M[

MD

a

ARRAA

KMC[NCG

1

edimpd[tot

scaTmis[oe

0d

Colloids and Surfaces B: Biointerfaces 94 (2012) 220– 225

Contents lists available at SciVerse ScienceDirect

Colloids and Surfaces B: Biointerfaces

j our na l ho me p age: www.elsev ier .com/ locate /co lsur fb

icelle-catalyzed reaction between ninhydrin and nickel dipeptide complexNi(II)–Gly-Tyr]+

ohd. Akram ∗, Dileep Kumar, Kabir-ud-Dinepartment of Chemistry, Aligarh Muslim University, Aligarh 202 002, U.P., India

r t i c l e i n f o

rticle history:eceived 5 November 2011eceived in revised form 24 January 2012ccepted 25 January 2012vailable online 3 February 2012

a b s t r a c t

The interaction of nickel dipeptide complex [Ni(II)–Gly-Tyr]+ with ninhydrin has been investigated in theabsence and presence of cationic cetyltrimethylammonium bromide (CTAB) and gemini (16-s-16, s = 4,5, 6) surfactants spectrophotometrically at 80 ◦C and pH 5.0. The product formed was the same and thereaction followed first- and fractional-order kinetics with respect to [Ni(II)–Gly-Tyr]+ and [ninhydrin],respectively, in both aqueous as well as micellar media. In the presence of CTAB, rate increased andreached up to a maximum, then decreased. Also, whereas typical rate constant (k ) increase and leveling-

eywords:icelle

atalysisNi(II)–Gly-Tyr]+

inhydrinTAB

�

off regions, just like CTAB, were observed with geminis, the latter produced a third region of increasingk� at higher concentrations. This unusual third-region effect of the gemini micelles is assigned to changesin their micellar morphologies. The micellar catalysis is explained in terms of pseudo-phase model. Thebinding constants and the values of activation parameters such as activation energy (Ea), enthalpy ofactivation (�H#) and entropy of activation (�S#) have been evaluated.

emini surfactants

. Introduction

Systems involving surfactants constitute a field of great inter-st due to their wide ranging uses in pharmaceutical formulations,etergent and petroleum recovery processes. Surfactant aggregates

n water and in apolar solvents have also been utilized to mimic theicro-environments of biomacromolecular ensembles. Therefore,

hysics, chemistry and biology meet at the frontier area of inter-isciplinary research on association colloids formed by surfactants1]. Various kinetic studies have been undertaken in micellar mediao elucidate the actual micellar rate effect caused by a compositef noncovalent interactions between the micelles on one hand andhe reaction and activated complex on the other.

Gemini or dimeric surfactants are amphiphilic molecules con-isting of two hydrophobic tails and two hydrophilic head groupsovalently connected through a spacer rather than one hydrophilicnd one/two hydrophobic group(s) of conventional surfactants.he gemini surfactants are superior on several counts than theironomeric counterpart single chain conventional surfactants. The

ncreasing studies of geminis than conventional surfactants areubjected due to their unusual solution and interfacial properties

2–7]. The greater efficiency and effectiveness of gemini micellesver comparable conventional surfactants make them highly cost-ffective as well as environmentally desirable.

∗ Corresponding author. Tel.: +91 571 2703515.E-mail address: drmohdakram@rediffmail.com (Mohd. Akram).

927-7765/$ – see front matter © 2012 Elsevier B.V. All rights reserved.oi:10.1016/j.colsurfb.2012.01.041

© 2012 Elsevier B.V. All rights reserved.

