low quality predictions (or measurements) can lead to ...€¦ · low quality predictions (or...
TRANSCRIPT
Low Quality Predictions (or Measurements) Can Lead to
Erroneous Decisions
John ComerTechnical Director
Sirius Analytical Ltd.
Talk given at ACD Users Meeting, Obernai, France, 25 October 2007
/412© 2007
Who are Sirius?
UK-based instrument company We make instruments for measuring physicochemical properties– pKa
– logP– Solubility
We analyse samples sent by customersWe support these activities with scientific research– Over 50 publications– Many new methods introduced
We specialise in accurate measurement
www.sirius-analytical.com
/413© 2007
Sirius and ACD
We have a copy of ACD ChemSketch v 4.55, May 2000We use it for drawing chemical structuresDatabase is useful for structures of drugs, and referencesWe use it for predicting pKa, logP and solubility, before designing experiments to measure them– How many pKas?– Which are acids, which are bases?– Approximate values?– Predicted logP often correlates with solubility, so helps us decide
whether to use cosolvent for pKa measurement
We published a paper citing our use of ACD software [1]Sirius RefinementPro software produces a file of measured data ready to be imported into ACD software
[1] Box, K. Bevan, C. Comer, J. Hill, A. Allen, R. Reynolds, D. High-throughput measurement of pKa values in a mixed-buffer linear pH gradient system. Anal. Chem. 2003 (75(4)) pp 883-892
/414© 2007
Themes covered by this talk
Variations in published solubility valuesSolubility definitionsReasons for incorrectly reported solubility valuesKinetic vs. equilibrium solubilityLoperamide – an example of an incorrect resultLogP vs. logS0, and predicting BCS classSolubility vs. pH, and the validity of Henderson-Hasselbalch
From next Monday, you will be able to download these slides from
http://www.sirius-analytical.com/downloads/Downloads/downloadsacd.htm
/415© 2007
Acknowledgements
Colleagues at Sirius– Karl Box, Tom Gravestock, Jon Mole, Ruth Allen (Chemistry)– Martin Stuart (Software)
Academic collaborators– Krisztina Takács-Novák, Edit Baka (Semmelweis University, Budapest)– Farah Huque (University of Cardiff)– Rebeca Ruiz (University of Barcelona)– Slavica Eric (University of Belgrade)
Not forgetting…– Tony Williams (ACD)
From next Monday, you will be able to download these slides from
http://www.sirius-analytical.com/downloads/Downloads/downloadsacd.htm
/416© 2007
How to measure solubility by shake-flask
Weigh sample, add buffer, stir for a long time (e.g. 24 hours or more) to create saturated solution
Sample must be present in excess,
i.e. precipitate always there
Separate solution and
solid, by
Sedimentation, Centrifugation or
Filtration
Check crystalline form
(polymorph) of solid
Analyse concentration in solution
Concentration = equilibrium solubility
Don’t forget to check pH of solution!
This information provides solubility at a
measured pH
/417© 2007
Variation of aqueous solubility in literature (25ºC)
