Interfacial activation

Normally, reactions take place at the oil-water interface by the movement of the lid allowing the substrate to access the active sites. This is a special phenomenon in lipases and is referred to as the interfacial activation. This distinguishes lipases from esterases. This phenomenon is often associated with the reorientation of the a-helical lid structure that in turn increases the surface hydrophobicity in the vicinity of the active site and subsequently exposes the site (Cleasby et al., 1992). Usually, substrates form an equilibrium between the monomeric, micellar, and emulsified states, which require a suitable model in order to study lipase kinetics. Sarda and Desnuelle (1958) documented the fact that interfacial activation is an enhancement of lipase activity and that lipase works effectively on the two-dimensional surface of the micelle. They proposed that lipase becomes activated 

Supercritical Fluids Technology in Lipase Catalyzed Processes 

This interfacial activation does not appear in all hydrolytic enzymes. For example, cutinase from fusarium solani, which hydrolyzes cutin, usually has the fatty acids -Ci6 and -C,8 and does not have a lid and does not exhibit these characteristics (Carvalho et al 1998 Schrag et al 1997). In addition, the presence of a lid does not always cause the enzyme to be active at the interface. The exception is lipase B from Candida antarctica that does not show interfacial activation despite having a lid (Kuo and Gardner, 2002 Uppenberg et al., 1994). Additionally, Staphylococcus hyicus and P. aeruginosa show an interfacial activation with some substrates but not with others (Verger, 1997). 

X-ray studies of the 3D structure of lipase confirmed the existence of only one domain, except in pancreatic lipases where there were two distinct domains a large N-terminal domain (amino-terminal domain) and a smaller C-terminal domain. The large N-terminal domain is a typical hydrolase that contains an active site with a catalytic triad formed by Ser, Asp, and His (van Tilbeurgh et al., 1992). 

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Figure 20 shows the plot of the surface tension vs. the logarithm of the concentration (or-lg c-isotherms) of sodium alkanesulfonates C,0-C15 at 45°C. In accordance with the general behavior of surfactants, the interfacial activity increases with growing chain length. The critical micelle concentration (cM) is shifted to lower concentration values. The typical surface tension at cM is between 38 and 33 mN/m. The ammonium alkanesulfonates show similar behavior, though their solubility is much better. The impact of the counterions is twofold First, a more polarizable counterion lowers the cM value (Fig. 21), while the aggregation number of the micelles rises. Second, polarizable and hydrophobic counterions, such as n-propyl- or isopropylammonium and n-butylammonium ions, enhance the interfacial activity as well (Fig. 22). Hydrophilic counterions such as 2-hydroxyethylammonium have the opposite effect. Table 14 summarizes some data for the dodecane 1-sulfonates.

The interfacial activity is determined by the sterical properties of the molecule. At the interface the spatial demand A0 of the hydrophobic part of the molecule is higher because of the second chain of the internal sulfonate compared with the terminal sulfonate. Thus, the surface concentration of the surfactant molecules is lower. That means that the hydrocarbon chains are laterally oriented and therefore cover the interface between the solution surface and air more completely. Because the ratio of the spatial demand of the head group to the volume of the alkyl chain governs the radius of the micellar surface, it... 

For long-chain alcohol esters it is interesting to see that the interfacial tension between a 0.01 wt % aqueous solution and octane or xylene has a minimum for ester sulfonates with a total 22 carbon atoms in the fatty acid chain and the ester chain [60]. The balance in length between the two chains has only a poor effect. Thus, a-sulfonated fatty acid esters with a total number of 22-26 carbon atoms in the molecule have excellent interfacial activities. To attain the same magnitude in the interfacial tension between linear alkylbenzenesulfonate (LAS) solution and octane, the required concentration of LAS is 0.1 wt %. This is 10 times the concentration needed for a-sulfonated fatty acid esters [60]. 

Hydrolysis of substrates is performed in water, buffered aqueous solutions or biphasic mixtures of water and an organic solvent. Hydrolases tolerate low levels of polar organic solvents such as DMSO, DMF, and acetone in aqueous media. These cosolvents help to dissolve hydrophobic substrates. Although most hydrolases require soluble substrates, lipases display weak activity on soluble compounds in aqueous solutions. Their activity markedly increases when the substrate reaches the critical micellar concentration where it forms a second phase. This interfacial activation at the lipid-water interface has been explained by the presence of a... 

Cutinase is a hydrolytic enzyme that degrades cutin, the cuticular polymer of higher plants [4], Unlike the oflier lipolytic enzymes, such lipases and esterases, cutinase does not require interfacial activation for substrate binding and activity. Cutinases have been largely exploited for esterification and transesterification in chemical synthesis [5] and have also been applied in laundry or dishwashing detergent [6]. 

In 1958 Sarda and Desnuelle [79] discovered the lipase activation at the interfaces. They observed that porcine pancreatic lipase in aqueous solution was activated some 10-fold at hydrophobic interfaces which were created by poorly water-soluble substrates. An artificial interface created in the presence of organic solvent can also increase the activity of the lipase. This interfacial activation was hypothesized to be due to a dehydration of the ester substrate at the interface [80], or enzyme conformational change resulting from the adsorption of the lipase onto a hydrophobic interface [42,81,82]. 

The weight percentage breakdown of fractions and subfractions obtained from fractionation of both the crude oil and shale oil samples are shown in Figure 3 and 4, respectively. The percentage recoveries of Fraction HI from the crude oU and shale oil samples were 16.5% and 24.1%, respectively. To investigate the interfacial activity of these subfractions upon reaction with alkali, IFT measurements were carried out with a 1% solution of each fraction in toluene against aqueous... 

Seifert, W.K. Howells, W.G. Interfacially Active Acids in a California Crude Oil, Anal. Chem. 1969, 41, 554. 