The application of ninhydrin for the estimation and detectionof amino acids/peptides has great importance in revealing latentfinger prints (due to the formation of colored product knownas Ruhemann’s purple, DYDA [8]). The color-forming ninhydrin-amino acids/peptides reaction have characteristics of commonaddition–elimination type reactions. As the color of purple dyefaints at room temperature, many attempts were carried out tostabilize it. Metal ion complex formations are the prominent inter-actions in nature. The effect of metal ions on this reaction were alsocarried out with the point of view of promoting the nucleophilicattack. The condensed product acts as a potential tridentate metalbinding ONO donor ligand producing stable five membered metalchelate. As a modification, therefore, interaction of ninhydrin withmetal–amino acid complexes were also tried, and the color yieldwas indeed affected [9–11]. Our group has made notable contribu-tion toward the enhancement of Ruhemann’s purple yield (henceincreased sensitivity) of ninhydrin-amino acid reaction involvingsurfactant micelles and solvents too [12–15]. However, studies onninhydrin-peptide reaction are scanty [16,17] and, therefore, insearch of enhanced utility, we have studied the [Ni(II)–Gly-Tyr]+ –ninhydrin reaction in micelle mediated aqueous medium undervarying experimental conditions. For this purpose we used cationicgemini surfactants alkanediyl-�, �-bis(dimethylcetylammoniumbromide) (16-s-16, s = 4, 5, 6). For comparison, the monomeric

cetyltrimethylammonium bromide (CTAB) was also used.

Proteins are a class of organic compounds which are presentin and vital to every living cell. In the form of skin, hair, callus,cartilage, muscles, tendons and ligaments, proteins hold together,

rfaces B: Biointerfaces 94 (2012) 220– 225 221

pitotAaa

2

2

9n(d(9pmi

2

mmccwrctN

B NM +Br-

1C

2

i(1atnca

2

id[

6005505004504000.0

0.2

0.4

0.6

0.8

1.0

1.2

edcba

Ab

so

rba

nce

λ (nm)

Fig. 1. Spectra of the reaction product of ninhydrin with [Ni(II)–Gly-Tyr]+ inabsence and presence of surfactants in acetate buffer solution (pH = 5.0) in (a)aqueous medium, (b) 16-6-16, (c) 16-5-16, (d) 16-4-16, and (e) CTAB. Reactionconditions: [16-s-16] = 30 × 10−5 mol dm−3 (s = 4, 5, 6), [CTAB] = 30 × 10−3 mol dm−3,[ninhydrin] = 6.0 × 10−3 mol dm−3, [Ni(II)–Gly-Tyr]+ = 3.0 × 10−4 mol dm−3, temper-

Mohd. Akram et al. / Colloids and Su

rotect, and provide structure to the body of multicelled organ-sm. In the form of enzymes, harmones, antibodies, and globulins,hey catalyze, regulate, and protect the body-chemistry. In the formf hemoglobin, myoglobin and various lipoproteins, they affecthe transport of oxygen and other substances within an organism.mino acids not only act as building blocks in protein synthesis butlso play a significant role in metabolism, microbiology, nutritionnd pharmaceuticals.

. Experimental

.1. Materials

CTAB (Merck, 99.0%), ninhydrin (Merck, 99.0%), Gly-Tyr (SRL,9.0%), acetic acid (Merck, 99.0%), sodium acetate (Merck, 99.0%),ickel nitrate hexahydrate (s.d. fine, 98.0%), 1,6-dibromohexaneFluka, >97.0%), 1,5-dibromopentane (Fluka, >98.0%), 1,4-ibromobutane (Fluka, >98.0%), N, N-dimethylhexadecylamineFluka, >95.0%), ethyl acetate (HPLC and spectroscopy grade,9.0%), and ethanol absolute (Merck, 99.8%) were used as sup-lied. An acetate buffer of pH 5.0 was used as solvent. The pHeasurements were made using a digital ELICO LI-122 pH meter

n conjunction with a combined electrode.

