20 3 EXPER IM EN TAL - Catalysis in the hydrolysis of esters and related compounds

a) Sources, P u rific a tio n , and P repa ra tion of M a te ria ls .

A ll reagents used in kin etic studies w ere of AnalaR grade.

AnalaR , p yrid in e was refluxed w ith solid potassium hydroxide before being c a re fu lly fra c tio n a lly d is tille d (b. pt. 115^) through a long V ig re u x column.

Reagent grade 1 ,4 -dioxan was refluxed over sodium w ire u n til the surface ^ S of the m etal was not fu rth e r discoloured during several hours. It was then

5 o

ca re fu lly d is tille d (b . pt. 101 ) fro m the sodium ^utilising a V ig re u x column.

Thus p u rifie d the dioxan was kept in a froze n state u n til needed, to prevent % 33

peroxide and aldehyde fo rm a tio n .

D euteriated w a ter w ith a stated deuterium content of 99.7% was supplied ï by K o ch-L igh t L a b o ra to rie s.

P repa ra tion of substituted phenyl chloroform âtes 4

The substituted phenyl chloroform âtes u tilis e d in this study are a ll

d known compounds^^’ and w ere prepared by methods already cited in the s

36 37 38

lite ra tu re ’ ’ . P h ysical data concerning these compounds is recorded in < Table 6, and the general p repa rative method is given below.

A solution of phosgene (0.14 mole) in toluene (125 m l) was cooled to - 10^ in a three-necked fla s k equiped w ith therm om eter, m agnetic s tir r e r , re flu x condenser, and dropping funnel. A solution of the substituted phenol

(0,12 mole) in N, N -d ie th yla n ilin e (0.12 mole) was added, w ith s tirrin g , w h ils t m aintaining the tem perature below 0^. If the substituted phenol was insoluble

—21—

in N, N -d ie th yla n ilin e , anhydrous AnalaR acetone (10-20 m l) was added to achieve solution. The reaction m ix tu re was then kept below 0^ fo r three hours, a fte r w hich tim e the tem perature was slow ly allowed to ris e to room tem pera tu re . The N, N -d ie th yla n ilin e hydrochloride form ed separated out as a p re cip itate and was rem oved by filtra tio n . The reaction m ix tu re , cooled to - 15? was then s tirre d vig o ro u sly and cold 5% h yd ro ch lo ric acid (7 5 m l) was added. The toluene la y e r was then separated, washed w ith ice w a ter (75 m l), and dried over anhydrous sodium sulphate. Once dried , the sodium sulphate was removed by filtra tio n , and the toluene was removed fro m the re s u ltin g solution by d is tilla tio n under reduced pressure. The substituted phenyl chloroform ate which rem ained was then p u rifie d by careful fra c tio n a l d is tilla tio n under reduced pressure.

P re p a ra tio n of phenyl azidoform ate

Sodium azide (0. 65g) dissolved in the m inim um quantity of 10% aqueous dioxan was added slow ly to a s tirre d solution of fre s h ly d is tille d phenyl

chloro fo rm ate (1.36 g) in dioxan (25 m l). The m ix tu re was s tirre d fo r 4 hours and the solvents rem oved under vacuum. The residue was dissolved in ether, washed tw ice w ith cold w a ter, and drie d (MgSO^). The ether was rem oved to leave a liq u id , w hich was assumed to be phenyl azidoform ate, ^ _max 2150 cm-1

(Ng). F u rth e r p u rific a tio n and id e n tifica tio n was not attempted because of the 39

danger of explosion .

b) K inetic Method

—22—

added to the b u ffer contained in a cuvette in the therm ostatted c e ll holder of a Unicam SP 500 spectrophotom eter. The change in absorbance w ith tim e was recorded, using a Unicam SP 22 c h a rt-re c o rd e r and a SP 505 program m e re co rd e r. The fir s t- o r d e r plots obtained w ere lin e a r over three h a lf-liv e s . Rate constants w ere calculated by the method of Swinbourne (Appendix 2).

