Eco-friendly biosynthesis and characterization of silver nanoparticles from Solanum lycopersicum (Tomato) peel waste and its application in disinfecting metallic surfaces

doi:10.21203/rs.3.rs-3100232/v1

Download PDF

Article

Eco-friendly biosynthesis and characterization of silver nanoparticles from Solanum lycopersicum (Tomato) peel waste and its application in disinfecting metallic surfaces

https://doi.org/10.21203/rs.3.rs-3100232/v1

This work is licensed under a CC BY 4.0 License

Version 1

posted

You are reading this latest preprint version

According to their special physio-chemical properties, Nanoparticles have gained worldwide attention as a new bio-alternative for chemical control agents. This investigation aims to the eco-friendly synthesis of nanosilver particles from tomato peel extract (TPE-AgNPs) and evaluates their characteristics and inhibitory activities against pathogenic bacteria and fungi as well as their role in metallic surface disinfecting. To initiate biosynthesis, tomato peel extract was mixed with silver nitrate (AgNO₃) solution until the color changes to reddish brown. Ultraviolet (UV-Visible) spectroscopy, Fourier-transform infrared (FTIR) spectroscopy, X-ray diffraction (XRD) analysis, and transmission electron microscopy (TEM) techniques were used to characterize biosynthesized TPE-AgNPs. Results recorded that obtained TPE-AgNPs had a strong score (238nm) of Plasmon resonance (SPR) by SPR of 4.5. Functional groups of carboxyl, hydroxyl, and phenolic groups existed and were detected by the FTIR spectrum. The synthesized TPE-AgNPs had an amorphous nature which was confirmed by XRD analysis. TEM analysis showed spherical TPE-AgNPs sized from 4.44-27.59nm. The biosynthesized TPE-AgNPs had a negative zeta potential of -68.44 mV. The inhibitory activities of synthesized TPE-AgNPs were evaluated against eleven microbial pathogenic using well diffusion method, inhibition zone diameter (IZD) was measured in centimeters. Results showed that B. subtilis and E.coli was the most sensitive pathogens with IZD of 4.0 and 0.92cm, respectively However, L. monocytogenes and S. sonnei were the most resistant pathogens with IZD of 0.92 and 0.90 cm, respectively. Synthesized TPE-AgNPs from tomato peels had good inhibitory potentials against pathogenic fungi with IZD of 3.0 and 0.92cm against A. solani and C. albicans, respectively. Applying the use of TPE-AgNPs as bio disinfectant significantly decreased the microbial load of metallic blades and proves its efficiency as a disinfectant agent after 120min. of contacting. So, more applications on disinfecting metallic surfaces such as dentistry are indeed needed.

Biological sciences/Biotechnology

Biological sciences/Microbiology

Biosynthesis

Silver nanoparticles

Tomato peels

Characterization

Pathogens

Biocontrol

Despite the need to their role as disinfectants and sterilizing agents, the mis handling of chemical disenfectants has led to an increase in antimicrobial resistance phenomenon and the inability to eliminate many of microorganisms. Therefore, it is necessary discover many biological compounds that help in reducing the spread of this phenomenon. Recently, multidrug resistant (MDR) microorganisms threats the public health all over the world as they are related to nosocomial infections. On the other hand, community -acquired infections have become prevalent because of some of MDR microorganisms especially, bacteria ¹. The spread of MDR bacteria into the environment is a crucial development and is associated with increased morbidity, mortality, healthcare costs, and antibiotic use². Pathogenic fungi threaten the lives of many microorganisms due to the increase in cases of infection and their transmission among infected patients³. Actual resistance to fungal treatment in some genera has become a serious problem due to their development of antibiotic resistance⁴. So, Therefore, it is very important to find alternative biological sources to avoid the problem of the development of antibiotic resistance in many microbes³. The most famous vegetable fruit, tomato (Lycopersicon esculentum), is well known for its content of vitamins A, B, and C, beta-carotene, phytosterols, and other nutrients⁵. Tomato juices and peels contain ingredients that have been shown to have anticancer, antimutagenic, antiangiogenic, antioxidant, antibacterial, and antiviral properties⁶. These ingredients also have a wide range of uses in the pharmaceutical, medical, cosmetic, and food industries. Also, Tomato peels were utilized in the biosynthesis process of AgNPs as a reducing and capping agent and showed good antibacterial activity towards resistant pathogens of B. subtilis, C. albicans, P. vulgaris, Pseudomonas sp., S. aureus⁷. The main constituents of tomato fruits include antioxidants (phenols, caretonides, leukopines, and glutathione) as well as proteins ⁸. These compounds play a key role in stabilizing and reducing the biosynthesized AgNPs. Tomato has an important role in the biosynthesis of silver and gold nanoparticles by using their callus and leaf extracts⁹. Moreover, many studies recommended the usage of fruit peels as a bio source for controlling pathogenic bacteria and fungi as an alternative safe source instead using harmful and toxic chemical fungicidal compounds and bactericidal preparations ¹⁰. So, the current study goal was to biosynthesize and characterize AgNPs from tomato peel extract as well as study its inhibitory potential against pathogenic bacteria, and fungi and apply it as a disinfectant for metallic surfaces.

