**1. Introduction**

Recent studies report that cinnamon essential oil (CEO) and rosemary essential oil (REO) have demonstrated antimicrobial activity, against microorganisms, due to their main components that are cinnamaldehyde and eucalyptus, respectively [1, 2].

The encapsulation of EOs has shown to provide stability to lipidic and volatile compounds, maintaining their antimicrobial activity. Encapsulation by emulsification has been recently studied to protect lipidic compounds in a micro- and nanoscale [3, 4]. The different compounds encapsulated in microemulsions are very stable during storage and can be released in specific conditions and dosage [5]. The controlled release of the encapsulated compounds can be influenced by different factors including environmental conditions, form and shape of particles, particle size, the bioactive solubility, and diffusivity [6, 7]. The release of the microencapsulated EO may determine their effectiveness depending on the selected application;

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*Technology, Science and Culture - A Global Vision*

[1] CONAGUA. Estadística del agua en México. México, D.F.: Comisión

[2] CONAGUA. Estadística del Agua en México. Cd. México: Comisión Nacional

[3] R. J. Brandes Company. Naturalized Streamflow Data. Austin, Texas: Texas Commission on Environmental Quality;

[4] USDA. Hydrology National Engineering Handbook: Time of Concentration. Washington D.C.: Natural Resources Conservation

[5] US Army Corps of Engineers. Hydrologic Modeling System, HEC-HMS. Quick Start Guide Version 3.5. Davis, CA, USA: Institute for Water Resources, Hydrological Engineering

[6] Legates DR, MacCabe GJ. Evaluating the use of "goodness-of-fit" measures in hydrologic and hydroclimatic model validation. Water Resources Research.

[7] Moriasi DN, Arnold JG, Van Liew MW, Bigner RL, Harmel RD, Veith TL. Model evaluation guidelines for systematic quantification of accuracy in watershed simulation. American Society of Agricultural and Biological Engineers. 2007;**50**(3):885-900

Nacional del Agua; 2003

**References**

del Agua; 2017

Service; 2010

Center; 2010

1999;**35**(1):233-241

2003

consequently, the development of mathematical models able to describe the release of these compounds establishes a very useful tool for the behavior prediction [6].

The objective of this study was to evaluate the release of CEO and REO encapsulated in microemulsions prepared by high-frequency ultrasound, setting a target concentration to be released, according to their antimicrobial activity against bacteria related to common illness by food consumption.

## **2. Methodology**

#### **2.1 Materials**

The REO was purchased at Hersol (Mexico) and the CEO at TECNAAL (Mexico). For the preparation of the oil-in-water (O/W) microemulsions, inulin (Fructagave PR95, Mexico) was used as stabilizer and Tween 80 (Sigma-Aldrich, USA) as emulsifying agent.

#### **2.2 Microemulsion formation**

The dispersed phase of CEO microemulsion was prepared with 10% (w/w) of CEO, while for REO microemulsion, it was prepared with 30% (w/w) of REO. The microemulsions were prepared using an ultrasound homogenizer (Cole Parmer, CP 505, USA). The continuous phase of the O/W CEO or REO microemulsions was formulated with 3% of Tween 80 and 5% of inulin.

#### **2.3 Particle size**

Particle size distributions of the microemulsions' droplets were measured using a particle analyzer by laser diffraction (Bluewave, Microtrac, USA).

#### **2.4 Release of encapsulated CEO and REO**

The amount of microemulsion dispersed in water was defined according to values reported in other studies [8, 9] of minimum inhibitory concentration (MIC) and minimum bactericide concentration (MBC) against bacteria of the CEO and REO; maximum concentrations of 15 and 80 μL/mL, respectively, were adjusted in this study. The microemulsions were added in water, maintaining the system agitated at 60 rpm and 25°C. The quantification of the EOs' release was made by dissolving 1 mL of sample in 9 mL of n-hexane, determining their absorbance in a spectrophotometer at 310 nm for the CEO and 225 nm for the REO.

#### **2.5 Mathematical modeling for release profiles**

The concentration data of the released EOs at different times were recorded and fitted to dynamic kinetic models, by optimizing the minimal square error. Eq. (1) was applied to describe a first-order kinetic model. A kinetic with an initial zero slope and a maximum slope at the inflection point (typical *S* shape response) was used to fit a second-order dynamic (Eq. (2)).

$$\mathbf{C}(t) = \mathbf{C}\_{\text{max}} \left[ \mathbf{1} - e^{-k\boldsymbol{\beta}} \right] \tag{1}$$

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**Figure 1.**

Ugarte et al. [10].

*Modeling of the Controlled Release of Essential Oils Encapsulated by Emulsification*

release, the concentration of 60 μL/mL was obtained after 360 min.

would be available after 210 and 420 min, respectively.

