

**143**

*The Potential Application of Nanoparticles on Grains during Storage: Part 2 – An Overview…*

of chitosan and its derivatives creates an electric field for an electrostatic interac

tion between the polycationic structure and the anionic components of the cell (i.e., lipopolysaccharide and cell surface proteins), thus altering cell permeability [118–123]. High pH enhance rapid protonation, which increase the positive charge density (polycationic activity) of chitosan. A positive correlation was established between charge density and the biocidal activity of quaternized chitosan [124–127]. The inhibition potential of chitosan could be incapacitated when the charge density is reduced [120] due to changes of pH values. A similar outcome was reported by Qin et al. [128]. The antimicrobial mechanism was associated with the interac

tion of the negatively charged cell membranes and the cationic NH3+groups of the chitosan derivative, which increase membrane permeability resulting in lysis [129] and leakage of macromolecules killing the cells. A carboxyfluorescein (CF)-loaded liposome study showed the effectiveness of lower molecular weight (LMW) chi

mellar vesicles [130]. Similarly, Ing et al. [131] reported that chitosan NPs prepared

and MICHMW = 0.5–1.2 mg/mL) compared to the solution form (MIC = 3 mg/mL for both MWs and species). The authors established a statistical linear relation

ship between MW and particle size/zeta potential, thus provided an avenue for the manipulation of physicochemical properties of NPs to maximize its ability to penetrate the cells, trigger leakage of intracellular component, eventually killing the

Researchers [132–135] proposed the fundamental mechanism contributing to interaction of negatively charged surface components of fungi and bacteria with the positively charged NH3+ groups of glucosamine (chitosan), which alters cell surface, and trigger leaking of intracellular substances, resulting in the impairment of vital physiological activities thus killing the microorganism. The inability of the second amino groups on N-acetylation of chitosan oligomers to donate positive chargere

sult in the inhibition of its fungistatic activity [136]. Therefore, the contribution of NH3+ groups to biocidal activity cannot be ignored and should carefully be consid

The outer membrane (OM), inner core of lipopolysaccharide (LPS) molecules,

phosphate and carboxyl, which contribute to the hydrophilic nature of the cell wall, thus creatin interaction of charges (electrostatic) with divalent cations.

advantage over Gram(+) bacteria. Therefore, breaching the integrity of the OM by

LMW chitosan showed higher efficiency perforate/penetrate the microbial cell compared to HMW chitosan, which interacts with DNA to change the translation and transcription profile of genes. Chitosan binds to DNA with accurate precision, denying the organism of normal DNA transcription and mRNA synthesis, resulting

On the other hand peptidoglycan (PG) and teichoic acid (TA) on the cell wall of Gram(+) bacteria have polyanionic group, which facilitates interaction via covalent bond with N-acetylmuramic acid in the PG layer, or via glycolipid- which links outer leaflet of the cytoplasmic membrane [139]. As documented by Kong et al. [120], the poly(glycerol phosphate) anion groups aid the structural stability of cell

compounds (antibiotics and toxic drugs), giving Gram(

chitosan could enhance its biocidal activity toward Gram(

wall in addition to some membrane-bound enzymes.

from different concentrations of LMW and high molecular weight (HMW)

μg/

*F*. *solani* (MICLMW= 0.86–1.2 mg/mL

−) bacteria are composed of anionic groups like

−) bacteria a comparative

−) bacteria [137, 138].

−) bacteria cells from macromolecules and hydrophobic

*C*. *albicans* (MICLMW = 0.25–0.86 mg/

≈7%) leakage of carboxyfluorescein found in the large unila

tosan on the cell membrane. The results showed that 0.75

showed efficient inhibitory activity against

mL and MICHMW = 0.6–1.0 mg/mL) and

fungi and extend safety of the grains.

ered to maximize the effects.

