ANTIEPILEPTIC DRUGS kit

ANTIEPILEPTIC DRUGS IN SERUM/PLASMA
LC-MS/MS KIT

Epilepsy is defined as seizure disorder induced by surge of electrical activity in the brain. Anticonvulsant or antiepileptic medications encompass a broad range of drugs that are primarily intended to control ongoing or anticipated future seizures as well as being beneficial in non-epileptic conditions such as psychiatric disorders and pain syndromes. Many antiepileptic drugs (AEDs) have a narrow therapeutic range thus relatively modest alterations of drug concentration in plasma can lead to loss response or toxic effects. Therefore, it is necessary to simultaneously determine AEDs including their active metabolite to improve treatments without undesirable side effects.

  • Highlights

    • Simultaneous measurement the concentrations of 21 antiepileptic drugs
    • Total run time is 10.5 min.
    • Just a few pipetting steps for the sample treatment-protein precipitate and shoot approach-no need for derivatization
    • Each drug safeguarded by its own internal standard
    • Consuming small volume of patient’s sample
    • Long life span of HPLC column
  • Matrix

    • Plasma
    • Serum

METHOD SCOPE

Jasem method ensures to simultaneously measure concentrations of 21 antiepileptic drugs in human serum with a run time of 10.5 minutes consuming small volume of patient’s serum/ plasma with only protein precipitation

Specimen: Human serum/plasma

Analysis Time: 10.5 min

Injection Volume: 5 µL

Sample Preparation


  • Step 1

    Pipette 50 µL of serum/plasma patient’s sample (calibrator/control) into a glass centrifuge tube

  • Step 2

    Add 30 µL of internal standard solution and vortex for 5 sec.

  • Step 3

    Add 400 µL of Reagent-1, vortex for 5 sec. then, centrifuge at 4000 rpm for 5 min.

  • Step 3

    Decant the clear supernatant into HPLC vial prior to injection

  • Total ion chromatogram of AED’s method

  • Retention Times of Compounds-Internal Standards

    Compound Name

    RT (Min.)

    Labelled-Internal Standard

    Levetiracetam

    3.87

    d6-Levetiracetam

    Gabapentin

    3.98

    d4-Pregabalin

    Pregabalin

    3.98

    d4-Pregabalin

    Ethosuximide

    3.98

    d4-Pregabalin

    Zonisamide

    4.36

    15N,d4-Zonisamide

    Lamotrigine

    4.46

    13C3-Lamotrigine

    Rufinamide

    4.54

    d4-Felbamate

    Felbamate

    4.55

    d4-Felbamate

    Primidone

    4.70

    d5-Primidone

    Lacosamide

    4.71

    d3-Lacosamide

    Carbamazepine Epoxide

    4.83

    d4-Oxcarbazepine

    Phenobarbital

    4.93

    d5-Phenobarbital

    Oxcarbazepine-Metabolite

    4.93

    d4-Felbamate

    N-Desmethylmethsuximide

    4.96

    d12-Topiramate

    Oxcarbazepine

    5.07

    d4-Oxcarbazepine

    Topiramate

    4.95

    d12-Topiramate

    Phenytoin

    5.21

    d4-Oxcarbazepine

    Tiagabine

    5.22

    d6-Tiagabine

    Carbamazepine

    5.35

    d10-Carbamazepine

    Valproic Acid

    6.19

    d6-Valproic Acid

    Stiripentol

    6.60

    d9-Stiripentol

  • Quantitation Limit

     

     

    Agilent 6460 LC-MS/MS

    Agilent 6470  LC-MS/MS

    Agilent Ultivo LC-MS/MS

     

    Analyte

     

    LOQ mg/L

     

    1

    Levetiracetam

    2,92

    0,49

    1,97

    2

    Gabapentin

    0,42

    0,09

    0,29

    3

    Pregabalin

    0,41

    0,06

    0,13

    4

    Ethosuximide

    2,07

    1,42

    5

    Zonisamide

    1,59

    0,47

    0,56

    6

    Lamotrigine

    1,09

    0,27

    0,24

    7

    Felbamate

    3,88

    1,32

    1,57

    8

    Primidone

    0,95

    0,30

    0,32

    9

    Lacosamide

    0,36

    0,13

    0,10

    10

    Oxcarbazepine Metabolite

    2,38

    0,14

    1,02

    11

    Phenobarbital

    3,07

    1,95

    1,42

    12

    Carbamazepine 10,11-epoxide

    0,46

    0,40

    23,11

    13

    Phenytoin

    0,73

    0,49

    87,49

    14

    Topiramate

    0,39

    0,17

    0,11

    15

    N-Desmethylmethsuximide

    3,36

    1,29

    1,83

    16

    Oxcarbazepine

    0,23

    4,11

    0,02

    17

    Tiagabine

    0,02

    0,01

    0,01

    18

    Carbamazepine

    0,87

    0,10

    0,24

    19

    Rufinamide

    2,31

    0,68

    2,15

    20

    Valproic acid

    3,74

    1,59

    2,87

    21

    Stiripentol

    0,64

    0,12

    0,06

  • Recovery

    Analyte

    Conc.