According to the release curves (**Figure 1**), the CEO release showed a constant rate in the first 90 min; followed by a plateau from 90 to 360 min. On the other hand, the REO release presented a delay release of 60 min, but then the release rate was constant until 420 min passed, presenting a decrease of the release rate until

The antimicrobial activity of EOs depends on their availability, in certain food, to inhibit spoilage microorganisms [8]. In **Figure 1**, it can be observed that the encapsulated CEO release of 10 μL/mL was obtained after 60 min, and for REO

Tomičić et al. [9] evaluated the antimicrobial activity of different EOs against *Listeria monocytogenes*. The authors reported that the MIC value with CEO was 2.56 μL EO/mL and with REO was 20.48 μL EO/mL. In addition, Kaskatepe et al. [8] reported MICs values of commercial and natural CEO between 0.39 and 12.5 μL EO/mL against *Escherichia coli*. For this study, these values were taken as a reference for the EOs release. In accordance with the release curve of CEO microemulsion (**Figure 1**), the MICs for both bacteria would be available in 30 min, whereas the MBC for *L. monocytogenes* will be released in 90 min and for *E. coli* in 60 min. Meanwhile, for the REO microemulsion, the MICs would be released after 120 min, for both bacteria. Finally, the concentration of MBC for *L. monocytogenes* and *E. coli*

Different factors can influence the rate at which the EOs is released: first, the amount of EO in the microemulsion; and second, the lipid droplet size in the microemulsion. The droplet size D50 determined in CEO and REO microemulsions were 1.99 and 3.45 μm, respectively. Since the CEO microemulsion droplet size was smaller than in REO microemulsion, it enhanced its release in the aqueous solution. As shown in **Figure 1**, the release profile for CEO shows a monotonic response with a rapid raising at the beginning of the process, and after a lapse of time, the CEO concentration tends to achieve a stationary value, such behavior was described by a first-order kinetic model. For REO, a change in the slope of the release curve, describing a *S*-shape response, can be observed. This behavior was fitted to a second-order dynamic model, as discussed by Cardoso-

For the CEO release, k1 was determined as 0.0166, while for REO liberation, the parameter k2 was calculated as 0.00708. To be dimensionally consistent, both

*(a) Release of CEO microemulsion (*■*) and (b) REO microemulsion (•), both in water at 60 rpm and*

*25°C. Continuous lines represent mathematical model estimation.*

*DOI: http://dx.doi.org/10.5772/intechopen.83691*

reaching a plateau at 600 min.

**3. Results**

$$\mathbf{C}(t) = \mathbf{C}\_{\text{max}} \left| \mathbf{1} - \left( k\_2 t + \mathbf{1} \right) e^{-k\_2 t} \right| \tag{2}$$

*Modeling of the Controlled Release of Essential Oils Encapsulated by Emulsification DOI: http://dx.doi.org/10.5772/intechopen.83691*


*2.3. Climatic Trend Analysis*

The non-parametric Mann Kendall (MK) test [47] was used to determine the significance of the climate trends, because the climatic data were not independent and normally distributed. The seasonal trends for Tmin, Tmax and Rainfall during the growing period with yield data were determined using a linear regression model. The free and open software package developed by the Finnish Meteorological Institute (MAKESENS) (https://en.ilmatieteenlaitos.fi/makesens) was used for the Mann Kendall (MK) test and Sen's slope estimator. The Sen's slope estimator allows for the significance of the trend to be analyzed. The MK test is robust, simple and frequently used in climate, environmental and hydrological studies [13,48–51]. The Sen's slope is a robust estimate of the underlying trend.


**Abstract**

Biological control is a potential nonchemical method to manage plant pathogens by beneficial microorganisms. To improve antagonistic potential of biocontrol agents, mutation by radiations, chemicals, and genetic manipulations has been used. Genetic techniques and ionizing radiation containing direct or indirect emissions play the greatest role for selection of useful microorganisms to enhance the efficiency of biological systems. Indeed, genetic engineering has a main role in increasing antimicrobial metabolites, host colonization ability, and endurance in micro-ecosystem. Genetic improvement can be achieved by protoplast fusion, genetic modification (GM), and chemical (genotoxic agents) and physical mutations. However, ultraviolet light and ionizing radiations can induce modifications in the genome of an organism. Irradiation, particularly gamma rays, is also applied for controlling postharvest diseases. Indeed, irradiation cannot completely eliminate pathogens, but it might result in cell injury and directly damage the chromosomal DNA of a living cell. This technology has been used for many reasons including disinfestation of foods, reducing foodborne pathogens, and extending shelf life many fruits, vegetables, and nuts. In the current review, we discuss advances in the radiation and molecular genetic techniques with the aim to improve antagonistic potential of microorganisms as it is applied to the suppression of plant pathogens.

**Keywords:** biocontrol agents, genetic engineering, ionizing radiations

#### **1. Introduction**

The risks associated with chemical residues on the leaves and fruits have highlighted the need for more useful and safer alternative control treatments. Biological control is a potential nonchemical mean to manage plant pathogens including fungi, bacteria, nematodes, or weeds by beneficial microorganisms such as fungi, yeast, or bacteria [1]. The virulence of pathogens probably changes if genetic mutations occur in the genes related to pathogenicity of microorganisms upon mutagenic treatments [2]. It is quite obvious that the application of ionizing radiations such as gamma rays and X-rays and genetic methods plays the greatest role in the development of techniques for the selection of useful microorganisms

[3]. Indeed, irradiation is the process of exposing an amount of energy in the form of speed particles or rays for improving food safety and reducing microorganisms that destroy agricultural crops. For instance, studies on transcriptional changes of *Salmonella typhimurium* using gamma irradiation showed that the expression of the virulence genes in irradiated mutants was reduced in comparison with non-irradiated controls [4]. It has been also shown that gamma irradiation alone or in combination with other methods can improve the microbiological safety [5]. Likewise, the wholesomeness (lack of teratogenicity, mutagenicity, and toxicity) of irradiated products has been studied extensively, in which food irradiation has not provided any evidence of increased threat of mycotoxin formation in irradiated food [6]. There is an increasing interest in commercial application of genetically modified microorganisms with improved biocontrol properties toward plant pathogens [7]. To that end, this chapter is an advanced survey reviewing the radiation and molecular genetic techniques with the aim to improve antagonistic potential of microorganisms as it is applied to the suppression of plant diseases.