The OM protects Gram (

in cell death [140–142].

and lipid components of Gram(

triggered moderate (








μL of LMW chitosan

*DOI: http://dx.doi.org/10.5772/intechopen.93213*

*Some selected studies on fungicidal activities of chitosan NPs.*

*The Potential Application of Nanoparticles on Grains during Storage: Part 2 – An Overview… DOI: http://dx.doi.org/10.5772/intechopen.93213*

of chitosan and its derivatives creates an electric field for an electrostatic interaction between the polycationic structure and the anionic components of the cell (i.e., lipopolysaccharide and cell surface proteins), thus altering cell permeability [118–123]. High pH enhance rapid protonation, which increase the positive charge density (polycationic activity) of chitosan. A positive correlation was established between charge density and the biocidal activity of quaternized chitosan [124–127]. The inhibition potential of chitosan could be incapacitated when the charge density is reduced [120] due to changes of pH values. A similar outcome was reported by Qin et al. [128]. The antimicrobial mechanism was associated with the interaction of the negatively charged cell membranes and the cationic NH3+groups of the chitosan derivative, which increase membrane permeability resulting in lysis [129] and leakage of macromolecules killing the cells. A carboxyfluorescein (CF)-loaded liposome study showed the effectiveness of lower molecular weight (LMW) chitosan on the cell membrane. The results showed that 0.75 μg/μL of LMW chitosan triggered moderate (≈7%) leakage of carboxyfluorescein found in the large unilamellar vesicles [130]. Similarly, Ing et al. [131] reported that chitosan NPs prepared from different concentrations of LMW and high molecular weight (HMW) showed efficient inhibitory activity against *C*. *albicans* (MICLMW = 0.25–0.86 mg/ mL and MICHMW = 0.6–1.0 mg/mL) and *F*. *solani* (MICLMW= 0.86–1.2 mg/mL and MICHMW = 0.5–1.2 mg/mL) compared to the solution form (MIC = 3 mg/mL for both MWs and species). The authors established a statistical linear relationship between MW and particle size/zeta potential, thus provided an avenue for the manipulation of physicochemical properties of NPs to maximize its ability to penetrate the cells, trigger leakage of intracellular component, eventually killing the fungi and extend safety of the grains.

Researchers [132–135] proposed the fundamental mechanism contributing to interaction of negatively charged surface components of fungi and bacteria with the positively charged NH3+ groups of glucosamine (chitosan), which alters cell surface, and trigger leaking of intracellular substances, resulting in the impairment of vital physiological activities thus killing the microorganism. The inability of the second amino groups on N-acetylation of chitosan oligomers to donate positive chargeresult in the inhibition of its fungistatic activity [136]. Therefore, the contribution of NH3+ groups to biocidal activity cannot be ignored and should carefully be considered to maximize the effects.

The outer membrane (OM), inner core of lipopolysaccharide (LPS) molecules, and lipid components of Gram(−) bacteria are composed of anionic groups like phosphate and carboxyl, which contribute to the hydrophilic nature of the cell wall, thus creatin interaction of charges (electrostatic) with divalent cations. The OM protects Gram (−) bacteria cells from macromolecules and hydrophobic compounds (antibiotics and toxic drugs), giving Gram(−) bacteria a comparative advantage over Gram(+) bacteria. Therefore, breaching the integrity of the OM by chitosan could enhance its biocidal activity toward Gram(−) bacteria [137, 138]. On the other hand peptidoglycan (PG) and teichoic acid (TA) on the cell wall of Gram(+) bacteria have polyanionic group, which facilitates interaction via covalent bond with N-acetylmuramic acid in the PG layer, or via glycolipid- which links outer leaflet of the cytoplasmic membrane [139]. As documented by Kong et al. [120], the poly(glycerol phosphate) anion groups aid the structural stability of cell wall in addition to some membrane-bound enzymes.

LMW chitosan showed higher efficiency perforate/penetrate the microbial cell compared to HMW chitosan, which interacts with DNA to change the translation and transcription profile of genes. Chitosan binds to DNA with accurate precision, denying the organism of normal DNA transcription and mRNA synthesis, resulting in cell death [140–142].