8 A. Avdeef, C. M. Berger, Pharm. Res. 17 (2000), pp. 85-89.11 S. H. Yalkowsky, Y .He, Handbook of Aqueous Solubility Results , CRC Press, Boca Raton, (2003)12 T. Higuchi, F. M. L. Shih, T. Kimura, J. H. Rytting, J. Pharm. Sci . 68 (1979), pp. 1267-1272 13 A. Fini, G. Fazio, G. Feroci, Int. J. Pharm . 126 (1995), pp. 95-10214 S. Pinsuwan, A. Li, S. Yalkowsky, J. Chem. Eng. Data 40 (1995), pp.623-62615 S. Pinsuwan, P. B. Myrdal, Y. C. Lee, S. H. Yalkowsky, Chemosphere 35(11) (1997), pp. 2503-251316 C. A. S. Bergström, K. Luthman, P. Artursson, Eur. J. Pharm. Sci. 22 (2004), pp387-39817 R. H. Levy, M. Rowland, J. Pharm. Sci. 60 (1971), pp. 1155-115918 P. Ruelle, U. W. Kesserling, J. Pharm. Sci . 87(8) (1998), pp. 998-101419 M. F. Powell, In Analytical Profiles of Drug Substances, K. Florey, Ed. Academic Press: San Diego, 1986, Vol. 29 K. Kawakami, K. Miyoshi, Y. Ida, Pharm. Res . 22 (2005), pp. 1537-154330 D. K. Madan, D. E. Cadwallader, J. Pharm. Sci. 62 (1973) pp.1567
Table from: Baka, E. Comer, J E A. Takács-Novák, K. Study of equilibrium solubility measurement by saturation shake-flask method using hydrochlorothiazide as model compound, J. Pharm. Biomed. Anal., 2007 (submitted)
CompoundSolubility range (mg/ml)
Solubility range (µg/ml)
Cholesterol [12, 30 ] 0.025, 2600Dexamethasone [11, 29] 89.1, 121.0Diclofenac [8. 12. 13] 0.6 - 2.4Digoxin [11,29] 28.0, 97.9Estradiol [11,29] 0.16, 5.00Hydrocortisone [11, 29] 280, 359Ibuprofen [8, 13, 14, 15] 20 - 80Indomethacin [11,19] 4.00, 14.0Lidocaine [16, 17, 18, 19] 2.30 - 4.60Progesterone [11, 29] 7.90, 200Riboflavine [11, 29] 66.0, 99.9
/418© 2007
Uncertainty of experimental data
Anthracene0.86 log units [2] 17 laboratories
NOH
OH
CH3
CH3 CH3
TerfenadinelogS = -6.17[6], -4.67[5,8], -7.74[7]
[1] Jorgensen, W L. Duffy, E M. Adv. Drug Deliv. Rev. 2002, 54, 355-366[2] Kishi, H. Hashimoto, Y. Chemosphere 18 (1989) 1749-1759][3] Pogliani, L. J. Chem. Inf. Comput. Sci. 1996, 36, 1082-1091[4] Dannenfelser, R M. Yalkowsky, S H. Sci. Total. Environ. 1991, 109-110, 625-628 [5] Analytical Profiles of Drug Substances[6] Bergström, C.A. Wassvik, U.N. Luthman, K. Artursson, P. J. Chem. Inf. Comput. Sci. 44 (2004) 1477-1488 [7] Sirius Analytical Ltd. – measured in-house[8] Rytting, E. Lentz, K. A. Chen, X –Q. Qian, F. Venkatesh, S. The AAPS Journal 7 (2005) E78-E105[9] Dollery, C. Therapeutic Drugs, 2nd edition 1998, Churchill Livingstone, London
N
NHN
NH
O
NH2
GuaninelogS = -3.58 [3,8], -1.86[4]
“The average uncertainty in experimental logS measurements for reasonably complex organic molecules is likely no better than 0.6 log unit” [1]
N
O
NO
Cl
LoperamidelogS= -7.13 [7], -4.38 [9]
/419© 2007
Software for prediction of solubility
% Compounds predicted within
0.42 0.4062.338.5AlogP98
0.61 0.6072.937.7Cerius2ADME0.51 0.4967.241.0WSKOWWIN0.42 0.39 66.434.4ADMEWORKS Pred.
0.53 0.5174.644.3Absolv 2
0.73 0.7273.142.9SPARC
0.35 0.3445.723.3CHEMICALC
0.57 0.5773.847.6QikProp
0.44 0.4277.962.3MOLPRO0.60 0.5978.754.9ESOL0.48 0.4681.146.7PredictionBase0.67 0.6681.151.6AlogS0.73 0.7285.259.0ACDLabs0.67 0.6586.059.8ChemSilico0.74 0.7386.959.0ADME Boxes0.76 0.7486.972.1Admensa0.82 0.8291.064.8SimulationPlus
R2 q2±1.0 log unit±0.5 log unitModel
Predictive abilities of some software for aqueous solubility prediction
Dearden’s study [1]Test set: 122 compounds [2]
“Prediction of aqueous solubility can be almost as good as
experimental measurements for most of the compounds”
[1] Dearden JC Expert Opin. Drug Discov. (2006) 31
[2] Rytting, E. Lentz, K. A. Chen, X –Q. Qian, F. Venkatesh, S. The AAPS Journal 7 (2005) E78