2.1. Chemicals and solutions

Silver nitrate (AgNO₃) was purchased from Sigma Aldrich, Germany. It was used for the silver solution preparation. Standard antibiotics for Nystatin, Ampicillin, Streptomycin, and Fluconazole (1000µg/mL) for yeast, gram-negative, gram-positive, and Fungi, respectively.

2.2. Tomato peel collection and aqueous extract preparation from Solanum lycopersicum

Tomato peel waste was collected from local markets located in Cairo governorate, Egypt, and packed in sterilized plastic bags, then rapidly transferred under cooling using an ice box to the Microbiology lab. at the Faculty of Agriculture, Ain Shams University, Cairo, Egypt. Tomato peel waste was kept at 4°C until further use. Extraction of TPE-AgNPs was carried out as follows: All tomato peels were washed three times with deionized water (DW). Twenty grams of tomato peels were blended in 80mL of deionized water using an electric blender (Moulinex, France). Thereafter, the blended tomato peel extract was kept dark for 24h. Filtration for the obtained extract was done using filter paper (Whatman No. 1). All filtrate was collected in screw tubes and kept at 4°C for further studies⁷.

2.3. Pathogenic microbial strains

In the current investigation, eleven pathogenic microbial strains (six bacterial and five genera) were collected from Agric. Microbiology Department, Faculty of Agriculture, Ain Shams University, Cairo, Egypt. The collected pathogens were, Bacillus subtilis ATCC 6633, Escherichia coli ATCC 8379, Klebsiella pneumoniae ATCC 700603, Salmonella typhi DSM 17058, Shigella sonnei DSM 5570 and Listeria monocytogenes NC 013768, Alternaria solani ATCC 62102, Aspergillus flavus ATCC 9643, Candida albicans DSM 1386, Fusarium oxysporum ATCC 62506, and Rhizopus oryazae ATCC 96382 ¹¹.

2.4. Standard inoculum

Standard inoculum for all tested bacterial pathogens was prepared by picking up a pure single colony of each tested bacterial culture and inoculating it into a conical flask (250mL) containing 50mL of Nutrient broth medium. After inoculation, all cultures were incubated for 24h at 30ºC at an agitation speed of 150 rpm on a rotary shaker. One milliliter of obtained cultures was diluted using sterilized water to reach 4.5–5.77×10⁵ colony forming unit (CFU/mL). On the other hand, Standard inoculum for fungal pathogens was prepared by scratching their agar slants in 10mL of sterilized water and collecting their spore suspensions. Collected spore suspensions were diluted to reach (1.1–1.3×10⁸ CFU/mL) ¹².

2.5. Biosynthesis of TPE-AgNPs from Solanum lycopersicum

In the present study, we prepared a silver nitrate (AgNO₃) aqueous solution and aqueous tomato peel extract for Ag⁺ reduction to Ag⁰ owing to the production of TPE-AgNPs. The TPE-AgNPs biosynthesis process was started by adding 20mL of tomato peel aqueous extract to 80mL of the AgNO₃ solution. A negative control is used by preparing tomato peel aqueous extract without adding AgNO₃ solution. The mixtures were then mixed vigorously and kept overnight on a rotary shaker at 150rpm at 30℃ in the dark. Change in color from pale yellow to reddish brown is a positive result ⁵.

2.6. Characterization techniques of the biosynthesized TPE-AgNPs from Solanum lycopersicum

The surface images of nanoparticles were obtained using Scanning electron microscopy (SEM) using VEGA 3-T Scan, Brno, Czech Republic, image resolution 512 by 442, image pixel size: 0.04µm, with a 20.0KV accelerating voltage⁶. The Fourier transform infrared spectroscopy (FTIR) analysis has been subjected to silver nanoparticles samples on Bruker VERTEX 80 (Germany), in the wave number range 4000–400cm^− 1, with resolution 4cm^− 1, refractive index 2.4. The stability of silver nanoparticles was analyzed by measuring the absorbance using the UV-Vis (JASCO Corp., V-570) spectrum at 200–800nm ¹³. The crystalline states of nanoparticles were obtained using powder X-ray diffraction (XRD) analysis using a Shimadzu-7000 X-ray diffractometer. The dried TPE-AgNPs were subjected to X-ray diffraction (D2 phaser Bruker 2nd Gen X-ray diffraction) patterns on glass substrate were recorded in a wide range of Bragg angles θ at a scanning rate of 2 θ angles/min and wavelength (λ = 1.5406) ⁸. The size distribution of particles and zeta potential charge was estimated via dynamic light scattering (DLS) using Zetasizer Nano Series (HT), Nano ZS, Malvern Instruments, UK. Dynamic light scattering was used to provide information about the TPE-AgNPs properties at “long wavelength” limited by the wavelength of light¹⁴. Zeta potential was measured as one of the characteristics of charged colloids that may be determined using phase analysis light scattering based on DLS and laser Doppler velocimetry, with count rate 286KHz, and wavelength (λ = 1.54 nm) at 30°C and viscosity 0.995cp ¹⁵.

