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Optimization of parameters for the gas chromatographic determination of polycyclic aromatic hydrocarbons
67
Analytrco Chlmlca Acta, 268 (1992) 67-71 Elsevler Science Publishers B V . Amsterdam
Optimization of parameters for the gas chromatographic determination of polycyclic aromatic hydrocarbons W R Trevehn, L H Vldal, M D Landgraf, I C E Srlva and M 0 0 Rezende Unwerszdade de Silo Paula, Instztuto de Fistca e Quimcca de Srio Carlos, Departamento de Quimrca e Fika Molecular, Au Dr Carlos Botelho 1465, Cx P 369, 13560 SZo Carlos SP (Brazd) (Received 5th February 1992, revised manuscnpt received 2nd Aprd 1992)
Ahstraet
The optmuzatlon of condltlons for the gas chromatographlc determination of sixteen polycychc aromatic hydrocarbons (PAHs) using the sphtless mJectlon mode 1sdescribed Parameters optlmlzed were the type and amount of solvent, the temperature of the mnjtion port and the column and the purge time A factonal design was used uutlally to ldentlfy the most important parameters Based on this mttlal study, xylene was chosen as the solvent and 100°C was chosen as the best value for temperature of the column The other parameters were optmuzed by using a sunplex method The optlmum condltlons found were 3 ~1 of solvent, 280°C for the mJectlon port and a purge time of 30 s With these condltlons, relatwe standard devlatlons for five rephcate determmattons of 2-ng amounts of PAHs were 3 2-10 8% Keywords Gas chromatography,
Optmuzatlon methods, Factonal design, Polycychc aromatic hydrocarbons, Simplex
Polycychc aromatic hydrocarbons (PAHs) represent a class of organic compounds identified as “unambiguous priority pollutants” by the US Envlronmental Protection Agency (EPA) [l] This class of compounds has been shown to contam several potent carcmogens [2] The occurrence and potential hazards of these compounds have stimulated research related to the determmatlon of PAHs m the environment Gwen the low concentrations of these species, it 1s important to use condltlons that optimize the separation efficiency and detection hmlts m envrronmental samples Grob and Grob [3,4] ldentlfled the solvent effect as one of the most lmportant factors for alkanes such as heptane, octane Correspondence to M 0 0 Rezende, Umversldade de Slo Paulo, Instltuto de Fislca e Quinuca de SCo Carlos, Departamento de Quinnca e Fiilca Molecular, Av Dr Carlos Botelho 1465, CxP 369, 13560 Sb Carlos SP (Brazil)
and nonane determined using gas chromatography (GC) with the sphtless mJectlon mode Cold trapping was orlgmally described as the mechamsm operating m the sphtless mJectlon mode Subsequent studies [4,5] showed that the solvent effect 1s important for both sphtless and on-column mjectlon Based on Grob and Grob’s studles, Jennmgs et al [6] described the theoretical basis of the solvent effect Cold trapping and the solvent effect were described as the two functions that led to the reconcentration of elutes on the top of the column The factors affecting splitless mjection are numerous, e g , inJection speed, liner size, matnx composition, presence of a liner plug [7], column temperature, volatlhty of the solvent, amount of solvent and tune of vaporlzatlon [8] Umvarlate studies, although useful, are seldom completely satisfactory for complex processes such as m GC that depend on multiple variables The
0003-2670/92/$05 00 0 1992 - Elsevler Science Pubhshers B V All nghts reserved
68
WR Trevelm et al /And
best condltlons are usually obtamed by multlvanate studies such as factorial design and simplex methods such as the sequential simplex method introduced by Spendley et al [9] and improved by Nelder and Mead [lo] In this study, a factorial design was used to identify the most important variables m GC to increase the sensltlvlty of sixteen PAHs and a simplex method was used to identify the optimum values for the most nnportant parameters, namely the volume of solvent, the temperature of the mjectlon port and the purge time