Chan, M. Interfacial Activity in Alkaline Flooding Enhanced Oil Recovery, Ph.D. Thesis, USC, Los Angeles, 1980. 

Wisniewski, M. Jakubiak, A. Szymanowski, J. Interfacial activity and rate of palladium(II) extraction with decyl pyridine monocarboxylates. J. Chem. Technol. Biotechnol. 1995, 63, 209-214. 

More advanced semiempirical molecular orbital methods have also been used in this respect in modeling, e.g., the structure of a diphosphonium extractant in the gas phase, and then the percentage extraction of zinc ion-pair complexes was correlated with the calculated energy of association of the ion pairs [29]. Semiempirical SCF calculations, used to study structure, conformational changes and hydration of hydroxyoximes as extractants of copper, appeared helpful in interpreting their interfacial activity and the rate of extraction [30]. Similar (PM3, ZINDO) methods were also used to model the structure of some commercial extractants (pyridine dicarboxylates, pyridyloctanoates, jS-diketones, hydroxyoximes), as well as the effects of their hydration and association with modifiers (alcohols, )S-diketones) on their thermodynamic and interfacial activity [31 33]. In addition, the structure of copper complexes with these extractants was calculated [32]. 

The observed rates of transfer are lower than those calculated by the correlation of Eq. 26 for organic molecules which themselves are surface-active, without specifically added long-chain molecules thus in the transference of (C4H9)4NI from water to nitrobenzene, of benzoic acid from toluene to water and the reverse, of diethylamine between butyl acetate and water, of n-butanol from water to benzene, and of propionic acid between toluene and water, the rates (44, 4 ) are of the order one-quarter to one-half those calculated by Eqs. (25) and (26). Since with these systems the solute itself is interfacially active, and therefore its monolayers should reduce the transfer of momentum, we interpret these findings as indicative that Ri and R2 are increased in this way. This is... 

Because the size of the emulsion droplets dictates the diameter of the resulting capsules, it is possible to use miniemulsions to make nanocapsules. To cite a recent example, Carlos Co and his group developed relatively monodisperse 200-nm capsules by interfacial free-radical polymerization (Scott et al. 2005). Dibutyl maleate in hexadecane was dispersed in a miniemulsion of poly(ethylene glycol)-1000 (PEG-1000) divinyl ether in an aqueous phase. They generated the miniemulsion by sonication and used an interfacially active initiator, 2,2 -azobis(A-octyl-2-methyl-propionamidine) dihydrochloride, to initiate the reaction, coupled with UV irradiation. 

Sander and Henze [50] have performed ac investigations of the adsorption potential of metal complexes at Hg electrode. Later, Sander etal. [51] have studied electrosorption of chromium - diethylenetriaminepentaacetic acid (DTPA) on mercury in 0.1 M acetate buffer at pH 6.2 using a drop-time method. The changes in the interfacial activity of the Cr(III)-DTPA complex with the bulk concentration obeyed the Frumkin adsorption isotherm. 

Lobacz et al. [52] have described partial adsorption ofTl+-cryptand (2,2,2) complex on mercury electrode. From voltocoulom-etry, cyclic voltammetry, and chrono-coulometry, it has been deduced that electroreduction of this complex proceeds via two parallel pathways from the solution and from the adsorbed states, which are energetically close. Also, Damaskin and coworkers [53] have studied adsorption of the complexes of alkali metal cations with cryptand (2,2,2) using differential capacity measurements and a stationary drop electrode. It has been found that these complexes exhibit strong adsorption properties. Novotny etal. [54] have studied interfacial activity and adsorptive accumulation of U02 " "-cupferron and UO2 - chloranilic acid complexes on mercury electrodes at various potentials in 0.1 M acetate buffer of pH 4.6 and 0.1 M NaCl04, respectively. 

In Ref 169, some peculiarities associated with adsorption of alkyne peroxides from DM F-water solutions onto the mercury electrode in the presence of tetraethylammonium cations have been described. Polarography and electrocapillary measurements were employed as the experimental techniques. It has been shown that interfacial activity of these peroxides was determined by the species generated as a result of associative interactions between peroxides and DMF and tetraethylammonium cations. 

Zampieri et al. [ 149], in order to circumvent the inherent problems of the earlier sedimentation studies, employed two different dyes (one water soluble and the other strong interfacially active) to monitor the association of water and surfactant with empty and filled RMs independently. They were able to estimate the sizes of filled and empty RMs based on water, protein, and surfactant balances by determining the individual Wg values for the two types of RMs. The conclusions arrived at were in sharp contrast to those of Levashov et al. [148], as it was shown that both the filled and empty RMs increased in size with the overall Wg and that neither the filled nor the empty RM size was the same after protein uptake. An assumption made by Zampieri et al. [149] is that the two dyes distributed between the RMs in proportion to water and surfactant, respectively. Hatton s group [152] suggested that this assumption may not be true based on their analyses of the substrate distribution effects and suggested that the statistical distribution of solutes over the micelle population may be skewed to one or the other of two types of RMs. 

In case of lipases, one of the simplest methods to combine an enzyme with an organic solvent is to coat the lipase with a lipid or surfactant layer before lyophilisation. It is estimated that about 150 surfactant molecules are sufficient for encapsulating one lipase molecule. Following this route the surfactant coated lipase forms reverse micelles with a minimum of water concentration. The modified lipases are soluble in most organic solvents, and the reaction rates are increased compared to the suspended hpases due to the interfacial activation [59,60]. 

The influence of surfactant on the catalytic activity of lipases in water is well known. The addition of surfactants can enhance the activity and enantioselectivity of these enzymes in aqueous solutions [94] due to the interfacial activation and due to the emulsification of hydrophobic substrates. 

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