Enzymatic erosion

Cuticular erosion results in the formation of imprints on the leaf surface that can easily be detected by scanning electron microscopy (A2.) Comparison of Erysiohe graminis with . cichoracearum demonstrated that . graminis erodes both barley and cucumber cuticles, but that 21. cichoracearum only eroded or left imprints on cucumber cuticles (A3.). These results suggest a level of host specificity, possibly for the enzymatic erosion of the cuticle, that may also be involved in the successful penetration of the leaf. 

Fig. 4.41 Contact mode AFM height images (the z-scale covers 200 nm) of PTMC film during enzymatic erosion. The images were captured between 80 and 340s and 340 and 600s after start of the hydrolysis, respectively. The arrows indicate the scan direction. Reproduced with permission from [71]. Copyright 2005. American Chemical Society...

Figure 16 Relation between the rate of enzymatic erosion and the degree of crystallinity (a) and the lamellar thickness (b) for melt-crystallized PHA films. Reproduced with permission from Abe, H. Doi, Y. Aoki, H. Akehata, T. Macromolecules31,1791. Copyright 1990 American Chemical Society.

Since the PHA depolymerases have ndo-hydrolase activity, as mentioned above, the random copolymers with 3HB unit as a major constituent are degradable by PHA depolymerase even though enzymatically inactive monomeric units are introduced. The effects of chemical structure of second monomer units and copolymer compositions on the rate of enzymatic erosion have been examined through the enzymatic degradation of solution-cast films of random copolymers of 3HB with various HA units in the presence of PHA depolymerase. " The enzymatic degradation of solution-cast films of these PHA copolymers was performed in an aqueous solution of purified PHA depolymerase from R. pickettii T1 at 37 °C. The rate of enzymatic erosion of solution-cast PHA films increased markedly with an increase in the fraction of second monomer units up to 10-20 mol.% to reach a maximum value followed by a decrease in the erosion rate (Figure 18). The highest rates of enzymatic erosion were 5-10 times that of the P(3HB) homopolymer film. 

Tanaka et al. and Iwata et performed enzymatic degradation of highly oriented P(3HB) monofilaments, in which both the 2/1 helix conformation (a-form) and planar zigzag conformation (p-form) existed. They demonstrated that the enzymatic erosion rate of the p-form is faster than the a-form for P(3HB) fibers. This result can also be explained in terms of conformational matching of the polymer chain with the active site of the enzyme. 

Pig. 3 Scanning electron micrographs of the PHB films crystrdlized at 120°C for 168 h from the melt before and after enzymatic erosion (Koyama and Doi 1997)... 

The degree of crystallinity of solvent cast copolymer films decrease from 60 to 18% when the fraction of 3-hydroxyhexanoic acid is increased from 0 to 25 niol-%. This finding suggests that the 3-hydroxyhexanoic acid units are excluded from the crystalline phase of 3-hydroxybutyric acid. The rates of enzymatic erosion increase markedly with the increase of 3-hydroxyhexanoic acid the... 

It is necessary to consider enzymatic erosion in any detailed discussion of this topic. This process takes place in two steps firstly, amide penetration through the membrane (transported by water) and pore walls secondly, the enzymatic reaction occurs and the amide reacts with the enzyme trapped within the matrix, producing maltose. The characteristic duration of this process is generally the sum of the penetration time (Tp) and the reaction time (Tj.). Therefore, because the reaction velocity (Pg) is proportional to the amide concentration Cg, the following is obtained ... 

Figure 3 Rates of enzymatic erosion of various PHA copolymer films in the aqueous solution of PHA depolymerase (from Akaligenes faecalis) at 37 C and pH 7.4. ( ) P(3HB-co-3HH), ( ) P(3HB-co-3HV), (o) P(3HB-c

Furthermore, it was reported that among the semicrystalline PLLA films with similar crystallinity, the films annealed at a higher temperature were more resistant to enzymatic degradation [14]. It is suggested that the lamellar crystal size of PLLA also plays a decisive role in the enzymatic degradation process. A similar dependence of the enzymatic erosion rate of semicrystalline PLLA on crystal size has been confirmed by Kurokawa et al. [28]... 

In addition to PLA stereocopolymers, PLA films with different stereochemical composition can be obtained by blending of PLA stereoisomers. The enzymatic degradability of both the enantiomeric blend films from PLLA and PDLA and the diastereoisomeric blend films of PDLLA with either PLLA or PDLA has been investigated by Tsuji and Miyau-chi [12]. It is of interest to note that the erosion rates of amorphous films show similar trends with respect to the overall Xlla between enantiomeric and diastereoisomeric blends (Figure 22.6). The enzymatic erosion rate of both blend films decreases with decreasing Xlla and approaches zero at LLA below 0.3, which is very similar to the result for the nonblended PLA films. Since the PLLA and PDLLA components are enzymatically hydrolyzable as opposed to PDLA, the content of hydrolyzable components in the blends with identical Xj t a values are apparently different between enantiomeric and diastereoisomeric blends. Therefore, such similarity in trends... 

To study the enzymatic degradation of the free amorphous region, completely amorphous PLLA thin film was prepared (see Section 22.3.2), and the erosion rate was directly monitored by using QCM in the nanogram per square centimeter regime [74]. The enzymatic erosion rate was dependent on the concentration of proteinase K, and the thin amorphous film of lOOnm thickness was completely hydrolyzed in 20 min when the concentration of the enzyme was >100 qg/mL. During the course of enzymatic degradation, even if the enzyme solution was replaced with a buffer solution (i.e.. 

FIGURE 22.15 Dependence of enzymatic erosion rate ( ) and adsorbed enzymes (O) on the concentration of proteinase K. Erosion rate was measured by QCM, whereas the adsorbed enzymes per 100 X lOOnm PLLA surface after enzymatic treatment for 7 min were measured from AFM images. Reprinted with permission from Ref. 74. Copyright 2005, American Chemical Society. 

The amount of film erosion increased proportionally with time for all the samples. The rate of erosion of solvent-cast film of P(3HB-co-8%-3HV) was 0.50 mg/h/cm and this value is five times faster than that of P(3HB) homopolymer solvent-cast film (0.10 mg/h/cm ). On the other hand, the rates of erosion of the 10 times and 16 times cold-drawn films were 0.38 and 0.32 mg/h/cm respectively, suggesting the effect of the crystallinity and long period on the rate of erosion. We reported the effect of crystallinity and solid state structure on enzymatic erosion of P(3HB) stretched films. In the case of P(3HB-co-8%-3HV), the enzymatic erosion rate seems to be strongly affected by the crystallinity and l[Pg.56]

---