EXPERIMENTAL
Reagents and standards
All solvents were glass distilled through a 150cm column The compounds studied were naphthalene, acenaphthene, acenaphthylene, fluorene, phenanthrene, anthracene, fluoranthene, pyrene, bendalanthracene, chrysene, benzo[ blfluoranthene, benzo[ klfluoranthene, benzo[ alpyrene, dlbenzo[a,h]anthracene, benzo[ ghzlperylene and mdeno[l,2,3_cdlpyrene (Supelco, Bellefonte, PA) A stock solution was prepared m a mixed benzene-methylene chrorlde (1 + 1, v/v> solvent contamg a nommal concentration of 2 mg ml-’ of each PAH Further dllutlons were made with benzene and xylene to give a fmal concentration of 2 pg ml-’ of each PAH Ahquots of the stock solution were measured using a 100~~1 Type 701 RN syringe (Hamilton, Reno, NV) Chromatographu:
condltwm
All experiments were done usmg a Model 5890 high-resolution gas chromatograph (Hewlett Packard, Palo Alto, CA) equipped with a flame lomzatlon detector (FID) The output of the FID was connected to an integrator (HP-33931 A fused-s&a capillary column coated with crosslmked Ultra-l methyl&cone (Hewlett-Packard) as the stationary phase (25 m x 0 32 mm 1 d , flhn thickness 0 17 pm> was used with splitless mjectlon Some expenmental conditions, mcludmg solvent, temperature of the mJectlon port, initial temperature of the column, amount of solvent
Chun Acta 268 (1992) 67-71
TABLE 1 Parameters used m factorial design Variable
Maxunum level ( + )
Mnumum level (-)
S = solvent P = time of purging V = amount of solvent T = uutlal column temperature I = injectIon port temperature
Benzene 05mm 10 /Ll 100°C 270°C
Xylene 1Omm 3O/.Ll 12Wc 290°C
and purge tnne, were varied during the process Other parameters, mcludmg flow-rate of the carrier gas (40 cm s-l), heating rate (5°C mm-‘) and fmal temperature (280’0, were the same m all experiments Peak areas for all PAHs were added and normahzed for each chromatogram Factonal design
Values of the parameters used m the factorial design are listed m Table 1 The combmatlons of these mmunum and maximum levels are shown m Table 2
TABLE 2 Combmatlons among the mmnnum and maximum levels of the parameters listed m Table 1 Experiment No E, E2 E3 E4 Es E6 E7 E8 E, EIO ‘%l
E 12 E13
E 14 E 1.5 El6
Variable S +
P
V
T
I
-
-
+
-
+
+
-
+
+
+ -
+ +
+ + + -
+ + + -
+ -
+ + + + + +
+ + + + +
+ + + +
+ + + +
+ + + + +
Total area (%) a 75 5 254 29 4 812 1000 73 1 274 320 45 4 418 786 75 0 43 2 318 989 82 9
B The total area for the stieen PAHs was obtamed by adding and normahzmg for each chromatogram
69
WR Trevelur et aL /AnaL Chun Acta 268 (1992) 67-71 TABLE 4
RESULTS AND DISCUSSION
Imtlal
Factor&
destgn
Relative values of areas for the several comblnations m the factorial design are included m Table 2 It is noted that the total area for expenments E,, E,-E,, Ell, Elz, El, and E,, are all greater than 50%, whereas the values for all other experunents are all less than 50% Accordingly, the former represent the most slgmflcant combmatlons Table 3 shows estunates of the factorial analysis from factorial design The order of relevance among the variables which can be observed 1s V>S>SVT>SP>VT>P>ST>SV>TI> PV > T > VZ > PT > SZ > I By comparing
the results expenment for E,, and E,, It 1s observed that lower values of T (effect = -0 4625) and I (effect = -0 9225) give a higher response Combmrng results for E, with these results, It 1s concluded that a lower value of S (effect = 167) results m a slgmflcantly smaller value for the relative area Therefore, It 1s concluded that larger values of S increase the sensltrvlty whereas smaller values of T and I give better sensitivity The factorial design shows that S, V and T are the most slgnlflcant variables Further, the vartable T was mmlmlzed and kept constant and the variable S was maxlmlzed and also kept constant
TABLE 3 Factorial analysis estimate Variable or mterachon
Effect
s