*Mycotoxins and Food Safety*

**142**

**Reference**

[110] [111] [112] [113]

Not reported

100 (MMW) and

*R*. *oryzae* CECT 2340, *A. alternata* CECT 20560,

2088

Molds and total flora isolated from strawberries (*R*. *Stolonifer*

and *B*. *cinerea*)

*R*. *solani* Kuhn*, F*. *oxysporum* (Schl.) *f.* sp. *Cucumernum owen,* 

*C*. *cucumerinum Ell.* Et Arthur, *B*. *cinerea Pers*., *C*. *orbiculare*

(Berk. & Mont.) *Arx*, *P*. *asparagi* (sacc.a) *Bubak*, *A*. *Kikuchiama* 

*Tanaka*, *P*. *italicum Wehmer*, *Fusarium oxysporum Schl. F. Sp.* 

*Uasinfectum* (Atk.) Snyd. & Hans,

*Kuhn., B*. *berengeriana de Not. f.* Sp*., Piricola* (Nose) *Koganezaea*

*et Sakuma*, *Sclerotinia sclerotiorum (Lib.) de Bary*, *Venturia* 

*nashicola Tanaka et Yamamoto*, *Gibberella zeae (Schw.) Petch* and

*Phytophthora infestabs* (Mont.)

*A*. *niger* *A*. *alternata f*. sp. *lycopersici*

0.1% or 1% (w/v)

100–6400 μg/mL

Coatings, films,

and liquid

Solutions

[114] [115]

**Table 2.**

*Some selected studies on fungicidal activities of chitosan NPs.*

Shrimp shell

Not reported

Not reported

90

*V*. *ctahliae Kleb., R*. *sclani* 

*A*. *niger* CECT

6 mL Final concentration before the

spraying was 0.02%, w/v

20, 30, 50, 100, and 150 mg/L

Solutions

97 (LMW)

85–89

(industrially made)

Not reported (industrial

made)

Not reported

80%

**Sources of chitosan** 

**Deacetylation** 

**Microorganisms**

*R*. *stolonifera*, *E*. *coli* DH5α strain

**Concentration**

1.0 mL

Coatings and

solutions

Films and

solutions

**Form applied**

**(%)**

75-85

**(CTS)**

Not reported

A decrease in the induction of β-galactosidase was observed when yeast cells were exposed to chitosan. A concentration of 0.35 mg/mL chitosan reduced β-galactosidase activity by 32%. An increased in concentration (1.25 mg/mL) further led to the reduction of enzyme activity. The control experiment did not follow the trend. Likewise, the treated cells showed that chitosan greatly influenced protein biosynthesis in the yeast [130]. Previous work [143] documented cell sensitivity to chitosan, which altered the deletions of genes involved in sphingolipid (e.g., *ipt1Δ*, *skn1Δ*, *lcb3Δ*) and ergosterol (e.g., *erg3Δ*, *erg5Δ*) biosynthesis. In 1981, Hadwiger et al. [144] detected chitosan within plant cytoplasm and nucleus within 15 min after application, which indicate that chitosan can efficiently penetrate the thicker cell wall (the reason for its detection) and potentially interfered with DNA transcription and translation. This study suggests that chitosan can easily penetrate microbial cells since plants have a thicker cell wall than microbes.

Moreover, looking at the time factor (15 min), it is evident that chitosan can quickly interact with fungi and bacteria cellular DNA with subsequent inhibition of DNA transcription, as well as RNA and protein synthesis [140, 145, 146], leading to cell death. Chitosan triggered transcriptional responses when introduced to *S*. *cerevisiae* strain X2180-1A (MATa SUC2 mal gal2 CUP1). T-Profiler analysis showed cis-regulatory motifs apart from the environmental stress response correlated positively with expression in the chitosan-treated sample. Cin5p, Crz1p, and Rlm1p were the transcription factors associated with identified binding sites. Genes participating in cell wall organization, biogenesis, and signal transduction were also triggered in the treated sample compared to the control [134]. Some factors influencing the antimicrobial activity of chitosan is discussed above; however, Kong et al. [120] and Hosseinnejad and Jafari [147] published an excellent reviews on these factors.