2.7. Antibacterial activity of the biosynthesized TPE-AgNPs from Solanum lycopersicum

2.7.1. Evaluation of antimicrobial activity using agar well-diffusion assay

For studying inhibitory potentials of TPE-AgNPs from tomato peels, Standard guidelines of CLSI were carried out using a good diffusion method. Briefly, Muller Hinton agar was prepared and poured into Petri dishes then kept till solidifying. 0.1mL of all bacterial and fungal standard inocula were spread individually onto the solidified MHA plates. Wells were made onto the inoculated MHA solidified agar using a sterilized cork porer (0.5cm in diameter). Each well was filled with 10µL with the prepared TPE-AgNPs against a control well filled with Ampicillin for G + ve bacteria, Streptomycin for G-ve bacteria, and Fluconazole for fungi). Incubation was carried out for all inoculated plates at 30℃ for 24h and 3-5d for bacteria and fungi, respectively. IZD was measured and recorded. Antimicrobial activity index (A.I) was calculated using the following eq:

$\text{A}.\text{I}=\frac{IZD of Ag-NPs extract }{IZD of standard antibiotic}$ ¹⁶

2.7.2. Minimum Inhibitory Concentration (MIC) of TPE-AgNPs from Solanum lycopersicum

MIC was done by the micro-diffusion method. Shortly, two-fold serial dilutions (1000, 500, 250, 125, 75, 50, 25, and 12.5µg/mL) were prepared from synthesized TPE-AgNPs and inoculated into welled MHA then incubated at 30 ℃ for 24h and 3-5d for bacteria and fungi, respectively. After incubation, IZD was measured. The MIC was determined as the lowest concentration of TPE-AgNPs inhibited microbial growth ¹⁷.

2.7.3. Assessment of minimum bactericidal concentration (MBC) of TPE-AGNPs from Solanum lycopersicum

According to the results of MIC, MBC was determined by reinoculating 0.1mL of the negative concentration results of MIC on MHA medium and incubating at 30℃ for 24h and 3-5d for bacteria and fungi, respectively. The total microbial count was determined in terms of colony forming unit (CFU/mL) and the MBC was defined as the lowest concentration of Ag-inhibited microbial growth ¹⁸.

2.7.4. Assessment of TPE-AgNPs mode of action

TPE-AgNPs mode of action was assessed according to the following equation:

$$Ag-NPs Mode of action=\frac{MBC value}{MIC value}$$

TPE-AgNPs is considered a bactericidal agent if MBC/MIC value is ≥ 4. On the other hand, it is considered bacteriostatic if the value is ≤ 2 ¹².

2.8. Application of TPE-AgNPs from Solanum lycopersicum as a disinfectant for metal tools.

Contaminated metal blades were submerged in a glass beaker containing 50mL of TPE-AgNPs solution (1000µg/mL) and left for 20 minutes. Interval blade samples were taken each 5 minutes and inoculated onto nutrient agar to estimate the total bacterial count expressed as CFU/mL. The negative control treatment was done by inoculating nutrient agar with contaminated blades without submerging them in TPE-AgNPs solution. All trials were carried out in triplicates.

2.9. Statistical analysis

All collected data were statistically analyzed using IBM® SPSS® Statistics software (2017). Tukey test at a P-value of 0.05 was applied.

3.1. Biosynthesis of AgNO₃ using tomato peel extract from Solanum lycopersicum

For the biosynthesis process, tomato peel aqueous extract and AgNO₃ (0.1M) were added to a glass beaker at a ratio of 1:1 and then left overnight at 30℃ in a rotary shaker at 150 rpm. Results recorded a change of the mixture color from pale yellow to brownish color indicating the reduction of Ag⁺ with brownish solution (after the reduction of Ag⁺), indicating the formation of silver nanoparticles as shown in Fig. 1.

3.2. Characterization of the synthesized TPE-AgNPs from Solanum lycopersicum

3.2.1. UV-Vis spectroscopy analysis

UV-Visible spectrophotometer analysis was used to confirm the synthesis of TPE-AgNPs. UV-visible spectrum data in Fig. 2 indicated a strong score (238nm) of Plasmon resonance (SPR) by SPR of 4.5.

3.2.2. X-Ray Diffraction (XRD)

X-ray diffraction (XRD) patterns of TPE-AgNPs detected two peaks of biosynthesized TPE-AgNPs from tomato peel extract at (1.54060mm), the peaks were observed in a range between (0–70⁰) and they were located at 4.5⁰ and 18.0⁰ which is in a range below 20⁰ as showing in Fig. 3.

3.2.3. Transmission Electron Microscopy (TEM)

The morphology and size of the biosynthesized TPE-AgNPs were observed by TEM images as illustrated in Fig. 4. TPE-AgNPs were spherical, monodisperse with particle sizes ranging from 4.44–27.59nm.

3.2.4. Fourier Transform Infrared Spectroscopy (FTIR) analysis

FTIR analysis was performed to characterize the biosynthesized TPE-AgNPs from tomato peels. Results in Table 1, Fig. 5 illustrated that seven independent peaks of the infrared spectrum were detected at 3307.64, 1636.16, 609.22, 591.67, 1042.64, 591.67and 609.22cm^–1. This spectrum clarifies a powerful band at 3307.64 cm^− 1 (corresponds to = C-H stretching of Alkenes and Alcohol or phenolic(; 1636.16 cm^− 1 (corresponds to the C = H stretching of Alkenes, and C = O stretching of Amides); 1456.99 cm^− 1 (corresponds to the C = C stretching of aromatic compounds); 1154.74 and 1042.64 cm^− 1 (corresponds to C-F stretching of alkyl and aryl halides), 609.22 cm^− 1 (corresponds to the C‒Br stretching of alkyl and aryl halides).