/4110© 2007
Rytting’s published solubility values
Rytting et al published two tables of measured solubilityTable 1 lists experimental logS of 321 drugs from the literatureTable 2 lists 122 logS values measured in Rytting’s lab. All samples are also in table 1, with slightly different resultsSearching through solubility results measured at Sirius, we found we had measured 25 of the Rytting compounds, using CheqSol methodology
Compound Sirius LogS
Rytting 321
Rytting 122
Amitriptyline -4.39 -4.46Chlorzoxazone -2.61 -2.83 -2.87Diclofenac -5.45 -5.10Flufenamic Acid -5.35 -4.62 -4.41Flurbiprofen -4.11 -3.74 -3.87Folic Acid -5.31 -5.44 -4.89Haloperidol -5.47 -4.43 -4.98Hydrochlorothiazide -2.68 -2.69 -2.78Ibuprofen -3.61 -3.42 -3.63Lidocaine -1.85 -1.77Mefanamic Acid -6.34 -3.77 -5.18Metoclopramide -3.59 -3.18Nadolol -1.57 -1.01 -1.01Naproxen -4.14 -4.16 -3.79Nitrofurantoin -3.33 -3.48 -3.49Norfloxacin -2.75 -3.06 -2.76Phenobarbital -2.28 -2.37Phenylbutazone -4.39 -2.64 -4.13Prochlorperazine -4.87 -4.40Propranolol -3.50 -0.71Quinine -2.81 -2.79 -2.43Sulfamerazine -3.10 -1.22 -3.11Sulfathiazole -2.70 -2.81 -2.18Sulindac -4.52 -5.00Terfenadine -7.74 -4.67 -4.87
/4111© 2007
y = 0.99x + 0.01R2 = 0.94
-8
-6
-4
-2
0
-8 -6 -4 -2 0LogS Sirius
LogS
Ryt
ting
Rytting 321 compounds
Linear (Rytting 321 compounds)
18 of Rytting’s Table 1 results compared with Sirius
Sirius results compare really well with 18 results from Rytting table 1, which were all values taken from the literature
/4112© 2007
18 Rytting Table 1 results plus 7 outliers
These outliers from Rytting Table 1 are all much higher than Sirius measured values
y = 0.99x + 0.01R2 = 0.94
-8
-6
-4
-2
0
-8 -6 -4 -2 0LogS Sirius
LogS
Ryt
ting
Rytting 321 compounds
Rytting 321 outliers
Linear (Rytting 321 compounds)
/4113© 2007
17 of Rytting’s Table 2 results compared with Sirius
These 17 results were measured by Rytting’s group using at least 24 hours shaking followed by 15 minutes centrifugation at 14,000 rpmSupernatants analysed by HPLC with UV detectionSome outliers high compared with Sirius measured values
y = 0.95x - 0.05R2 = 0.93
-8
-6
-4
-2
0
-8 -6 -4 -2 0LogS Sirius
LogS
Ryt
ting
Rytting 122
Rytting 122 outliers
Linear (Rytting 122)
/4114© 2007
Rytting tables 1 and 2 vs. Sirius
Who’s right – Sirius or the published values?Before trying to answer that question, let’s look at some definitions
y = 0.99x + 0.01R2 = 0.94
-8
-6
-4
-2
0
-8 -6 -4 -2 0LogS Sirius
LogS
Ryt
ting
Rytting 321 compounds
Rytting 321 outliers
Rytting 122 compounds
Linear (Rytting 321 compounds)
Terfenadine
Mefan-amic acid
PhenylbutazonePropranolol
Sulfamerazine
Haloperidol
Flufenamic acid
/4115© 2007
What does the word “solubility” mean?Thousands of references in PubMed, June 2007
To be sure of what we are measuring, we drafted our own definitions
Solubility 63,949Biological solubility 8,135Native solubility 1,813Water solubility 1,795Natural solubility 1,736Aqueous solubility 1,177Buffered solubility 397Equilibrium solubility 105Intrinsic solubility 62Turbidimetric solubility 35Thermodynamic solubility 30Unbuffered solubility 21Kinetic solubility 9
Dissolve excess sample in water, no buffer. Sample controls the solution pH.
How many people actually measure this pH?