167 0245 0 3875 6 6875 002 -0 19 -75x10-3 - 0 4625 024 -063 - 0 4975 0 3475 046 -0 735 -09225
P SP V sv PV SPV T ST PT SPT VT SVT PLT I
condltlonsfor smplex optrrmzatlon Variables
Constants
(rmn)
S
T (“0
v (/.Ll)
P
Xylene
100
10-30
05-10
I (“Cl 270-290
The maximum of the parameter V and the mmunum value of the parameter I, their own mteractlons and their mteractlons with other parameters can be obtained from factorial design Smpkx optwnlzation The mltlal conditions
for snnplex optmuzatlon are listed m Table 4 Based on these, the chromatographlc conditions to obtain the maxnnum sensltlvlty using Nelder and Mead’s algonthm [lo] were investigated Table 5 shows the progress of the simplex and Fig 1 shows some chromatograms which represent the importance of the simplex optumzatlon of the mJectlon mode with more dilute samples The mitral simplex with vertices V,, V,, V, and V, was based on the mdlcatlon supplied by factorial design Followmg the algorithm, vertex V, was rejected as it gave the worst response (66 6%), and vertex V, was obtained by reflection of V, through the experiments containing the vertices V,, V,, V, and V, However, m vertex V, the vanable V attams an unreal value which lies outside the optlmlzed surface So this vertex was contracted, obtaining a new system of vertices with V,, V,, V, and V,, thus generating an area value equal to 75 0% The algorithm continued to Vls, because the best response was obtained with V, V, and V, indicate a small increase m the response only due to the purge time, but when compared with V, and V,, both responses are similar, consldermg the analytical error, which demonstrates that the purge time is not a very important variable Concluswns The largest contribution was due to the amount of solvent, as expected Xylene increases the area values mainly of the late-elutmg peaks Another
70
WR Trevehn et al /Anal
Chtm. Acta 268 (1992) 67-71
TABLE 5 Progress of the simplex Vertex No
Variables v (rl)
p (mm)
I (“Cl
Vl
20 30 20 20 27 31 23 33 23 30 34 26 32 28 31
0 70 050 050 050 067 072 0 55 042 0 63 0 62 054 061 0 56 0 60 057
280 280 285 280 283 277 283 284 281 280 281 281 281 281 281
v2
V3 v4
v, V6 a Vl Vs a v9 VlO Vll v12 v13 v14 VlS
Retained vertices
Response (%I
1,2,3,4 1,2,3,5 _a 1,2,5,7 _a 275,799 2,5,9, 10 _= 2,5, 10, 12 _a 2,5, 10, 14 _a
718 1000 669 66 6 88 6 _a 75 0 _= 79 3 97 1 _a 82 2 _= 88 2 _a
’ The variable V attams an unreal value
0
1 1 ?
0
10
Fig 1 Gas chromatogram of the stieen
20
30
40 *
PAHs (a) after the best optlmlzation W2 condltlon), (b) under the V, condition
WR Treveh et al /Anal
71
Chm Acta 268 (1992) 67-71
nnportant parameter IS the mltlal column temperature, whereas the purge tnne and mJectlon port temperature are less significant These results show that the sensltlvlty 1s strongly mcreased by adoptmg the optimum chromatographlc conditions To determine trace constituents m a sample rt 1s essential to establish the best conditions for analysis, for which chemometrics 1s a useful tool The authors thank FINEP/PADCT for fmanclal support and Wagner R Trevelm thanks CAPES for grants The authors are also grateful to Dr Horatio A Mottola for reading and correcting the manuscript
REFERENCES 1 Samphng and Analysis Procedures for Screenmg of Industrial Effluents for Pnor~ty Pollutants, US Envtromnental Protection Agency, Environmental Momtormg and Support Laboratory, Cmcmnah, OH, 1977 2 M J Shear, Am J Cancer, 26 (1936) 322 3 K. Grob and K Grob, Jr, J Chromatogr SCI, 7 (1969) 584 4 K. Grob and K. Grob, Jr, J Chromatogr , 94 (1974) 53 5 K. Grob and K Grob, Jr, J Hugh Resolut Chromatogr Chromatogr Commun , 1 (1978) 57 6 W G Jennmgs, R R Freemann and T A Rooney, J High Resolut Chromatogr Chromatogr Commun, 1 (1978) 275 7 R D De Veaux and M Szelewslu, J Chromatogr SCI,27 (1989) 513 8 K Grob, ClassIcal Spht and Sphtless InJectIon m Capillary Gas Chromatography, Huthlg, Heidelberg, 1986 9 W Spendley, G R Hext and F R Hlmsworth, Technometrlcs, 4 (1962) 441 10 J A Nelder and R Mead, Comput J , 7 (1965) 308