Table 1

FTIR-spectrophotometer absorption peaks of the prepared TPE-AgNPs from *Solanum lycopersicum* and their corresponding functional groups.
No.	Peak Position	Group	Class
1	3307.64	O-H stretching	Alcohol or phenolic
2	1636.16	NH₂-C = O stretching	Amides
3	609.22	= C = H bonding	Alkyne
4	591.67	= C = H bonding	Alkyne
5	1042.64	N-O stretching	Nitro compound
6	591.67	C-Cl stretching	Alkyl halide compound
7	609.22	C-Cl stretching	Alkyl halide compound

3.2.5. Dynamic Light Scattering (DLS)

In the current study, the mean diameter of the biosynthesized TPE-AgNPs from tomato peels was 168.7nm, as shown in Fig. 6.

3.2.6. Zeta potential and particle size

Zeta potential is used to measure the whole net charge found on the TPE-AgNPs surfaces related to the ion charges found in the same electric field. Figure 7 shows the charge distribution potential on the TPE-AgNPs surface. As seen, TPE-AgNPs had a negative zeta potential − 68.44mV owing to the nonionic characteristics of TPE-AgNPs capping molecules.

3.3. Inhibitory activity of biosynthesized TPE-AgNPs from Solanum lycopersicum.

Results in Table 2 illustrated that showed that all tested pathogenic bacteria were highly susceptible to AgNPs, while fungi were resistant. B. subtilis, S. typhi, and S. sonii were the most sensitive bacterial strains to the biosynthesized TPE-AgNPs from tomato peels with inhibition zone diameters (IZD) of 3.8, 2.6, and 2.6mm, respectively. However, no IZD formation and activity index (AI) was observed in the case of A. solani ATCC 62102, A. flavus ATCC 9643, F. oxysporum ATCC 62506, and R. oryazae ATCC 96382.

Table (2)

Inhibitory activity and activity index (AI) of clinical pathogenic bacterial and fungal strains after incubation at 37°C and 28°C for 24 and 96 h, respectively.

Pathogen strains	Inhibition zone diameter (cm)			Activity Index (AI)
Pathogen strains	Antibiotic (1000µg/mL)	TPE-AgNPs (1000µg/mL)		Activity Index (AI)
G^+ ve Bacteria
B. subtilis ATCC 6633	4.0^a ± 0.86		3.8^b ± 0.41	0.95
L. monocytogenes NC 013768	0.92^g ± 0.24		1.1^f ± 0.55	1.19
G^− ve Bacteria
E. coli ATCC 8379	0.92^g ± 0.36		1.0^f ± 0.14	1.08
K. pneumoniae ATCC 700603	1.00^f ± 0.85		2.0^e ± 0.77	2.00
S. typhi DSM 17058	0.91^g ± 0.22		2.6^d ± 0.11	2.86
S. sonnei DSM 5570	0.90^g ± 0.24		2.6^d ± 0.30	2.88
Fungi and yeasts
A. solani ATCC 62102	3.0^c ± 0.50		0.00	0.00
A. flavus ATCC 9643	2.8^d ± 0.82		0.00	0.00
F. oxysporum ATCC 62506	2.1^e ± 0.66		0.00	0.00
R. oryazae ATCC 96382	1.90^e ± 0.50		0.00	0.00
C. albicans DSM 1386	0.92^g ± 0.40		1.1^f ± 0.50	1.19

Standard antibiotics were streptomycin, ampicillin, and fluconazole against G^+ ve bacteria, G^− ve bacteria, and fungi, respectively.*mm = milli meter, AI = activity index, and SE (±) = standard error. Values in the same column followed by the same letter do not significantly differ from each other, according to Duncan (1955) at a 5% level.

3.4. Minimum inhibition concentration (MIC) for TPE-AgNPs from Solanum lycopersicum

Results in Table 3 show the MIC values of biosynthesized TPE-AgNPs (1000 to 12.5µg/mL) against the tested pathogenic bacteria and fungi strains. The MIC value was 250µg/mL for E. coli, K. pneumoniae, S. typhi, L. monocytogenes, and S. sonnei, while it was 75.0µg/mL for B. subtilis. It is shown that 100% of the antibacterial spectrum activity of biosynthesized TPE-AgNPs from tomato peel extract was from 1000 to 250µg/mL, whereas at concentrations of 125, and 75µg/mL the activity was 16.7%. Whereas at a concentration of 50-12.5µg/mL the antibacterial spectrum showed no activity.

The TPE-AgNPs antifungal activity against C. albicans with MIC at 500µg/mL while the other fungal strains exhibited no antifungal effect, as shown in Table 3. At the concentrations of 1000 − 500µg/mL, 80.0% of the spectrum of activity was attained for all fungi and yeast strains. In addition, concentrations ranging from 250 − 12.5µg/mL did not display any activity against the tested fungi and yeast strains.

Table (3)

Minimum inhibitory concentration (MIC) of TPE-AgNPs from Solanum lycopersicum against clinical pathogenic bacterial and fungal strains after incubation at 37°C and 28°C for 24 and 96h, respectively.