/4116© 2007
Solubility definitionsKinetic (or Turbidimetric) Solubility is the concentration of a compound in solution at the time when an induced precipitate first appears
– This measurement is popular at early stage assessment– The result obtained is often significantly higher than the equilibrium solubility
Equilibrium (or Thermodynamic) Solubility is the concentration of compound in a saturated solution when excess solid is present, and solution and solid are at equilibrium– This is the true solubility of a molecule– It is traditionally measured by shake-flask, which is accurate but slow
Nowadays, high throughput shake-flask measurements are made in parallel– Other novel techniques have been developed e.g. Sirius CheqSol
Intrinsic Solubility S0 is the equilibrium solubility of the free acid or base form of an ionizable compound at a pH where it is fully un-ionized
Intrinsic solubility must be used for training software to predict solubility of ionisable drugs
/4117© 2007
Reasons for incorrectly reported intrinsic solubility
Shake-flask problemsResult reported was not measured at pH where sample was unionisedPrecipitate was present, but it was a poorly soluble salt, not the unionised sampleIncorrect separation, e.g. sample passed through filterNot waiting long enough for equilibrium
Other problemsNatural solubility result was reportedKinetic solubility result was reportedResult measured in a solvent was reportedSolubility reported for a different polymorph
/4118© 2007
Kinetic solubility and Equilibrium solubilitySome compounds can’t form super-saturated solutions– When the pH is right, they fall out of solution immediately– Their kinetic and equilibrium solubilities are identical
Other compounds form super-saturated solutions– Their kinetic solubilities are higher than their equilibrium solubilities
0
1000
2000
3000
4000
5000
6000
7000
8000
9000
0 2000 4000 6000 8000
Equilibrium solubility, µg/mL
Kin
etic
sol
ubili
ty, µ
g/m
L
ChasersNon-chasersLinear (Non-chasers)
0
200
400
600
800
1000
1200
1400
1600
0 200 400 600 800 1000 1200 1400 1600
Chasers
Non-chasers
/4119© 2007
Salt solubility reported, not intrinsic solubility
Monoprotic basepKa values used– 8.90 (measured at Sirius)
N
O
NO
Cl
Loperamide has been reported in the literature to have higher solubility than we found– Reported: LogS0 = -4.38 [1], equivalent to 41.7µM– Measured at Sirius: LogS0 = -7.13, equivalent to 0.073µM
[1] Dollery, C. Therapeutic Drugs, 2nd edition 1998, Churchill Livingstone, London
Loperamide
/4120© 2007
Loperamide pKa measurement
R2 = 1.00
pH-metric measurementSix titrations in water-methanol (14-55%)Result 8.90 by YS extrapolationpsKa values at methanol percentages used in CheqSol experiments are interpolated from YS graph
/4121© 2007
Loperamide solubility exampleSolubility measured in water-methanol mixtures (6 titrations, 20-55% methanol)
This example shows experiment done in 34.1% methanol
Interpolated psKa of 8.46
Top graph – chasing equilibrium
Bottom graph – Bjerrum plot, showing Precipitation Point and CheqPoint.
Sample is a Chaser, because its kinetic solubility (calculated from the precipitation point) is higher than its intrinsic solubility (calculated from the CheqPoint)
/4122© 2007
Loperamide solubility extrapolation
Intrinsic solubility– Extrapolated LogS = -7.13– Equivalent to 0.073µM
Natural solubility and pH– 0.83µM, pH 7.67,
determined by mass balance calculation
Solubility at pH 7.4– 2.39µM, interpolated from
solubility-pH profile
Salt solubility – see next slide
R2 = 0.99
Intrinsic solubility, S0
Solubility at pH 7.4
Natural solubility
Salt solubility
/4123© 2007
Loperamide verdict
Loperamide is extremely insoluble in free base formIt’s also poorly soluble in salt formWe measured solubility of several loperamide salts
Solubility Profile of Loperam ide with Various Counter-Ions
-8
-7
-6
-5
-4
-3
-2
-1
0
2 4 6 8 10 12
pH
LogS
0.1M mesylate
0.15M mesylate
0.025M HCl
0.075M HCl and0.025M Besylate0.15M HCl
0.05M Besylate
0.07M Benzoate
0.15M Besylate
0.15M Benzoate
0.15M Tosylate
Loperamide solubility reported in the literature – LogS of -4.38, equivalent to 41.7µM
We believe that the reported solubility is for a salt form of loperamide
/4124© 2007
What can we learn from logP vs. logS0?We measured PAMPA permeability and logP of 39 drugsAll marketed compoundsResults show that PAMPA permeability is proportional to logP [1]– Diverse set of 39 drug-like
compounds– LogP alkane-water measured
pH-metrically using Sirius GLpKa
[1] K. Box, J. Comer, F. Huque, Correlations between PAMPA permeability and log P. In Pharmacokinetic profiling in drug research: biological, physicochemical and computational strategies. Wunderli, H. Kraemer, S. Folkers, G. Testa, B., editors, Verlag Helvetica Chimica Acta, 2006, pp 243-257
/4125© 2007
-10
-8
-6
-4
-2
0
2
-2 0 2 4 6 8 10
Log P
Log
S 0
Melting point above 140ºC. Compounds can all form supersaturated solutions
Melting point below 140ºC. Can supersaturate
Melting point below 140ºC. Cannot supersaturate. Most have CNS activity
For points between grey lines: LogS0 = -1.03 LogP – 0.53
High solubility region
Low solubility region
Measured LogP vs. measured logS0 for 84 drugs
Pramoxine
/4126© 2007
-10
-8
-6
-4
-2
0
2
-2 0 2 4 6 8 10
Log P
Log
S
12
34
CLASS 1 CLASS 3
CLASS 2 CLASS 4
More permeable (higher logP)
Mor
e so
lubl
e
Comparing results with Biopharmaceutics Classification (BCS)
No BCS data for 5 compounds marked yellow.