.2. Synthesis and characterization of gemini surfactants

The gemini (alkanediyl-�, �-bis(dimethylcetylammonium bro-ide)) surfactants were synthesized as follows. A 1:2.1 equivalentixture of corresponding �, �-dibromoalkane with N, N-dimethyl-

etylamine in dry ethanol was refluxed at 80 ◦C for 48 h. Afterompletion of the reaction (as monitored using TLC), the solventas removed under vacuum and the solid thus obtained was

ecrystallized four to five times from ethyl acetate to obtain pureompounds. Purity of all the gemini surfactants was checked onhe basis of C, H, N analysis, which was further characterized by 1HMR [18,19].

r(CH2)sBr

C16 H (NMe2)

(dry ethanol, 80 C, 48 h)0

33 (CH2)sN +

Me2

Br-

C16H33

(where s = 4, 5, 6)

.3. Spectra

The UV–visible spectra of the reaction mixture contain-ng [ninhydrin] (6.0 × 10−3 mol dm−3) and [Ni(II)–Gly-Tyr]+

3.0 × 10−4 mol dm−3), recorded using SHIMADZU-model UV mini240 spectrophotometer, are shown in Fig. 1. It is evident that thebsorbance is higher in presence of surfactants which may be dueo strong association of the product with micelles. Further, witho change in the absorption maxima (�max = 400 nm, Fig. 1), it isoncluded that the same product is formed in both systems (i.e.,queous and micellar).

.4. Job’s method of continuous variations

The Job’s method of continuous variations was performedn aqueous and in micellar media (CTAB and geminis) for theetermination of the composition of reaction product betweenNi(II)–Gly-Tyr]+ complex and ninhydrin. It was found that one

e2

33H6

ature = 80 ◦C.

mole of metal complex associates with one mole of ninhydrin togive the reaction product (Fig. 2).

2.5. Kinetic measurements

[Ni(II)–Gly-Tyr]+ complex was made in situ by mixing 1:1 solu-tions of nickel nitrate and glycyl-tyrosine in a three-necked reactionvessel, placed in a thermostated oil-bath. To prevent evaporation,the reaction vessel was fitted with double-surface water condenser.For stirring and maintaining an inert atmosphere, pure nitrogen gas(free from CO2 and O2) was bubbled through the reaction mixture.The resulting solution was left 30 min to complete complexa-tion. The reaction was then started with the addition of thermallyequilibrated ninhydrin of required volume; the zero-time wastaken when half of the ninhydrin solution had been added. In allthe kinetic runs pseudo-first-order conditions were maintainedby using ≥10-fold excess of [ninhydrin] over [Ni(II)–Gly-Tyr]+.

At definite time intervals, aliquots of the reaction mixture werewithdrawn and absorbance was measured at �max (=400 nm). Thepseudo-first-order rate constants in aqueous (kobs, s−1) and inmicellar medium (k� , s−1) were calculated by using a computer

222 Mohd. Akram et al. / Colloids and Surfaces B: Biointerfaces 94 (2012) 220– 225

1.00.80.60.40.20.0

0.00

0.04

0.08

0.12

e

dc

b

a

ΔA

40

0 n

m

Mole fraction of ninhydrin

Fig. 2. Plots of �A400 nm vs. mole fraction of ninhydrin for determination of compo-sition of the product formed by the reaction of [Ni(II)–Gly-Tyr]+ with ninhydrin in(c

pe

2c

c(tawt

(

(

3

3

(ianobs

O

O

O

+

+

+

OH2

k

K

+

OH2

NH

CH2

O

Ni

H2O O

N O

CHC

O

C

O

P

O

O

Ni

H2O O

NH2

O

NH

NH

CH2

CHCC

O

O

O

Ni

H2O OH2

NH2

CH2

CHCC

O

O

A

B

N

CH2

PhOH

CH2

PhOH

CH2

PhOH

Ph =

at constant values of [Ni(II)–Gly-Tyr]+ (3.0 × 10−4 mol dm−3), tem-

a) aqueous medium, (b) 16-6-16, (c) 16-5-16, (d) 16-4-16, and (e) CTAB. Reactiononditions: [16-s-16] = 30 × 10−5 mol dm−3, [CTAB] = 30 × 10−3 mol dm−3.

rogramme. Further details of kinetic measurements can be foundlsewhere [9–17].