Pathogenic bacteria		MIC (µg/mL) of TPE-AgNPs
Pathogenic bacteria		1000 (control)	500	250		125	75		50	25	12.5
B. subtilis ATCC 6633		-	-	-		-	-		+	+	+
E. coli ATCC 8379		-	-	-		+	+		+	+	+
K. pneumoniae ATCC 700603		-	-	-		+	+		+	+	+
L. monocytogenes NC 013768		-	-	-		+	+		+	+	+
S. typhi DSM 17058		-	-	-		+	+		+	+	+
S. sonnei DSM 5570		-	-	-		+	+		+	+	+
The spectrum of activity (%)		6/6	6/6	6/6		1/6	1/6		0/6	0/6	0/6
The spectrum of activity (%)		100	100	100		16.7	16.7		0	0	0
Pathogenic fungi	MIC (µg/mL) of TPE-AgNPs
Pathogenic fungi	1000 (control)		500	250	125		75	50	25	12.5
A. solani ATCC 62102	+		+	+	+		+	+	+	+
A. flavus ATCC 9643	+		+	+	+		+	+	+	+
C. albicans DSM 1386	-		-	+	+		+	+	+	+
F. oxysporum ATCC 62506	+		+	+	+		+	+	+	+
R. oryazae ATCC 96382	+		+	+	+		+	+	+	+
The spectrum of activity (%)	4/5		4/5	0/5	0/5		0/5	0/5	0/5	0/5
The spectrum of activity (%)	80.0		80.0	0.00	0.00		0.00	0.00	0.00	0.00

-No growth, + positive growth.

3.5. Minimum lethal concentration (MLC) for TPE-AgNPs from Solanum lycopersicum

Values of minimum lethal concentration MLC (MBC and/or MFC) for tomato peels silver nanoparticles (TPE-AgNPs) are presented in Table 4, The MBC value was exhibited at 500µg/mL for E. coli, K. pneumoniae, and S. typhi, L. monocytogenes, and S. sonnei, while it was 125.0µg/mL with B. subtilis. The results clearly showed 100% of the antibacterial spectrum activity of TPE-AgNPs at concentrations ranging from 1000 to 500µg/mL, whereas, at concentrations of 250 and 125µg/mL, the activity was 16.7%. However, concentrations of 75–12.5µg/mL had no observed inhibitory activity. The MIC of TPE-AgNPs against C. albicans was 1000µg/mL, while the other fungal strains exhibited no antifungal effect, as shown in Table 4. At the concentration of 1000µg/mL, 80.0% of the spectrum of activity was attained for all fungi and yeast strains. In addition, concentrations ranging from 500 − 12.5µg/mL did not display any activity against the tested fungi and yeast strains.

Table (4)

Minimum lethal concentration (MLC) of TPE-AgNPs against clinical pathogenic bacterial and fungal strains after incubation at 37°C and 28°C for 24 and 96 h, respectively.

Pathogenic bacteria		MBC (µg/mL) of AgNPs
Pathogenic bacteria		1000 (control)	500	250		125	75		50	25	12.5
B. subtilis ATCC 6633		-	-	-		-	+		+	+	+
E. coli ATCC 8379		-	-	+		+	+		+	+	+
K. pneumoniae ATCC 700603		-	-	+		+	+		+	+	+
L. monocytogenes NC 013768		-	-	+		+	+		+	+	+
S. typhi DSM 17058		-	-	+		+	+		+	+	+
S. sonnei DSM 5570		-	-	+		+	+		+	+	+
The spectrum of activity (%)		6/6	6/6	1/6		1/6	0/6		0/6	0/6	0/6
The spectrum of activity (%)		100	100	16.7		16.7	0.00		0.00	0.00	0.00
Pathogenic fungi	MFC (µg/mL) of AgNPs
Pathogenic fungi	1000 (control)		500	250	125		75	50	25	12.5
A. solani ATCC 62102	+		+	+	+		+	+	+	+
A. flavus ATCC 9643	+		+	+	+		+	+	+	+
C. albicans DSM 1386	-		+	+	+		+	+	+	+
F. oxysporum ATCC 62506	+		+	+	+		+	+	+	+
R. oryazae ATCC 96382	+		+	+	+		+	+	+	+
The spectrum of activity (%)	4/5		0/5	0/5	0/5		0/5	0/5	0/5	0/5
The spectrum of activity (%)	80.00		0.00	0.00	0.00		0.00	0.00	0.00	0.00

ـــ= No growth, + = growth.

3.6. Tomato peels silver nanoparticles (TPE-AgNPs) mode of action

Finally, it could be observed that the mode of action of tomato peels silver nanoparticles (TPE-AgNPs) against pathogenic bacterial and fungal strains is shown in Table 5. Results indicated that the TPE-AgNPs have a bactericidal and fungicidal effect with MBC or MFC/ MIC ≤ 2 toward 7 strains B. subtilis, E. coli, K. pneumoniae, L. monocytogenes, S. typhi, S. sonnei, and C. albicans.

Table (5)

Minimum inhibitory concentration (MIC) and minimum lethal concentration (MLC) of TPE-AgNPs on pathogenic bacterial and fungal strains after incubation at 37 and 28°C for 24 and 96h, respectively.