Analytrco Chlmlca Acta, 268 (1992) 67-71 Elsevler Science Publishers B V . Amsterdam
Optimization of parameters for the gas chromatographic determination of polycyclic aromatic hydrocarbons W R Trevehn, L H Vldal, M D Landgraf, I C E Srlva and M 0 0 Rezende Unwerszdade de Silo Paula, Instztuto de Fistca e Quimcca de Srio Carlos, Departamento de Quimrca e Fika Molecular, Au Dr Carlos Botelho 1465, Cx P 369, 13560 SZo Carlos SP (Brazd) (Received 5th February 1992, revised manuscnpt received 2nd Aprd 1992)
Ahstraet
The optmuzatlon of condltlons for the gas chromatographlc determination of sixteen polycychc aromatic hydrocarbons (PAHs) using the sphtless mJectlon mode 1sdescribed Parameters optlmlzed were the type and amount of solvent, the temperature of the mnjtion port and the column and the purge time A factonal design was used uutlally to ldentlfy the most important parameters Based on this mttlal study, xylene was chosen as the solvent and 100°C was chosen as the best value for temperature of the column The other parameters were optmuzed by using a sunplex method The optlmum condltlons found were 3 ~1 of solvent, 280°C for the mJectlon port and a purge time of 30 s With these condltlons, relatwe standard devlatlons for five rephcate determmattons of 2-ng amounts of PAHs were 3 2-10 8% Keywords Gas chromatography,
Optmuzatlon methods, Factonal design, Polycychc aromatic hydrocarbons, Simplex
Polycychc aromatic hydrocarbons (PAHs) represent a class of organic compounds identified as “unambiguous priority pollutants” by the US Envlronmental Protection Agency (EPA) [l] This class of compounds has been shown to contam several potent carcmogens [2] The occurrence and potential hazards of these compounds have stimulated research related to the determmatlon of PAHs m the environment Gwen the low concentrations of these species, it 1s important to use condltlons that optimize the separation efficiency and detection hmlts m envrronmental samples Grob and Grob [3,4] ldentlfled the solvent effect as one of the most lmportant factors for alkanes such as heptane, octane Correspondence to M 0 0 Rezende, Umversldade de Slo Paulo, Instltuto de Fislca e Quinuca de SCo Carlos, Departamento de Quinnca e Fiilca Molecular, Av Dr Carlos Botelho 1465, CxP 369, 13560 Sb Carlos SP (Brazil)
and nonane determined using gas chromatography (GC) with the sphtless mJectlon mode Cold trapping was orlgmally described as the mechamsm operating m the sphtless mJectlon mode Subsequent studies [4,5] showed that the solvent effect 1s important for both sphtless and on-column mjectlon Based on Grob and Grob’s studles, Jennmgs et al [6] described the theoretical basis of the solvent effect Cold trapping and the solvent effect were described as the two functions that led to the reconcentration of elutes on the top of the column The factors affecting splitless mjection are numerous, e g , inJection speed, liner size, matnx composition, presence of a liner plug [7], column temperature, volatlhty of the solvent, amount of solvent and tune of vaporlzatlon [8] Umvarlate studies, although useful, are seldom completely satisfactory for complex processes such as m GC that depend on multiple variables The
0003-2670/92/$05 00 0 1992 - Elsevler Science Pubhshers B V All nghts reserved
68
WR Trevelm et al /And
best condltlons are usually obtamed by multlvanate studies such as factorial design and simplex methods such as the sequential simplex method introduced by Spendley et al [9] and improved by Nelder and Mead [lo] In this study, a factorial design was used to identify the most important variables m GC to increase the sensltlvlty of sixteen PAHs and a simplex method was used to identify the optimum values for the most nnportant parameters, namely the volume of solvent, the temperature of the mjectlon port and the purge time
EXPERIMENTAL