T A B LE 6

P h ysical data concerning substituted phenyl C hloroform âtes

The wavelengths used w ere as shown in Table 6. The io n ic strength was kept constant by the addition of K C l. Spectra w ere recorded on a Unicam SP 800 spectrophotom eter. A Beckman ’Research’ pH m e te r w ith calom el electrode and general purpose glass electrode was used fo r the m easurem ent

Substituent B .P . B .P . L it. % Y ie ld A (obs)nm Reference

4

H - 78-80° (22 mm) (22 mm)80° 84 274 ! 36 % 4 -M e - 118-20 (40 mm) (30 mm)108 89 282 36 1 3 -M e - 95-97 (20 mm) (22 mm)103 78 275 37 4-M eO - 132-34 (25 mm) (24 mm)132 87 293 34 4 -C l- 114-15 (20 mm) (20 mm)114 80 274 38 ^ 4-NO - 80-81°G (M .P .) 8 0 -l°M P 96 400 36

—23—

of pH, Values of pD , w ere obtained by using the re latio nship, given previo usly, 18

24"

C H A P T E R TW O

NUCLEOPHILIC AND GENERAL BASE CATALYSIS BY PYRIDINE AND M ETHYLPYRIDINES IN THE HYDROLYSIS

..V.

-2 5 - 1. INTRODUCTION O , O

Il r L .

Il

i

h

Y i ^

H

(Scheme 4) 9_Il

o

_Il

CH—c — OAr --- > CH—C +- OAr

3 t 3 I

B

" ■B'*"

H g O

fast

Ç

(2)

C Hg-C—OH 4- BH*^

(Scheme 5)

Numerous investigations^^ have been made of base-catalysed ester [ hyd ro lysis. In p a rtic u la r, the mechanism of h yd ro lysis of a ry l acetates has 4 been examined in great de ta il ; these compounds are w idely used as substrates

in enzym atic studies. The two mechanism s w hich have been generally proposed

fo r the base-catalysed h yd ro lysis of such compounds are (1) general base catalysis i (Scheme 4), where attack of a w ater m olecule on the carbonyl group of the a ry l

ester is assisted by p a rtia l bond fo rm a tio n to the base (B) and, (2) nucleophilic catalysis (Scheme 5) where the base attacks the ester d ire c tly to give an in te r m ediate w hich is ra p id ly hydrolysed.

« 26-

In spite of extensive study, however, it is s till not possible to p re d ict w ith ce rta in ty whether any p a rtic u la r catalysing base w ill act as a nucleophilic o r a general base catalyst. The w o rk in this section of the thesis was undertaken in o rd e r to investigate the c rite ria concerned w ith this p lu ra lity of m echanism .

20 21

I t has p re vio u sly been shown, by Gold and his co -w o rke rs ’ , that the two mechanism s m ay occur concurrently in ce rta in instances. The

simultaneous existence of the two m echanisms was dem onstrated by trapping the interm ediate produced in the nucleophilic pathway and com paring the amount of interm ediate form ed w ith the amount of reactant consumed. T h e ir general

conclusion was that the tendency fo r nucleophilic catalysis to occur increases as the leaving group becomes b e tte r (i. e. the lo w er the pK^ of the phenol). I t was shown, fo r instance, that the acetate-catalysed h yd ro lysis of 2 ,3-dinitrophenyl acetate (pK^ = 4. 96) is 100% nucleophilic w h ils t the corresponding reaction fo r 4-m ethylphenyl acetate (pK^ = 10. 26) is 100% general base catalysed. I t is fo r phenols of interm ediate pK^ ( and hence of leaving tendency) that both pathways may occur ; fo r example, the acetate-catalysed h yd ro lysis of 4-nitrophe nyl acetate was found to be 70% nucleophilic and 30% general base catalysis. Gold

and co-w orkers suggest that the v a ria tio n of m echanism can be predicted, to some extent, by consideration of the differences in pK^ values fo r the acetate ion and the phenoxide ion ; where the phenoxide ion is much m ore basic than the acetate ion ( A p K ^ greater than 3), the nucleophilic m echanism becomes

in sig n ifica n t. Gold et a l. point out, fro m consideration of Scheme 6, that the breakdown of the interm ediate by the alterna tive routes -1 and 2 w ill depend

-2 7 -

1 I

? "

ArO.CO.Me-f- AcO ;? = ± A rO -C -M e

|

OAg

A r O 4- Ac O

(Scheme 6)

upon the anionic s ta b ility of the alterna tive leaving groups (acetate o r phenoxide) w hich, fo r com parisons w ith in a se rie s w ith close s tru c tu ra l s im ila ritie s in the leaving groups involved, can be equated w ith the b a sicity of the species. F o r

the acetate ester of a v e ry acidic phenol, the reverse step (-1) w ill be re la tiv e ly 4 in sig n ifica n t compared w ith the favoured breakdown by route 2. U nder these

circum stances, nucleophilic catalysis should be im p ortant, although the

nucleophilic component w ill decrease as the quotient (k. k ) / (k + k ) decreases._{1 z} _-1 _z In term s of such an analysis the c o rre la tio n of the onset of nucleophilic catalysis w ith increasing leaving tendency can be understood, although there seems to be no obvious reason why general base catalysis should cease. The analysis of Gold and his co -w o rke rs im p lic itly assumes that general base catalysis occurs w ith a ll este ri^b u t that once nucleophilic catalysis becomes