.6. Determination of critical micellar concentration (cmc) byonductivity measurements

The critical micellar concentration (cmc) values of cationicetyltrimethylammonium bromide (CTAB) and dicationic gemini16-s-16, s = 4, 5, 6) solutions were determined using conduc-ivity measurements. The experiments were carried out at 30 ◦Cnd 80 ◦C under varying conditions, i.e., water, water + ninhydrin,ater + [Ni(II)–Gly-Tyr]+, water + ninhydrin + [Ni(II)–Gly-Tyr]+ and

he respective cmc values are:

(a) CTAB (× 103): 0.91, 0.89, 0.72 and 0.66 (at 30 ◦C); 1.37, 1.28, 1.38and 1.07 mol dm−3 (at 80 ◦C).

b) 16-6-16 (× 103): 0.043, 0.039, 0.045 and 0.038 (at 30 ◦C); 0.058,0.060, 0.050 and 0.059 mol dm−3 (at 80 ◦C).

(c) 16-5-16 (× 103): 0.034, 0.033, 0.046 and 0.043 (at 30 ◦C); 0.055,0.053, 0.044 and 0.056 mol dm−3 (at 80 ◦C).

d) 16-4-16 (× 103): 0.032, 0.031, 0.034 and 0.041 (at 30 ◦C); 0.043,0.044, 0.048 and 0.049 mol dm−3 (at 80 ◦C).

. Results and discussion

.1. Dependence of reaction rate on pH

The preliminary studies of [Ni(II)–Gly-Tyr]+

3.0 × 10−4 mol dm−3) and [ninhydrin] (6.0 × 10−3 mol dm−3)nteraction were made in the pH range 4.0–6.0 in aqueousnd in micellar media ([CTAB] = 30 × 10−3 mol dm−3 or [gemi-is] = 30 × 10−5 mol dm−3) at 80 ◦C. It was found that the value

f rate constant increased sharply in pH up to 5.0 and thereafterecame almost constant in both the media. Consequently, all theubsequent kinetic measurements were made at pH 5.0.

Scheme 1.

3.2. Dependence of reaction rate on [Ni(II)–Gly-Tyr]+

To see the effect of [Ni(II)–Gly-Tyr]+ complex on the reactionrate, the kinetic runs were carried out with different concentra-tions varying from 2.0 × 10−4 mol dm−3 to 4.0 × 10−4 mol dm−3

at fixed [ninhydrin] (6.0 × 10−3 mol dm−3), temperature(80 ◦C) and pH (5.0) in the absence and presence of micelles([CTAB] = 30 × 10−3 mol dm−3 or [geminis] = 30 × 10−5 mol dm−3)(Table 1). It was found that the order of the reaction with respect to[Ni(II)–Gly-Tyr]+ complex is unity in the two systems (i.e., aqueousand micellar media). Therefore, the rate law is given as Eq. (1):

rate = d[P]dt

= (kobs or k� ) [Ni(II)–Gly-Tyr]+ (1)

3.3. Dependence of reaction rate on [ninhydrin]

The plots of rate constants vs. [ninhydrin] of experiments car-ried out with different [ninhydrin] (range: 6–40 × 10−3 mol dm−3)

perature (80 ◦C) and pH (5.0) in aqueous and in micellar media([CTAB] = 30 × 10−3 mol dm−3 and [geminis] = 30 × 10−5 mol dm−3)(Table 1) are non-linear passing through the origin. This confirms

Mohd. Akram et al. / Colloids and Surfaces B: Biointerfaces 94 (2012) 220– 225 223

Table 1Dependence of pseudo-first-order rate constants (kobs or k�) on [Ni(II)–Gly-Tyr]+, [ninhydrin] and temperature for the reaction of [Ni(II)–Gly-Tyr]+ with ninhydrin at pH 5.0.