Pathogenic bacteria	MIC (TPE-AgNPs µg/mL)	MBC (TPE-AgNPs µg/mL)	MBC/MIC Ratio	Mode of action
Bacteria
B. subtilis ATCC 6633	75	125	1.7	+
E. coli ATCC 8379	250	500	2	+
K. pneumoniae ATCC 700603	250	500	2	+
L. monocytogenes NC 013768	250	500	2	+
S. typhi DSM 17058	250	500	2	+
S. sonnei DSM 5570	250	500	2	+
Fungi
A. solani ATCC 62102	0	0	0	0
A. flavus ATCC 9643	0	0	0	0
C. albicans DSM 1386	500	1000	2	+
F. oxysporum ATCC 62506	0	0	0	0
R. oryazae ATCC 96382	0	0	0	0
Bactericidal/Fungicidal (+) = ≤ 2 and Bacteriostatic/fungistatic (-) effect = ≥ 4.

3.7. Application of TPE-AgNPs from Solanum lycopersicum as a disinfectant for metal tools

Contaminated metal blades were submerged in a glass beaker containing 50mL of TPE-AgNPs solution (1000µg/mL) and left for 120min. To test the disinfectant efficiency of TPE-AgNPs. Results in Fig. 8 and Fig. 9 shows a significant decrease in total bacterial count in contaminated blades treated with TPE-AgNPs at a concentration of 1000µg/mL, leading to the complete elimination of microbial contaminants after 120 min. of contacting time.

Not only does the misuse and mishandling of chemical disinfectants harm workers in the industry, but also, it promotes the increase in the number of multidrug-resistant (MDR) microorganisms (MDR). Therefore, it is indeed needed to find new more bio alternatives for chemical cleansers and disinfectants to decrease the phenomenon of MDR. Biosynthesis of nanoparticles is considered a promising technique to play an important role in inhibiting pathogenic bacteria and fungi as well as disinfecting many surfaces as well as metallic ones.

Nanoparticles are characterized by having small diameters smaller than 1µm and have two different morphological shapes such as crystalline and amorphous one¹⁹. In our study, the morphological shape of TPE-AGNPs was spherical and monodisperse with a particle size range from 4.44-27.59nm. In a study on the average nano particle size of the pomegranate peel, it was 8.0 nm⁴, while that of the orange peel was 14.0 nm²⁰. UV- visible spectrophotometry is used to detect the spectrum of the biosynthesized TPE-AgNPS. We recorded a strong score (238nm) of Plasmon resonance (SPR) with an SPR of 4.5. Also, in a similar study on olive, UV-Visible spectrophotometric readings were recorded at a wavelength of 238nm²¹. It could be noted that the biosynthesized NPs surrounded by amino acids, proteins, and other essential metabolites are present in the extract which gives it a negative charge ¹⁵. XRD analysis shows X-ray diffraction (XRD) patterns of TPE-AgNPs detected two peaks of biosynthesized TPE-AgNPs from tomato peels extract at 1.54060mm, the peaks were observed in a range between (0–70⁰) and they were located at 4.5⁰ and 18.0⁰ which is in a range below 20⁰. In another study using the bio-reduction method, the XRD result was four peaks ranging between 10-25nm, they were pure crystalline and had an average size of 7.0nm ²²which is approximately similar compared to the current results. Functional groups of carboxyl, hydroxyl, and phenolic groups existed by the FTIR spectrum. Many researches reveal absorption bands in 1936, 1882, 1803,1764, 1656, 1602, 1510, 1392, 1222, 1188, 1085cm^− 1. In another study of ²³. The vibrational bands correspond to the bonds such as C = O, C-C, C-N, C-O-C, C-O-H, and C-Cl which were in the region range of 600-3822cm^− 1. Zeta potential is used to measure the negative net charge on the particle’s surface. We had a total net charge estimated by -68.44mV owing to the nonionic characteristics of TPE-AgNPs capping molecules. The results are consistent with previous studies that reported a strong peak for TET-AgNPs was the most stable due to differences in the distribution of capping polyphenolic constituents present in the extract²⁴. Moreover, the antifungal effect of biosynthesized AgNPs with fruit peel extracts reduces the mycelial growth of A. solani⁴.

We found that TPE-AgNPs could inhibit the microbial growth that contaminates metallic blades after contacting time of 120min.

In summary, biosynthesized silver nanoparticles (AgNPs) in this study utilizing the aqueous tomato peel extract in a suitable and ecologically friendly method. The presence of phenolic chemicals, glutathione, and carotenoids in tomato peels has been shown to have the ability to reduce and stabilize the development of silver nanoparticles that can be used as antimicrobial agents. Several tests were conducted for the physiochemical characterization of the biosynthesized silver nanoparticles, with excellent results for the UV visible spectrophotometer, XRD, FTIR, TEM, and Zeta potential. Then the antimicrobial effect of these nanoparticles was examined against 11 different strains of bacteria and fungi through the agar well diffusion method the B. subtilis ATCC 6633, E. coli ATCC 8379, K. pneumoniae ATCC 700603, L. monocytogenes NC 013768, S. typhi DSM 17058, S. sonnei DSM 5570, and C. albicans DSM 1386. TPE-AgNPs could inhibit the microbial growth that contaminates metallic blades after contacting time of 60min. This indicated that the biosynthesized nanoparticles have good compatibility when used to treat bacteria and yeast infections with eukaryote organisms.