30 possible, it is the predom inant pathway. In a re vie w of this topic Johnson

quotes Brj6nstedJp values of ca. 0. 5 and ca. 0. 8 fo r general base and nucleophilic catalysis. Hence, the nucleophilic pathway is lik e ly to be much m ore effective than the general base catalysed reaction and i t is c le a r that, except in a few special b o rd e rlin e cases, i t would be d iffic u lt to detect the general base catalysed reaction when nucleophilic catalysis occurs. Under experim ental conditions

—28—

one w ill generally detect only the predom inant pathway, as was noted in Chapter 1 ; those reactions in vo lvin g both pathways m ust be ve ry fa s t if the general base catalysis pathway is to be detected. In view of this c rite rio n , this study has necessitated the use of stopped-flow spectrophotom etry.

The ejqperimental w ork designed to fa c ilita te the observation of the two separate pathways was based on the follow ing rationa le : p yrid in e , an effective

catalyst in the h yd ro lysis of acetic anhydride , is rendered c a ta ly tic a lly inactive 29 42

by substitution of a m ethyl group at the 2-position ’ , due to s te ric hindrance to nucleophilic attack. The same effect is observed if substitution occurs in the

anhydride, fo r B u tle r and Gold have shown that p yrid in e is not a catalyst fo r the h yd ro lysis of 2 ,2-dim ethylpropionic anhydride. P y rid in e is also a catalyst

fo r the h yd ro lysis of a ry l acetates and if the m echanism is the same then, by analogous reasoning to that above, the a b ility of p yrid in e to act as a nucleophile in this reaction can also be rem oved by substitution at the 2-position. Any general base catalysed reaction w hich occurs concurrently w ith the nucleophilic catalysis w ill then be the only reaction and, under these circum stances, it w ill be possible to observe and characterise this reaction pathway.

:: 4

4

_#₆ I -2 9 - I A 1 2. RESULTS AND DISCUSSION

The h yd ro lysis of 4-nitrophe nyl acetate in a series of p yrid in e and |

I

3-m e th ylp yrid in e b u ffe rs, prepared by h alf neu tralisation of the p yrid in e base % w ith h yd ro ch lo ric acid, was examined at three tem peratures and the re su lts a re 4 given in Table 7. F rom these re su lts i t was found that a p lo t of lo g k(obs)

against concentration of fre e (i. e. unprotonated) p yrid in e and 3-m e th ylp yrid in e $

is lin e a r in a ll cases, w ith no in te rce p t (Figure 6 and 7). The ca ta lytic | coefficients, given at the foot of each colum n in this table, w ere used to calculate 4 activation param eters fo r the two reactions (Table 11).

The n o n -p articipation of a w ater m olecule in the ra te -d e te rm in in g step

of a nucleophilic ca ta lytic process means that, as an in d ica to r of nucleophilic | catalysis, the ra tio k(HgO)/k(DgO) should have a value of about unity^^; this 4 c rite rio n should, however, be used w ith caution, although it has a fa ir ly wide

v a lid ity . The ra te of p yridine-catalyse d hydrolysis of 4-nitrophe nyl acetate in DgO at 25. 0° was m easured and the re su lts are given in Table 8. The ca ta lytic coefficient is 1. 86 x 10 ^1 m ol ^s ^ and k(H O) / k(D O) fo r the reaction is thus 1. 0. T his re s u lt provides good evidence that the reaction exhibits nucleophilic catalysis.

Another c rite rio n fo r nucleophilic catalysis has been proposed by Gold 21

and his co -w o rke r who found that in the acetate-catalysed h yd ro lysis of a ry l acetates, nucleophilic catalysis is associated w ith a sm a ll negative entropy of

-1 -1

activation (-10 cal m ol K ) w h ils t , fo r general base catalysed reactions the value is much m ore negative (-30 cal m ol K ). The absolute values o f / \ S

—30- ,

■i

Table 7

}

The h yd ro lysis of 4-nitrophenyl acetate in buffered solution at various tem peratures

I = 0. 5 M [PKPA] ^ = ca. lO "^ M 4 - 1 4 10 k(obs) s r o n--- ^ i T C 25.5 35.5 45.0 | Ç 0.080 1.96 4.56 8.42