Reagents and standards
All solvents were glass distilled through a 150cm column The compounds studied were naphthalene, acenaphthene, acenaphthylene, fluorene, phenanthrene, anthracene, fluoranthene, pyrene, bendalanthracene, chrysene, benzo[ blfluoranthene, benzo[ klfluoranthene, benzo[ alpyrene, dlbenzo[a,h]anthracene, benzo[ ghzlperylene and mdeno[l,2,3_cdlpyrene (Supelco, Bellefonte, PA) A stock solution was prepared m a mixed benzene-methylene chrorlde (1 + 1, v/v> solvent contamg a nommal concentration of 2 mg ml-’ of each PAH Further dllutlons were made with benzene and xylene to give a fmal concentration of 2 pg ml-’ of each PAH Ahquots of the stock solution were measured using a 100~~1 Type 701 RN syringe (Hamilton, Reno, NV) Chromatographu:
condltwm
All experiments were done usmg a Model 5890 high-resolution gas chromatograph (Hewlett Packard, Palo Alto, CA) equipped with a flame lomzatlon detector (FID) The output of the FID was connected to an integrator (HP-33931 A fused-s&a capillary column coated with crosslmked Ultra-l methyl&cone (Hewlett-Packard) as the stationary phase (25 m x 0 32 mm 1 d , flhn thickness 0 17 pm> was used with splitless mjectlon Some expenmental conditions, mcludmg solvent, temperature of the mJectlon port, initial temperature of the column, amount of solvent
Chun Acta 268 (1992) 67-71
TABLE 1 Parameters used m factorial design Variable
Maxunum level ( + )
Mnumum level (-)
S = solvent P = time of purging V = amount of solvent T = uutlal column temperature I = injectIon port temperature
Benzene 05mm 10 /Ll 100°C 270°C
Xylene 1Omm 3O/.Ll 12Wc 290°C
and purge tnne, were varied during the process Other parameters, mcludmg flow-rate of the carrier gas (40 cm s-l), heating rate (5°C mm-‘) and fmal temperature (280’0, were the same m all experiments Peak areas for all PAHs were added and normahzed for each chromatogram Factonal design
Values of the parameters used m the factorial design are listed m Table 1 The combmatlons of these mmunum and maximum levels are shown m Table 2
TABLE 2 Combmatlons among the mmnnum and maximum levels of the parameters listed m Table 1 Experiment No E, E2 E3 E4 Es E6 E7 E8 E, EIO ‘%l
E 12 E13
E 14 E 1.5 El6
Variable S +
P
V
T
I
-
-
+
-
+
+
-
+
+
+ -
+ +
+ + + -
+ + + -
+ -
+ + + + + +
+ + + + +
+ + + +
+ + + +
+ + + + +
Total area (%) a 75 5 254 29 4 812 1000 73 1 274 320 45 4 418 786 75 0 43 2 318 989 82 9
B The total area for the stieen PAHs was obtamed by adding and normahzmg for each chromatogram
69
WR Trevelur et aL /AnaL Chun Acta 268 (1992) 67-71 TABLE 4
RESULTS AND DISCUSSION
Imtlal
Factor&
destgn
Relative values of areas for the several comblnations m the factorial design are included m Table 2 It is noted that the total area for expenments E,, E,-E,, Ell, Elz, El, and E,, are all greater than 50%, whereas the values for all other experunents are all less than 50% Accordingly, the former represent the most slgmflcant combmatlons Table 3 shows estunates of the factorial analysis from factorial design The order of relevance among the variables which can be observed 1s V>S>SVT>SP>VT>P>ST>SV>TI> PV > T > VZ > PT > SZ > I By comparing
the results expenment for E,, and E,, It 1s observed that lower values of T (effect = -0 4625) and I (effect = -0 9225) give a higher response Combmrng results for E, with these results, It 1s concluded that a lower value of S (effect = 167) results m a slgmflcantly smaller value for the relative area Therefore, It 1s concluded that larger values of S increase the sensltrvlty whereas smaller values of T and I give better sensitivity The factorial design shows that S, V and T are the most slgnlflcant variables Further, the vartable T was mmlmlzed and kept constant and the variable S was maxlmlzed and also kept constant
TABLE 3 Factorial analysis estimate Variable or mterachon
Effect
s
167 0245 0 3875 6 6875 002 -0 19 -75x10-3 - 0 4625 024 -063 - 0 4975 0 3475 046 -0 735 -09225