Author contribution

BTA, SME, BH, SHA conceived and designed the research. BTA, SME, BH, SHA, EA,EH,SA, HM, AG conducted experiments and collected data. BTA, SME, BH, SHA analyzed and interpreted microbiological data, BTA, SME, BH, SHA revised the manuscript. All authors wrote the draft manuscript, reviewed and edited the manuscript. All authors read and approved the manuscript.

Conflicts of interest

The authors have no competing interests to declare relevant to this article’s content.

Acknowledgment

The authors would like to thank New Programs Administration, Faculty of Agriculture, Ain Shams University for providing all lab facilities.

Data availability

The raw data and analyzed data used during the current study available from the corresponding author on reasonable request.

All microbial pathogens were provided by Agricultural Microbiology Department, Faculty of Agriculture, Ain Shams University, Cairo, Egypt and was deposited in the following strain providers

subtilis ATCC 6633 was from ATCC collection https://www.atcc.org/products/6633
coli ATCC 8379 was from ATCC collection https://www.atcc.org/products/8379 and was deposited in GenBank with taxonomy ID: NCBI:txid 481805 https://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgiTPE-AgNPs id=481805
pneumonia ATCC 700603 was from ATCC collection https://www.atcc.org/products/700603
monocytogenes was deposited in Genbank with gene accession number NC_013768 https://www.ncbi.nlm.nih.gov/nuccore/NC_013768
typhi DSM 17058 was from DSM collection https://www.dsmz.de/collection/catalogue/details/culture/DSM-17058
sonii DSM 5570 was from DSM collection https://www.dsmz.de/collection/catalogue/details/culture/DSM-5570
solani ATCC 62102 was from ATCC collection https://www.atcc.org/products/62102
flavus ATCC 9643 was from ATCC collection https://www.atcc.org/products/9643
albicans DSM 1386 was from DSM collection https://www.dsmz.de/collection/catalogue/details/culture/DSM-1386
oxysporum ATCC 62506 was from ATCC https://www.atcc.org/products/62506
oryzae ATCC96382 was from ATCC collection https://www.atcc.org/products/96382

Ethical Statement

This article does not contain any studies with human participants or animals performed by any of the authors

Frieri, M., Kumar, K. & Boutin, A. Antibiotic resistance. Journal of Infection and Public Health vol. 10 369–378 Preprint at https://doi.org/10.1016/j.jiph.2016.08.007 (2017).
Murugaiyan, J. et al. Progress in Alternative Strategies to Combat Antimicrobial Resistance: Focus on Antibiotics. Antibiotics vol. 11 Preprint at https://doi.org/10.3390/antibiotics11020200 (2022).
Mansoor, S. et al. Fabrication of Silver Nanoparticles Against Fungal Pathogens. Frontiers in Nanotechnology vol. 3 Preprint at https://doi.org/10.3389/fnano.2021.679358 (2021).
Mostafa, Y. S., Alamri, S. A., Alrumman, S. A., Hashem, M. & Baka, Z. A. Green synthesis of silver nanoparticles using pomegranate and orange peel extracts and their antifungal activity against Alternaria solani, the causal agent of early blight disease of tomato. Plants 10, (2021).
Carbone, K. et al. Microwave-assisted synthesis of catalytic silver nanoparticles by hyperpigmented tomato skins: A green approach. LWT 133, (2020).
Zia, M. et al. Green synthesis of silver nanoparticles from grape and tomato juices and evaluation of biological activities. IET Nanobiotechnol 11, 193–199 (2017).
Carbone, K. et al. GREEN SYNTHESIS OF SILVER NANOPARTICLES FROM HYPERPIGMENTED TOMATO SKINS AND PRELIMINARY EVALUATION OF THE INSECTICIDAL ACTIVITY. https://www.researchgate.net/publication/349590157 (2020).
Chand, K. et al. Green synthesis characterization and antimicrobial activity against: Staphylococcus aureus of silver nanoparticles using extracts of neem, onion and tomato. RSC Adv 9, 17002–17015 (2019).
SI, A. et al. Sustainable preparation of gold nanoparticles via green chemistry approach for biogenic applications. Materials Today Chemistry vol. 17 Preprint at https://doi.org/10.1016/j.mtchem.2020.100327 (2020).
Kumar, S. et al. Plant extract mediated silver nanoparticles and their applications as antimicrobials and in sustainable food packaging: A state-of-the-art review. Trends in Food Science and Technology vol. 112 651–666 Preprint at https://doi.org/10.1016/j.tifs.2021.04.031 (2021).
Hemdan, B. A., El Nahrawy, A. M., Mansour, A. F. M. & Hammad, A. B. A. Green sol–gel synthesis of novel nanoporous copper aluminosilicate for the eradication of pathogenic microbes in drinking water and wastewater treatment. Environmental Science and Pollution Research 26, 9508–9523 (2019).
Galal, G. F., Abd-Elhalim, B. T., Abou-Taleb, K. A., Haroun, A. A. & Gamal, R. F. Toxicity assessment of green synthesized Cu nanoparticles by cell-free extract of Pseudomonas silesiensis as antitumor cancer and antimicrobial. Annals of Agricultural Sciences 66, 8–15 (2021).
Yap, Y. H. et al. Green Synthesis of Silver Nanoparticle Using Water Extract of Onion Peel and Application in the Acetylation Reaction. Arab J Sci Eng 45, 4797–4807 (2020).
Mohamed, M. A., Jaafar, J., Ismail, A. F., Othman, M. H. D. & Rahman, M. A. Fourier Transform Infrared (FTIR) Spectroscopy. Membrane Characterization 3–29 (2017) doi:10.1016/B978-0-444-63776-5.00001-2.
Abu-Hussien, S., Sameh, M. & Abo El-Naga, M. Optimization of Polyunsaturated Fatty Acids (PUFAs) Production from Olive Cake by Pseudomonas fluorescens NBRC14160 using Response Surface Methodology and their Application in Kareish Cheese Manufacture. Arab Universities Journal of Agricultural Sciences 0, 0–0 (2021).
Ijaz, I. et al. Green synthesis of silver nanoparticles using different plants parts and biological organisms, characterization and antibacterial activity. Environmental Nanotechnology, Monitoring and Management vol. 18 Preprint at https://doi.org/10.1016/j.enmm.2022.100704 (2022).
Valdez-Salas, B. et al. Promotion of surgical masks antimicrobial activity by disinfection and impregnation with disinfectant silver nanoparticles. Int J Nanomedicine 16, 2689–2702 (2021).
Abd-Elhalim, B. T. et al. Enhancing durability and sustainable preservation of Egyptian stone monuments using metabolites produced by Streptomyces exfoliatus. Sci Rep 13, 9458 (2023).
Cheng, J. et al. Preparation of a multifunctional silver nanoparticles polylactic acid food packaging film using mango peel extract. Int J Biol Macromol 188, 678–688 (2021).
Mohamed, Y. M. A. & Elshahawy, I. E. Antifungal activity of photo-biosynthesized silver nanoparticles (AgNPs) from organic constituents in orange peel extract against phytopathogenic Macrophomina phaseolina. Eur J Plant Pathol 162, 725–738 (2022).
Alowaiesh, B. F., Alhaithloul, H. A. S., Saad, A. M. & Hassanin, A. A. Green Biogenic of Silver Nanoparticles Using Polyphenolic Extract of Olive Leaf Wastes with Focus on Their Anticancer and Antimicrobial Activities. Plants 12, (2023).
Asif, M. et al. Green Synthesis of Silver Nanoparticles (AgNPs), Structural Characterization, and their Antibacterial Potential. Dose-Response 20, (2022).
Ansari, M. et al. Green Synthesized Silver Nanoparticles: A Novel Approach for the Enhanced Growth and Yield of Tomato against Early Blight Disease. Microorganisms 11, (2023).
Weng, X., Yang, K., Owens, G. & Chen, Z. Biosynthesis of silver nanoparticles using three different fruit extracts: Characterization, formation mechanism and estrogen removal. J Environ Manage 316, (2022).