/4127© 2007
-10
-8
-6
-4
-2
0
2
-2 0 2 4 6 8 10LogP
LogS
Measured values from previous slides
/4128© 2007
-10
-8
-6
-4
-2
0
2
-2 0 2 4 6 8 10LogP
LogS
Predicted values calculated using a popular program (not ACD!)
Tendency for solubility values to be calculated too high
Terfenadine
/4129© 2007
-10
-8
-6
-4
-2
0
2
-2 0 2 4 6 8 10LogP
LogS
Folic acid
Loperamide
Am
anta
dine
Dilti
azem
Piroxicam
Glipizide Meclofenamic acid
Mefenamic acidWar
farin
Probe
necid
Predicted properties can differ significantly from measured values
Biggest errors are in predicted solubility
Errors lead to mis-prediction of BCS class
MeasuredPredicted
/4130© 2007
-10
-8
-6
-4
-2
0
2
-2 0 2 4 6 8 10LogP
LogS
Amantadine
Diltiazem
1
2
111333222444
High permeability
Hig
h so
lubi
lity
Measured class 1 – predicted class 2
Measured■ Predicted
/4131© 2007
Measured 2 and 4 – predicted 1 and 3
-10
-8
-6
-4
-2
0
2
-2 0 2 4 6 8 10LogP
LogS
Folic acid
Loperamide
Piroxicam
GlipizideMeclofenamic acid
Mefenamic acidW
arfa
rinPr
oben
ecid
1
2
3
4 Measured■ Predicted
111333222444
High permeability
Hig
h so
lubi
lity
/4132© 2007
Solubility vs. pH for promethazine, a weak base with one pKa– weak acid would be mirror-image
LogS = the concentration of unionized sample in solution in equilibrium with excess undissolved sampleGraph based on Henderson-Hasselbalch principles, drawn using pKa and intrinsic solubility (S0)
[H][H])(KSS a0 +
=
logS
pH (Concentration scale)
-4
-3
-2
-1
0
1
4 6 8 10 12
Intrinsic solubility
pKa
Solubility and pH for ionisable molecules
a
a0
K[H])(KSS +
=
acid base
If the sample has more than one pKa, or it forms an insoluble salt, the equations are more complicated
N
S
NCH3CH3
CH3
/4133© 2007
Henderson Hasselbalch vs. Shake-flask
Green lines are calculated from pKaand CheqSol S0 values measured at SiriusBlue points are shake-flask data [1]Agreement is good where samples are unionizedWhy does diclofenac solubility deviate from the green line at higher solubilities?