104 [Ni(II)–Gly-Tyr]+ (mol dm−3) 103 [ninhydrin] (mol dm−3) Temp. (◦C) 105 kobs (s−1) 105 k� (s−1)

Aqueous CTABa 16-6-16 16-5-16 16-4-16b

2.0 6 80 10.6 43.3 42.7 43.2 44.22.5 10.9 43.0 42.6 43.3 44.33.0 10.6 42.9 42.6 43.3 44.53.5 10.9 42.7 42.6 43.4 44.44.0 10.7 42.8 42.7 43.4 44.53.0 6 80 10.6 42.9 42.6 43.3 44.5

10 20.1 54.0 61.0 80.2 83.015 31.8 77.5 77.0 93.0 97.120 41.5 98.0 85.1 100.8 109.325 51.0 107.2 92.2 110.1 115.030 60.5 136.0 97.0 115.4 120.135 70.0 153.5 102.3 120.1 124.540 75.6 160.1 105.0 122.7 127.9

3.0 6 70 4.8 29.0 34.6 35.1 38.475 9.5 34.5 35.7 39.8 40.780 10.6 42.9 42.6 43.3 44.585 21.6 56.1 46.6 55.3 55.690 31.9 60.9 50.1 60.5 64.3

ti

3

si(a(rtt

4

oam

TDw

a [CTAB] = 30 × 10−3 mol dm−3.b [16-s-16] = 30 × 10−5 mol dm−3.

hat the order of reaction with respect to [ninhydrin] is fractionaln both the media.

.4. Dependence of reaction rate on temperature

To see the dependence of reaction rate on temperature, aeries of kinetic runs were made at different temperatures rang-ng from 70 ◦C to 90 ◦C in aqueous as well as in micellar media[CTAB] = 30 × 10−3 mol dm−3 and [geminis] = 30 × 10−5 mol dm−3)t constant [ninhydrin] (6.0 × 10−3 mol dm−3), [Ni(II)–Gly-Tyr]+

3.0 × 10−4 mol dm−3) and pH (5.0) (Table 1). Using variation of theate constants with temperature, the values of activation parame-ers were evaluated by employing a linear least squares regressionechnique.

. Reaction in absence of micelles

Results of the kinetic studies carried out under pseudo-first-rder conditions of excess [ninhydrin] over [Ni(II)–Gly-Tyr]+ inqueous medium are given in Table 1. It has earlier been found thatetal peptide–ninhydrin reaction proceeds through formation of

able 2ependence of reaction rate on concentration of surfactants for reaction of ninhydrin (6.0ith calculated values (k� cal) at 80 ◦C and pH 5.0.

103 [CTAB] (mol dm−3) CTAB 105 [16-s-16] (mol dm−3) 16-6-

105 k� (s−1) 105 k� cal (s−1) 105 k

0 10.6 – 0 10.6

5.0 33.9 36.9 5.0 11.3

10.0 38.5 37.8 10.0 18.8

15.0 40.7 42.9 20.0 31.6

20.0 41.7 44.6 30.0 42.6

25.0 42.6 41.7 40.0 45.0

30.0 42.9 45.0 50.0 46.8

35.0 42.6 40.4 60.0 48.8

40.0 42.4 41.4 80.0 53.6

45.0 42.2 41.8 100.0 57.7

50.0 42.0 42.4 250.0 63.0

55.0 41.6 40.3 400.0 65.0

60.0 41.2 37.6 600.0 68.0

65.0 40.8 38.9 1000.0 71.0

70.0 40.0 38.3 1500.0 75.1

75.0 39.8 39.3 2000.0 79.2

80.0 39.2 36.4 2500.0 85.0

3000.0 96.0

a ternary labile complex in which both (the peptide and ninhy-drin) are connected to the same metal ion (template mechanism[20]). The presence of metal ion brings the reactive groups togetherwhich provides much better chance for their combination withinits coordination sphere. On the basis of the above, it can be con-cluded that the reaction occurs in two kinetically distinguishablesteps (Scheme 1). Firstly, a fairly rapid ternary labile complex for-mation between ninhydrin and [Ni(II)–Gly-Tyr]+ occurs and then, aslower condensation of amino group to carbonyl group takes place.Therefore, the rate equation is given as:

d[P]dt

= kK[N][A]1 + K[N]

(2)

On comparing with Eq. (1), Eq. (2) can be written as Eq. (3):

kobs = kK[N]1 + K[N]

(3)

([N] is the total concentration of ninhydrin) which can be re-arranged as:

1kobs

= 1k

+ 1kK[N]

(4)

× 10−3 mol dm−3) with [Ni(II)–Gly-Tyr]+ (3 × 10−4 mol dm−3) and their comparison

16 16-5-16 16-4-16

� (s−1) 105 k� cal (s−1) 105 k� (s−1) 105 k� cal (s−1) 105 k� (s−1) 105 k� cal (s−1)

– 10.6 – 10.6 –– 13.8 – 14.2 –

17.6 22.7 18.7 22.8 17.230.8 37.4 38.3 37.0 36.141.6 43.3 42.7 44.5 43.843.7 52.6 50.1 55.2 54.548.4 56.3 55.4 57.8 59.348.3 60.4 60.5 61.1 59.754.7 64.9 65.4 68.6 70.256.7 68.2 70.4 71.9 73.964.3 72.3 71.7 75.0 77.869.6 75.0 77.4 78.7 84.170.1 76.7 77.3 82.2 85.1

– 78.3 – 87.6 –– 82.0 – 93.2 –– 88.0 – 105.1 –– 95.0 – 115.3 –– 108.0 – 133.0 –

224 Mohd. Akram et al. / Colloids and Surfaces B: Biointerfaces 94 (2012) 220– 225

3000200010008060402000

20

40

60

80

100

120

140

100806040200

10

15

20

25

30

35

40

45

105k ψ

(s-1

)

103 [CTAB] (mol dm

-3)

III

II

I

c

b

a

10

5k ψ

(s

-1)

105 [16 -s-16] (mol dm

-3)

Fig. 3. Dependence of reaction rate constant (k�) on [gemini] for the reac-tion of ninhydrin with [Ni(II)–Gly-Tyr]+ in presence of surfactants in acetatebuffer solution (pH = 5.0) in (a) 16-6-16, (b) 16-5-16, (c) 16-4-16, and (Inset)f[

Atacat

5

icT(4arbbrtosctp[I

fma

tpa

+

Nw + Dn NDn

+KN

KSA w + Dn ADn

with the s = 4 gemini) with a resultant decrease in k . Thus the

or variation of [CTAB]. Reaction conditions: [ninhydrin] = 6.0 × 10−3 mol dm−3,Ni(II)–Gly-Tyr]+ = 3.0 × 10−4 mol dm−3, temperature = 80 ◦C.

ccordingly, a plot of 1/kobs vs. 1/[N] yielded straight line with posi-ive intercept (1/k) and slope (1/kK). The values of k (2.9 × 10−3 s−1)nd K (9.0 mol−1 dm3) were thus evaluated in aqueous medium. Thealculated values of rate constants (kcal), obtained by substituting knd K in Eq. (3), are in close agreement with the kobs, which supportshe proposed mechanism and confirms the validity of Eq. (3).

. Reaction in presence of surfactant micelles

The effect of surfactants on the reaction rate was exam-ned by varying the amount of CTAB/gemini at fixed reactiononditions (i.e., [ninhydrin] = 6.0 × 10−3 mol dm−3, [Ni(II)–Gly-yr]+ = 3.0 × 10−4 mol dm−3, temperature = 80 ◦C and pH = 5.0)Table 2). The values of rate constants (k�) increased from 10.6 to2.9 × 10−5 s−1, on increasing [CTAB] from 0 to 30 × 10−3 mol dm−3

nd further increment in [CTAB] had a decreasing effect on theate (Fig. 3, Inset). The plot of k� vs. [CTAB] is perfectly generaleing common characteristic of bimolecular reactions catalyzedy micelles [21–25]. A detailed investigation revealed that theeaction follows first- and fractional-order kinetics with respecto [Ni(II)–Gly-Tyr]+ and [ninhydrin], respectively. Thus, the orderf reaction with respect to [Ni(II)–Gly-Tyr]+ and [ninhydrin] is theame as that in aqueous medium. With the geminis, however, rateonstant (k�) first increases (Fig. 3, part I), remains constant upo certain concentration (part II) (the characteristics of part I andart II are just like the monomeric conventional counterpart CTAB)21–25], and then increases at higher concentrations sharply (partII).