I

0.16 3.68 7.70 4 Î 0.24 5.26 10.2 20.3 4 4 0.32 6.88 12.8 î 0.40 8.32 15.8 30.6 3 -1 -1 10 k 1 m ol 8 1.88 3.42 6.80 f r e e ^ 0.080 0.16 0.24 0.32 0.40 , 3 , , -1 -1 10 k 1 m ol s

I

6. 50 8.70 16.0

₁

i. 8.70 17.7 34.6 J 12.0 24.5 46.3 _■| 14.7 30.1 51.4 19.6 36.5 66.7 I 4.46 8.12 14.9 "1

—31“

(Figure 6 ; V a ria tio n of k(obs) w ith p yrid in e concentration fo r the hyd ro lysis of PK PA at 25. 5°)

JÛ

0-2

0 - 4

(F igure 7 ; V a ria tio n of k(obs) w ith 3 -m e thylpyridine concentration fo r the h yd ro lysis of PNPA at 25. 5^)

—32—

"i:

found in this present study do not correspond w ith those found fo r the acetate-

catalysed reactions. T his c rite rio n , although not co n firm in g the m echanism as | being nucleophilic ca talysis, w ill, however, be used la te r to indicate a change j in m echanism . In view of the size of the kinetic isotope effect fo r the reaction , "I

and the previous study of the ro le of p yrid in e in the h yd ro lysis of acetic anhydride, there seems to be little doubt that the present reactions exhibit nucleophilic catalysis.

The substitution of a m ethyl group at the 2-position o f p yrid in e prevents nucleophilic catalysis fo r s te ric reasons, as has been previo usly mentioned ; if , then, 2-m e thylpyridine accelerates the hyd ro lysis of 4-nitrophe nyl acetate, the mechanism m ust be other than nucleophilic catalysis. The pyridine-catalyse d reaction of 4-nitrophe nyl acetate is not its e lf a p a rtic u la rly fa st reaction, and it was found that w ith 2-m e thylpyridine no other reaction, apart fro m the spontaneous h yd ro lysis, could be detected. However, if the general-base

catalysis pathway is only a sm all fra c tio n of the total reaction, then that reaction may be too slow to detect. T h is study, using 4-nitrophe nyl acetate, established that the pyridine-catalyse d ester hyd ro lysis is nucleophilic ; s im ila r experim ental evidence using a m ore re a ctive ester would have required p ro h ib itiv e ly la rg e quantities of D^O to enable the kin etic isotope effect to be m easured in a stopped- flow apparatus.

A m ore reactive ester, 2 ,4 -din itro phenyl acetate, was chosen fo r fu rth e r study. Its ra te of hyd ro lysis in the presence of pyrid in e , 3 -m e thylpyridine, and 4 -m e th ylp yrid in e was examined by use of a stopped-flow spectrophotom eter.

i

-33-

Table 8

H yd rolysis of 4 -nitrophe nyl acetate in D^O at 25^ Tables 9-11 I = 0.5 Ivl [2, 4-D N P A ] = ca. lO "^ M Table 9 4-m e th ylp yrid in e k (obs) s ^ T°C 'TsTS ' m Ts 4 4 .5 ^

U-CHsCgH^N]

M

0.036 0.34 0.50 0.78 0.040 0.48 0.71 1.08 0.054 0. 51 0.85 1.26 0.072 0.63 0.94 1.43 0.20 1.94 3.22 4.96 0.40 3.38 I = 0.5 M [p N P A ]^ = ca 1 0 "^ M

f

[c H N ] _ M 0.11 0.26 0.36 0.46 5 5** fre e 10^ k(obs) s” ^ 2.02 4.90 6.80 8.20

H yd ro lysis of 2 ,4-din itro phenyl acetate catalysed by various p yrid in e bases

—34— Table 10 3-m e thylpyrldlne k (obs) 8 ^ 0_ /--L:"" % ...--- T C 25.2 34.8 44.5 " 0.168 .106 . 165 .264 0.336 .210 .323 .505 0.440 .275 .425 .662 Table 11 P y rid in e k (obs) s ^ T°C ' 25.2 34.8 4 4 .5 fre e 0.028 0.047 0.088 0.120 0.175 0.281 0.500 0.770 0.250 0.410 0.680 1.070 0.280 0.465 0.775 1.240 0.375 0.595 1.100 1.580 4

In document Catalysis in the hydrolysis of esters and related compounds (Page 30-69)