    Agilent 6460 LC-MS/MS

    Agilent 6470 LC-MS/MS

    Agilent Ultivo  LC-MS/MS

    % Recovery

    % Recovery

    % Recovery

    Carbamazepine

    4,67 mg/L

    99,45

    106,83

    115,00

    Carbamazepine Epoxide

    1,66 mg/L

    87,26

    112,75

    114,00

    Ethosuximide

    21,2 mg/L

    105,06

    103,00

    Felbamate

    18,80 mg/L

    98,15

    106,86

    110,00

    Gabapentin

    4,65 mg/L

    99,46

    104,27

    103,00

    Lacosamide

    2,43 mg/L

    99,91

    106,91

    107,00

    Lamotrigine

    5,0 mg/L

    94,33

    114,6

    111,00

    Levetiracetam

    11,9 mg/L

    107

    105,47

    114,00

    N-Desmethylmeth suximide

    9,89 mg/L

    96,63

    106,17

    102,00

    Oxcarbazepine

    0,87 mg/L

    95,32

    95,49

    95,00

    Oxcarbazepine Metabolite

    7,68 mg/L

    99,61

    107,81

    114,00

    Phenobarbital

    9,65 mg/L

    93,42

    97,14

    98,00

    Phenytoin

    4,91 mg/L

    98,58

    115,33

    107,00

    Pregabalin

    1,51 mg/L

    98,13

    112,6

    113,00

    Primidone

    5,6 mg/L

    93,32

    119,83

    113,00

    Stiripentol

    3,32 mg/L

    95,68

    99,85

    102,00

    Tiagabine

    0,06 mg/L

    101,33

    99,81

    97,00

    Topiramate

    3,26 mg/L

    86,65

    97,62

    98,00

    Valproic Acid

    22,60 mg/L

    98,72

    111,59

    105,00

    Zonisamide

    8,29 mg/L

    96,26

    100,59

    100,00

    Rufinamide

    8,93 mg/L

    97,82

    100,86

    90,00

  • Repeatability

    Analyte

    Conc.

    Agilent 6460

    Agilent 6470

    Agilent Ultivo

    LC-MS/MS

    LC-MS/MS

    LC-MS/MS

    % CV

    % CV

    % CV

    Carbamazepine

    4,67 mg/L

    4,41

    0,86

    1,17

    Carbamazepine Epoxide

    1,66 mg/L

    12,85

    0,95

    1,04

    Ethosuximide

    21,2 mg/L

    2,5

    1,64

    Felbamate

    18,80 mg/L

    7,47

    1,69

    3,18

    Gabapentin

    4,65 mg/L

    4,17

    0,92

    1,24

    Lacosamide

    2,43 mg/L

    6,13

    1,08

    1,1

    Lamotrigine

    5,0 mg/L

    6,43

    1,92

    2,32

    Levetiracetam

    11,9 mg/L

    5,46

    1,08

    4,24

    N-Desmethylmeth suximide

    9,89 mg/L

    8,55

    8,55

    3,35

    Oxcarbazepine

    0,87 mg/L

    6,19

    0,95

    0,99

    Oxcarbazepine Metabolite

    7,68 mg/L

    6,12

    1,9

    2,87

    Phenobarbital

    9,65 mg/L

    13,9

    6,52

    2,53

    Phenytoin

    4,91 mg/L

    4,91

    1,88

    1,62

    Pregabalin

    1,51 mg/L

    8,39

    1,11

    1,51

    Primidone

    5,6 mg/L

    7,58

    1,2

    1,96

    Stiripentol

    3,32 mg/L

    7,57

    0,88

    0,42

    Tiagabine

    0,06 mg/L

    4,26

    2,79

    3,65

    Topiramate

    3,26 mg/L

    5,86

    1,92

    1,36

    Valproic Acid

    22,60 mg/L

    4,22

    8,5

    2,98

    Zonisamide

    8,29 mg/L

    5,74

    1,24

    2,14

    Rufinamide

    8,93 mg/L

    5,84

    1,56

    7,16

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