The value of rate (k�) should remain constant in part I as [sur-actant] is lower than the cmc values. The observed catalytic effect

ay, therefore, be due to preponement of micellization by reactantsnd/or presence of premicelles [26].

The observed enhancement in the reaction rate is quantitatively

reated on the basis of the pseudo-phase model (Scheme 2), pro-osed by Menger and Portnoy [27], and developed by Romsted [28]nd Bunton [23].

Productsk'mk'w

Scheme 2.

Although several kinetic equations based on this generalScheme have been developed, the most successful appears to bethat of Romsted [28] who suggested an expression (Eq. (5)), whichtakes into account the solubilization of both the reactants into themicelles as well as the mass action model.

k� = kw[N] + (KSkm − kw) MSN [Dn]

1 + KS[Dn](5)

In the above equation, kw and km are the second-order rate con-stants, referring to bulk and micellar pseudo-phases, respectively,KS and KN are the binding constants of the [Ni(II)–Gly-Tyr]+ com-plex and ninhydrin to the cationic micelles, MS

N being the molarityof ninhydrin bound to the micellar headgroups, [Dn] represents themicellized surfactant (=[surfactant] − cmc), and NDn and ADn aremicellized ninhydrin and [Ni(II)–Gly-Tyr]+ complex, respectively.In order to find out KS and km, the non-linear least squares tech-nique was used for Eq. (5). This process gave the value of leastsquares, i.e., ˙d2

i(di = k� obsi − k� cali) at ith KS. The calculation was

repeated for different values of KS and the best value was the onefor which ˙d2

iwas minimum. The KS, thus obtained, was used to

obtain the best value of km.Most of the ionic micelle mediated reactions are believed to

occur either inside the Stern layer or at the interface between micel-lar surface and bulk water solvent [25]. From purely electrostaticconsiderations, ninhydrin (due to the presence of an electron cloud[29]) can be assumed to reside predominantly in the Stern layer.The micellar surface can attract or repel ionic species due to elec-trostatic interactions whereas hydrophobic interactions can bringabout the incorporation of reactants into micelles. The observedcatalysis is due to increased concentration of both ninhydrin and[Ni(II)–Gly-Tyr]+ complex in the Stern layer. The decrease in rateconstant beyond [CTAB] > 30 × 10−3 mol dm−3 was slow. It couldbe due to: (i) competition between the bromide and ninhydrin onthe micellar surface, and/or (ii) segregation of the reactants in thedifferent micelles as they are diluted in an increasing number ofaggregates.

Gemini micelles provide much better environment for theninhydrin-[Ni(II)–Gly-Tyr]+ reaction as compared to their anal-ogous counterpart (CTAB micelles). The reason for that is thepresence of spacer in the geminis which decreases the water con-tent in the aggregates making the environment less polar and thuscausing rate increase (see Scheme 1). Menger et al. [30] have opinedthat due to proximity of positive charges in gemini micelles anionbinding at surfaces is increased at the expense of binding of H2O.The behavior in part II is same for all the three geminis but val-ues of k� at all concentrations are in the order: s = 4 > 5 > 6 (Fig. 3).This is not for the first time but best results with 16-4-16 wereobtained earlier also [14]. It is well known that, to minimize itscontact with water, a spacer longer than the ‘equilibrium’ dis-tance between two-N+Me2 head groups (the ‘equilibrium’ distanceoccurs at s = 4 in 16-s-16 geminis [18,31]) tends to loop towardsthe micellar interior. This increased looping of the spacer (for s > 4)will progressively make the Stern layer more wet (in comparison

�

results are consonant with the earlier findings that increase in thewater content of the reaction environment has an inhibiting effect[32–35].

Mohd. Akram et al. / Colloids and Surfaces B: Biointerfaces 94 (2012) 220– 225 225

Table 3Thermodynamic parameters, rate and binding constant values for the reaction of [Ni(II)–Gly-Tyr]+ and ninhydrin in presence of CTAB and gemini surfactants at pH 5.0.