Propranolol
Diclofenac
[1] Shake flask data in the next four slides is from Avdeef, A. Berger, C M. Brownell, C. Pharm. Res. 2000, 17(1), 85-89
BasepKa = 9.54S0 = 81µg/mL (314µM)
O
NH
OH
CH3
CH3
NH
OH
O Cl
Cl
AcidpKa = 3.99
S0 = 0.9µg/mL (4.1µM)
/4134© 2007
Deviations from solubility-pH profile
The graph may deviate at higher logS values for two possible reasons:– Either the ionized sample may
precipitate as a salt with a counter-ion
– Or all the compound used in the experiment may dissolve (“Solubility Limit”)
logS
pH
-5
-4
-3
-2
-1
0 2 4 6 8
Either: poorly soluble salt
or:all sample dissolved
/4135© 2007
Reaching the solubility limit
9.4 g/litre
29.6 g/litre
296 g/litre
Although it was not confirmed in the publication, it is likely that the solubility limit was reached in this example The concentration at the red points is shownIt is impractical to do shake-flask solubility experiments at very high concentrations, especially for new chemical entities available only in small quantities
Diclofenac
0.00075 g/litre
/4136© 2007
Solubility vs. pH for four drugsKetoprofen(acid, one pKa)
O
O
OH
CH3
800 g/litre
Hydrochlorothiazide (acid, two pKas)
94.2 g/litre
SSNH
O O
NH
O
O
NH2
Cl
146 g/litre
Ibuprofen(acid, one pKa)
O
OH
CH3
CH3
CH3
Famotidine (ampholyte, two pKas)
7.4 g/litre
/4137© 2007
Chlorprothixene
0.0000511M
Promethazine
0.0000499M
Pramoxine
0.0000485M
Does Henderson-Hasselbalch always draw true solubility-pH profiles?
A recent publication [1] suggested that Henderson-Hasselbalch did not always predict solubility-pH profiles accurately.
We re-measured properties of some molecules and found that the published profiles may have been based on questionable values for pKa or solubility
The next slides show results for chlorprothixene, promethazine and pramoxine
Shake flask solubilities were measured by HPLC with UV. Note the low UV signal for pramoxine
[1] Bergström, C A S. Luthman, K. Artursson, P. Accuracy of calculated pH-dependent aqueous drug solubility, Eur. J. Pharm. Sci., 2004, 22, 387-398
/4138© 2007
New profile (Sirius 2007): pKa = 9.52, LogS = -0.76
Published profile:pKa = 8.4 [1], logS = 0.2
Solubility of benzenesulfonate salt, (Sirius, 2007)LogS = 2.26
pH 12
No. of buffers: 10
[1] Encyclopedia of Therapeutic Drugs, Churchill Livingstone, New York, 1991
-1
0
1
2
3
4
0.0 2.5 5.0 7.5 10.0 12.5pH
LogS
(
S =
[B] +
[BH
+]).
Verdict: Sirius red profile fits all shake-flask points except at pH 12. It’s a coincidence that the published profile fits the mid-pH points, as the pKa and solubility are both questionable
Published black line = “best fit”
/4139© 2007
0
1
2
3
4
5
0.0 2.5 5.0 7.5 10.0 12.5pH
LogS
(
S =
[B] +
[BH
+]).
New profile (Sirius 2007) pKa = 8.99, LogS = 1.81
Published profile:pKa = 9.1 [1], logS = 0.3
pH 12.07
No. of buffers: 11
Verdict: shake-flask data fits Sirius profile much better than published profile in mid-pH region
[1] Encyclopedia of Therapeutic Drugs, Churchill Livingstone, New York, 1991
/4140© 2007
2
3
4
5
6
7
0.0 2.5 5.0 7.5 10.0 12.5pH
LogS
(
S =
[B] +
[BH
+]).
New profile (Sirius 2007) pKa = 7.12, LogS = 2.98
Published profile:pKa = 6.2 [1], logS = 2.5
Verdict: Nothing fits well. Different pKas used. Pramoxine is a liquid at high pH. UV absorbance is low. Could these properties make it hard to measure by shake-flask?
pH 12.08
No. of buffers: 9
[1] Casanovas, A M. Labat, C. Courriere, P. Oustrin, J. 1982, Eur. J. Med. Chem. 17, 333-337
/4141© 2007
Conclusions
There may have been shortcomings in experimental techniques used in the past to measure PhysChem propertiesIt can be risky to use published values without checking themIt’s useful when customers can improve predictions by making use of good quality results to train softwarePeople must work harder to get good experimental resultsThe danger: Too much automation, Not enough thinkingGood experimental design is critical for getting the right resultACD software helps us to design good experiments
Sirius – Instruments and PhysChem Analytical Services
Don’t forget! Download these slides from
http://www.sirius-analytical.com/downloads/Downloads/downloadsacd.htm