensitivity.
               If the test sample gels, report as greater than or equal to the test sensitivity.

          Note. The results at 20 min will allow release of the formulation for injection and the 60 min
          samples will confirm the results.




  Chemist_____________________________Initials_________________Date______________




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                       Page 30 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                        Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892


                                                          SOP #QA308


                  Analysis of the Purity of Fallypride and Tosyl-Fallypride by HPLC


Approved by: _____________________________Initials________ Date: _____________
              Victor W Pike, Ph.D., Chief, NIMH, PET Radiopharmaceutical Sciences


                Purpose: to determine the purity of Tosyl-Fallypride by HPLC



The HPLC System used in this method:
          HPLC pump: System Gold 126 Solvent Model (Beckman Coulter)
          UV detector: System Gold 166 Detector (Beckman Coulter)
          Sample Injector: Rheodyne with 200 µLloop
The HPLC conditions used in this method:
          Column: Prodigy ODS (10 µm, 4.6 X 250 mm, 100 A)
          Mobile Phase: Acetonitrile-5 mM ammonium formate (50: 50 v/v) at 2 mL/min
          Flow rate: 1 mL/min.
          PDA: 254 nm
Procedure
          1. Weigh 1−2 mg of precursor (tosyl-fallypride) in vial (20 mL size)
          2. Dissolved the sample in of acetonitrile (10−20 mL). The concentration of this solution is
               ~ 1 µg/µL.
          3. Inject 10 µLof the solution of tosyl-fallypride into HPLC and run for 30 min.
          4. Use the Beckman method “Fallypride_Analysis_SecondSys” to interrogate the
               Chromatograph, and get the Tosyl-fallypride purity (UV area percentage).




Chemist_____________________________Initials_________________Date__________




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                     Page 31 of 34
       [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                          PROCEDURES
PET Radiopharmaceutical Sciences Section,                                                 Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892




                                                      Attachment
This attachment lists the materials and equipments, which are used in the SOPs on Page 1.
Materials:
                                                                                           Quality grade and
Item    Name                                                       Name of supplier        catalog #
1       4,7,13,16,21,24-Hexaoxa-1, 10-diazabicyclo                 Aldrich Chem. Co.       Anhydrous, 98%
        [8.8.8] hexacosane (Kryptofix 222)                         1001 West St. Paul      Cat. # 29110-1G
                                                                   Ave., Milwaukee, WI
                                                                   53233
2       Potassium carbonate                                        Aldrich Chem. Co.       99.995%
                                                                   1001 West St. Paul      Cat. # 367877-10G
                                                                   Ave., Milwaukee, WI
                                                                   53233
3       Acetonitrile (anhydrous)                                   Aldrich Chem. Co.       Anhydrous, 99.8%
                                                                   1001 West St. Paul      Cat. # 271004-100
                                                                   Ave., Milwaukee, WI     ML
                                                                   53233
4       Acetonitrile                                               Burdick and Jackson,    HPLC Grade, ACS
                                                                   B&J Brand               Cat. # 017-4
5       Water                                                      Burdick and Jackson,    HPLC Grade, ACS
                                                                   B&J Brand               Cat. # 4218
6       Acetone                                                    Fisher Scientific       HPLC Grade, ACS
                                                                                           Cat #: 4218
7       Triethylamine                                              Aldrich Chem. Co.       HPLC Grade, ACS
                                                                   1001 West St. Paul      Cat #: 471283-
                                                                   Ave., Milwaukee, WI     500ML
                                                                   53233
8       Sodium acetate                                             Aldrich Chem. Co.       99+% ACS
                                                                   1001 West St. Paul      Cat. #: 241245-
                                                                   Ave., Milwaukee, WI     500G
                                                                   53233




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                            Page 32 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                                Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892




9       Acetic acid                                                Malinckodt             99+% ACS
                                                                   Chemicals              Cat. # V193-14
10      Luna C18 (10 m; 100 Å; 250 x 21.2 mm)                     Phenomenex             Cat. # 00G-4094-
                                                                                   th
                                                                   2320 W. 205 St.        P0
                                                                   Torrance, CA 90501-
                                                                   1456
11      Fallypride                                                 ABX,                   > 90%
                                                                   Wilhelm-RÖnsch Str.    Cat. # 156.0025
                                                                   9. D-01454
                                                                   Radeberg, Germany
12      Tosyl fallypride                                           ABX,                   > 90%
                                                                   Wilhelm-RÖnsch Str.    Cat. # 155.0002
                                                                   9. D- 01454
                                                                   Radeberg, Germany
13      Ethyl alcohol (200 proof)                                  The Warner-Graham      CAS # 64-17-5
                                                                   Company,
                                                                   Cockeysville, MD
14      Sodium Chloride for Injection (0.9% w/v;                   American               USP
        USP                                                        Pharmaceutical         CAT#:918610
                                                                   Partner, Inc.
15      Sterile empty vial (10 mL)                                 Abbott laboratories    USP
                                                                                          Cat. # 5816-11
16      Sep-pak Cartridges Plus C18                                Waters Corporate       Cat. # WAT020515
                                                                   34 Maple Street,
                                                                   Milford,
                                                                   MA. 01757 USA
17      Millex filter (25 mm; 0.22 m pore size)                   Millipore              Cat. # SLGVR35LS
18      Millex GV filter (25 mm; 0.22 m pore size;                Millipore              Cat. # SLGV004SL
        4 mm diameter)




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                           Page 33 of 34
     [18F]FALLYPRIDE FOR INJECTION: STANDARD OPERATING
                        PROCEDURES
PET Radiopharmaceutical Sciences Section,                                                Date of review: 05/17/04
Molecular Imaging Branch,
National Institute of Mental Health,
National Institutes of Health,
Bldg. 10, Rm. B3 C338,
Bethesda, MD 20892




     Instruments and equipment:
         Operation/Function                   Manufacturer                    Model               Serial #
     Radiosynthesis                     General Electric               TRACERlab FXFN       14390
     HPLC purification                  Beckman Coulter                System Gold 126      342-2187
                                                                       Solvent Model
     UV absorbance detection            Beckman Coulter                System Gold 166      332-2187
     in HPLC purification                                              Detector
     Radioactivity detection in         Bioscan                        Flow Counter         0409-316
     HPLC purification




Document 5. [18f]Fallypride for Injection: Standard Operating Procedures                            Page 34 of 34

				
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