Page 84 - GTM-2-3

Global Translational Medicine HPLC-UV lacosamide quantification

action, which justifies its efficacy, involves the modulation method for quantifying lacosamide in human plasma,
of slow sodium channels . Lacosamide is indicated for use intended for therapeutic monitoring purposes.
[1]
in adults, adolescents, and children. Its pharmacokinetics
include excellent bioavailability, a linear pharmacokinetic 2. Materials and methods
profile, and low plasma protein binding, with 60% hepatic 2.1. Critical reagents and solvents
[2]
elimination and 40% renal elimination .
Lacosamide standard (generously donated by local industry
While some authors have questioned the utility of [origin: Sigma-Aldrich, Ref. No. SML3059, purity >98%,
therapeutic drug monitoring in the general population, Saint Luis, Missouri, USA]); propranolol hydrochloride
citing the perceived predictability of plasma concentration in standard (used as an internal standard, purchased from
relation to the dose [3,4] , others have documented the benefits INAME, Control No. 190031, purity 100.2%, Buenos
of therapeutic drug monitoring. They argue for the necessity Aires, Argentina); HPLC-grade acetonitrile, HPLC-
of measuring anticonvulsant concentrations to characterize grade methanol, analytical-grade hydrochloric acid, and
individual pharmacokinetics and optimize patient drug ethyl acetate (SINTORGAN, Villa Martelli, Buenos Aires
regimens [5,6] . Preclinical studies have indicated the potential Argentina); MilliQ water; and monobasic potassium
for plasma level variations if lacosamide clearance is phosphate salt (CICARELLI, Buenos Aires, Argentina).
affected . Clinical evidence also supports the need for
[7]
dose adjustments in patients co-administered with enzyme 2.2. Equipment
inducers or those experiencing impaired hepatic or renal Analytical balance (METTLER TOLEDO, model MS
function due to critical illnesses . Furthermore, patients 105), vortex mixer (Scientific Industries, Genie2 G560),
[8]
with dynamic physiology (the elderly, children, and pregnant magnetic stirrer (Thermo Scientific Cimarec Basic model
women) present a challenge in establishing a single reference S194615), ultrasonic bath (TestLab, model TB-010 TA),
range, which remains a topic of debate in these populations . thermostatic bath (FAC, model Ballus), vacuum filtration
[9]
Some of the reported analytical methods for the system (Millipore), vacuum pump (Pascal, model PC-75),
determination of lacosamide include ultraviolet spectroscopy water purifier (Simplicity Millipore), centrifuge (IEC
(UVS), high-performance liquid chromatography Centra-7R), -80°C freezer (Revco), pH meter (COLE
(HPLC) with UV detection (HPLC-UV) , thin-layer PARMER, model 05669-29), and air compressor (Total
[10]
chromatography (HPTLC), and HPLC-mass spectrometry . Tools, industrial TTAC2506).
[11]
However, the existing methods for the determination of
lacosamide in human plasma exhibit various limitations, 2.3. Chromatographic system
especially in low-resource settings where affordable The analysis was performed using a THERMO
analytical methods are a key factor for obtaining versatile SCIENTIFIC ULTIMATE 3000 HPLC system equipped
and reliable results. Challenges associated with these with an autoinjector, column oven, and a diode array UV
methods include the absence of an internal standard [12,13] , detector. A Phenomenex Kinetex C18, 100 A column with
the necessity for more costly or unavailable reagents due to dimensions of 150 mm × 4.6 mm and a particle diameter
supply shortages, and the potential inefficiency of proposed of 2.6 µm was selected as the stationary phase. The mobile
sample treatments. The absence of an internal standard can phase (MP) consisted of a 70:30 v/v ratio of 10 mM
affect the accuracy and precision of measurements, as well potassium phosphate buffer (KH PO ) and acetonitrile,
2
4
as the reproducibility of the results. The analyte recovery pH adjusted to 3.5 using hydrochloric acid (HCl). An
from the matrix is a significant source of variation in the isocratic elution method was employed with a flow rate of
analytical process; thus, it is critical for the success of the 0.6 ml/min, an injection volume of 20 µL, and detection at
method and requires an adequate sample treatment. An a wavelength of 210 nm. Each sample had a total elution
unsatisfactory analyte recovery can cause a systematic time of 10 min.
error in the method, which affects not only accuracy and
precision but also sensitivity and selectivity. Furthermore, 2.4. Method optimization
obtaining inaccurate or imprecise analytical results may The optimized conditions encompassed the following
have implications for therapeutic drug monitoring, drug parameters: The choice of the bonded phase (C8, C18), the
dosing recommendations, and patient safety. composition of the mobile phase (organic solvent type and
These challenges prompted the development of an internal percentage, buffer type and pH), operational parameters such
method with a focus on regional accessibility. This work as flow rate, temperature, and wavelength, the nature of the
presents the development, optimization, and validation of a internal standard, and the method for preparing the biological
simple, fast, cost-effective, reliable, and efficient HPLC-UV sample (either precipitation or liquid-liquid extraction).

Volume 2 Issue 3 (2023) 2 https://doi.org/10.36922/gtm.1265