No competing interests reported.

Download PDF

Version 1

posted

You are reading this latest preprint version

Eco-friendly biosynthesis and characterization of silver nanoparticles from Solanum lycopersicum (Tomato) peel waste and its application in disinfecting metallic surfaces

Status:

Version 1

Abstract

Figures

1. Introduction

2. Materials and methods

2.1. Chemicals and solutions

2.2. Tomato peel collection and aqueous extract preparation from Solanum lycopersicum

2.3. Pathogenic microbial strains

2.4. Standard inoculum

2.5. Biosynthesis of TPE-AgNPs from Solanum lycopersicum

2.6. Characterization techniques of the biosynthesized TPE-AgNPs from Solanum lycopersicum

2.7. Antibacterial activity of the biosynthesized TPE-AgNPs from Solanum lycopersicum

2.7.1. Evaluation of antimicrobial activity using agar well-diffusion assay

2.7.2. Minimum Inhibitory Concentration (MIC) of TPE-AgNPs from Solanum lycopersicum

2.7.3. Assessment of minimum bactericidal concentration (MBC) of TPE-AGNPs from Solanum lycopersicum

2.7.4. Assessment of TPE-AgNPs mode of action

2.8. Application of TPE-AgNPs from Solanum lycopersicum as a disinfectant for metal tools.

2.9. Statistical analysis

3. Results

3.1. Biosynthesis of AgNO3 using tomato peel extract from Solanum lycopersicum

3.2. Characterization of the synthesized TPE-AgNPs from Solanum lycopersicum

3.2.1. UV-Vis spectroscopy analysis

3.2.2. X-Ray Diffraction (XRD)

3.2.3. Transmission Electron Microscopy (TEM)

3.2.4. Fourier Transform Infrared Spectroscopy (FTIR) analysis

3.2.5. Dynamic Light Scattering (DLS)

3.2.6. Zeta potential and particle size

3.3. Inhibitory activity of biosynthesized TPE-AgNPs from Solanum lycopersicum.

3.4. Minimum inhibition concentration (MIC) for TPE-AgNPs from Solanum lycopersicum

3.5. Minimum lethal concentration (MLC) for TPE-AgNPs from Solanum lycopersicum

3.6. Tomato peels silver nanoparticles (TPE-AgNPs) mode of action

3.7. Application of TPE-AgNPs from Solanum lycopersicum as a disinfectant for metal tools

Discussion

Conclusion

Declarations

References

Additional Declarations

Status:

Version 1

3.1. Biosynthesis of AgNO₃ using tomato peel extract from Solanum lycopersicum