Parameters and constants Aqueous CTAB 16-6-16 16-5-16 16-4-16

Ea (kJ mol−1) 140.1 36.2 27.8 22.3 19.6�H#(kJ mol−1) 137.2 33.3 24.9 19.4 16.6−�S# (JK−1 mol−1) 302.0 308.3 308.8 309.0 309.2102 km (s−1)a – 1.4 3.4 3.9 3.7

−1 3 a 0

5

btsroeflfsCttmmdaisi[[ried

6

memmoTawcTlvan

7

cTtma

[[[

[[[[[[[[[[[[

[

[

[[

[[

[

[[[[

KS (mol dm ) – 24.KN (mol−1 dm3)a – 48.

a At 80 ◦C.

After leveling-off, further increase at higher [geminis] is proba-ly associated with a change of micellar structure. It is well knownhat in aqueous solution above cmc, the surfactant moleculeself-assemble into aggregates. In general, the aggregates areotund, globular micelles. However, under appropriate conditionsf concentration, salinity, temperature, presence of counterions,tc., spherical micelles can undergo uniaxial growth to formexible worm-like micelles [1]. Further, in case of gemini sur-

actants, the extent of aggregate growth and the variations ofhapes of micelles depend on the spacer chain length(s) as well.ryo-TEM measurements from solutions of 16-s-16 clearly showhe transitions upon increasing s with the sequence of struc-ures of micelles being: vesicles + elongated micelles → elongated

icelles → spheroidal micelles [36]. Thus, the micellar growth isore pronounced the shorter the spacer unit, which is most likely

ue to the increasing geometrical constraints in the formation ofggregates with decreasing length of the spacer unit. Microviscos-ty and SANS data also support the argument that, within the geminiurfactants, micellar morphology tends to be less ellipsoidal withncreasing s [31]. The occurrence of structural changes at highergemini] have been confirmed by 1H NMR spectral studies also19]. Obviously, change in aggregate morphology provides differenteaction microenvironment (less polar) [37]; hence the k�-valuesncrease sharply. This is true for all the three geminis, only thextent of increase in k� (depending upon spacer chain length) isifferent.

. Activation parameters

The variation of activation parameters in CTAB and geminiicelles as compared to aqueous is as expected, because one might

xpect electrostatic attraction between the reactants and cationicicelles to reduce the activation enthalpy when reactants are in theicellar phase. The values of energy of activation (Ea), enthalpy

f activation (�H#) and entropy of activation (�S#) are given inable 3. A comparison with those of aqueous medium values reveals

large decrease in energy of activation and enthalpy of activationith a substantial negative entropy. The negative value of �S# indi-

ates the existence of compact activated state in the micellar phase.he negative entropy of activation is also a characteristic of rateimiting formation of an intermediate complex. In addition, a loweralue of activation energy clearly suggests the catalytic role of CTABnd geminis (catalysts lower the activation energy and provide aew reaction path).

. Conclusions

Kinetic experiments between ninhydrin and nickel-dipeptideomplex were carried out at fixed concentrations of [Ni(II)–Gly-

yr]+ (3.0 × 10−4 mol dm−3), [ninhydrin] (6.0 × 10−3 mol dm−3),emperature (80 ◦C) and pH 5.0 at �max = 400 nm spectrophoto-

etrically in aqueous and in presence of micellar media (CTABnd geminis). While presence of CTAB micelles do catalyze the

[[

130.0 120.0 96.070.4 66.9 75.9

reaction, the dicationic gemini surfactant micellar media are moreeffective to accelerate the reaction. Further, the enhancement bygeminis occurs at concentrations ca.100 times less than CTAB. Thus,whereas the sensitivity of the ninhydrin reaction stands increased,use of a very small amount of 16-s-16 gemini surfactants suggestsan increased sensitivity and a less environmental impact too.

Acknowledgements

Authors are thankful to UGC (India) for financial assistance (toDK) and Emeritus Fellowship (to KU).

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