1. J. Elek , A. Simon, J. Török, A. Csontos, Cs. Ballai
APPLICATION AND VALIDATION OF MID-RANGE INFRARED SPECTROSCOPY IN GLP ENVIRONMENT
J. Elek , A. Simon, J. Török, A. Csontos, Cs. Ballai
WSC10, Samara

2. Typical analytical workflow
method development
validation (EP, USP, ICH, etc.)
analysis GLP, GMP
documentation, archivation

3. Purpose of this presentation
Why this method was made:
for a certain analysis
Why it was presented in Hungary:
analists do not trust multivariate methods
Playing around with statistics
Faliure of instrument distributors „one button” solutions
Dominance of HPLC and GC
„counterstrike”
Why it was presented Here:
„János, you do not know much about chemometrics, but you play the guitar a nice way, so you should come back for the next meeting too.” Beata Walczak, 2012, Drakino

4. The infrared spectroscopy
Infrared domain: 780 nm –1000 μm, (10 cm-1– cm-1)
Discovery: Early 1800’s (Sir William Herschel)
Identification of the first absorption bands:
Systematic examinations, Coblentz 1903 (2-3 hours per spectrum)
The golden age: structure elucidation
Renaissance: multivariate analysis(discriminant analysis, quanification, PCA, LDA, PLS, etc.)

5. Ethanol IR spectrum

6. Ethanol NIR spectrum

7. What is the goal?
regression
discrimination

8. Analitical nightmare
Quantitative determination of an oily, high viscosity, mulit-component test item from arachis oil.
Sensitive to hydrolysis
Non volatile
Normal phase chromatography???
ATR FT-IR spectroscopy
Unscrambler X

9. ATR-technique Versatile Fast No sample preparation
Easy to clean (cross-contamiation)

10. Absorbtion IR spectrum of the test item

11. TI vs peanut oil

12. Calibration series

13. Calibration series 1st derivative

14. PLS-DA plot

15. Calibration line (validated)

16. Validation parameters
Repeatability
ccal (mg/g)
L4_001
5.3018
L4_002
5.2958
L4_003
5.1785
L4_004
5.3107
L4_005
5.2403
L4_006
5.2768
L4_007
5.2274
RSD%
0.9
Target C (mg/mL)
cNom (mg/g)
ccalc (mg/g)
Recovery (%)
RSD% 5
5.75
5,68
98.8
1.6
5.62
5.54
98.6
1.3
5.61
5.33
94.9
2.8
100
106.20
106.43
100.2
0.4
106.75
106.84
100.1
0.1
106.71
106.77
Date R2
RMSECV
Deviation of the individual calibration points %
clow
chigh
Feb 19
1.00
0.07
0.23
Feb 20.
0.04
0.24
Feb 24.
0.08
0.50
March 5
0.05
0.67

17. Achilles-heel: specificity
During method development in 2.5 mg/ml solution

18. Regression coefficients (no derivation)

19. Weighted regresion coefficients

20. Achilles-heel: specificity

21. Summary
The quantitative MIR analysis can replace the frequently used chromatographic techniques in most cases
Minor need for sample preparation
Fast, cheap
With careful use of chemomertic techniqes the methods can be validated according to the mainly applied guidlines (ICH, EP, USP, etc.)
Until these algorithms run in R and Matlab it will always be a very hard work to convince the „simple” analysts (and the authorities) about the adequacy of their use.