P SP V sv PV SPV T ST PT SPT VT SVT PLT I
condltlonsfor smplex optrrmzatlon Variables
Constants
(rmn)
S
T (“0
v (/.Ll)
P
Xylene
100
10-30
05-10
I (“Cl 270-290
The maximum of the parameter V and the mmunum value of the parameter I, their own mteractlons and their mteractlons with other parameters can be obtained from factorial design Smpkx optwnlzation The mltlal conditions
for snnplex optmuzatlon are listed m Table 4 Based on these, the chromatographlc conditions to obtain the maxnnum sensltlvlty using Nelder and Mead’s algonthm [lo] were investigated Table 5 shows the progress of the simplex and Fig 1 shows some chromatograms which represent the importance of the simplex optumzatlon of the mJectlon mode with more dilute samples The mitral simplex with vertices V,, V,, V, and V, was based on the mdlcatlon supplied by factorial design Followmg the algorithm, vertex V, was rejected as it gave the worst response (66 6%), and vertex V, was obtained by reflection of V, through the experiments containing the vertices V,, V,, V, and V, However, m vertex V, the vanable V attams an unreal value which lies outside the optlmlzed surface So this vertex was contracted, obtaining a new system of vertices with V,, V,, V, and V,, thus generating an area value equal to 75 0% The algorithm continued to Vls, because the best response was obtained with V, V, and V, indicate a small increase m the response only due to the purge time, but when compared with V, and V,, both responses are similar, consldermg the analytical error, which demonstrates that the purge time is not a very important variable Concluswns The largest contribution was due to the amount of solvent, as expected Xylene increases the area values mainly of the late-elutmg peaks Another
70
WR Trevehn et al /Anal
Chtm. Acta 268 (1992) 67-71
TABLE 5 Progress of the simplex Vertex No
Variables v (rl)
p (mm)
I (“Cl
Vl
20 30 20 20 27 31 23 33 23 30 34 26 32 28 31
0 70 050 050 050 067 072 0 55 042 0 63 0 62 054 061 0 56 0 60 057
280 280 285 280 283 277 283 284 281 280 281 281 281 281 281
v2
V3 v4
v, V6 a Vl Vs a v9 VlO Vll v12 v13 v14 VlS
Retained vertices
Response (%I
1,2,3,4 1,2,3,5 _a 1,2,5,7 _a 275,799 2,5,9, 10 _= 2,5, 10, 12 _a 2,5, 10, 14 _a
718 1000 669 66 6 88 6 _a 75 0 _= 79 3 97 1 _a 82 2 _= 88 2 _a
’ The variable V attams an unreal value
0
1 1 ?
0
10
Fig 1 Gas chromatogram of the stieen
20
30
40 *
PAHs (a) after the best optlmlzation W2 condltlon), (b) under the V, condition
WR Treveh et al /Anal
71
Chm Acta 268 (1992) 67-71
nnportant parameter IS the mltlal column temperature, whereas the purge tnne and mJectlon port temperature are less significant These results show that the sensltlvlty 1s strongly mcreased by adoptmg the optimum chromatographlc conditions To determine trace constituents m a sample rt 1s essential to establish the best conditions for analysis, for which chemometrics 1s a useful tool The authors thank FINEP/PADCT for fmanclal support and Wagner R Trevelm thanks CAPES for grants The authors are also grateful to Dr Horatio A Mottola for reading and correcting the manuscript
REFERENCES 1 Samphng and Analysis Procedures for Screenmg of Industrial Effluents for Pnor~ty Pollutants, US Envtromnental Protection Agency, Environmental Momtormg and Support Laboratory, Cmcmnah, OH, 1977 2 M J Shear, Am J Cancer, 26 (1936) 322 3 K. Grob and K Grob, Jr, J Chromatogr SCI, 7 (1969) 584 4 K. Grob and K. Grob, Jr, J Chromatogr , 94 (1974) 53 5 K. Grob and K Grob, Jr, J Hugh Resolut Chromatogr Chromatogr Commun , 1 (1978) 57 6 W G Jennmgs, R R Freemann and T A Rooney, J High Resolut Chromatogr Chromatogr Commun, 1 (1978) 275 7 R D De Veaux and M Szelewslu, J Chromatogr SCI,27 (1989) 513 8 K Grob, ClassIcal Spht and Sphtless InJectIon m Capillary Gas Chromatography, Huthlg, Heidelberg, 1986 9 W Spendley, G R Hext and F R Hlmsworth, Technometrlcs, 4 (1962) 441 10 J A Nelder and R Mead, Comput J , 7 (1965) 308