Antibacterial activity of green synthesized selenium nanoparticles using Vaccinium arctostaphylos (L.) fruit extract

Figures & data

Figure 1. Determination of the minimum inhibitory concentration (MIC) for green synthesized SeNPs, extract, and ciprofloxacin against S. aureus, E. coli, and C. diphtheria. Following the incubation period, resazurin dye was added. Column 1: a negative control (the natural blue/purple color of resazurin reduced to red-colorless). The plate contains concentrations ranging from 0.5 mg/ml at the highest point to 0.03125 mg ml⁻¹ at the lowest point, both of which were achieved using double serial dilution. The MIC value was calculated using the concentration of combinations in Column 8, which did not change color significantly. Wells content varied in concentration from 0.05 to 0.03125 mg/ml.

Figure 2. (A) demonstrated a reduction of Se (IV) to Se by changing the color of the SS solution to ruby red. (0). (B) UV-vis spectra of green SeNPs synthesized with V. arctostaphylos (L.) fruit extract (10% w/v).

Figure 3. FT‐IR spectrum analysis of green synthesized SeNPs by using fruit extract of V. arctostaphylos (L.) as a capping agent.

Table 1. Absorption peaks observed in the FTIR spectra, the corresponding functional groups and interpretations

No.	Absorption Peaks (cm^−1)	Functional Groups/Interpretation	References
1	3474.87	Formation of a bond between selenium ions and hydroxyl groups (-OH(R-O-H)) and/or amine groups (−NH2) in the fruit extract of V. arctostaphylos L.	Safdar et al. (2023)
2	1645.18	Involvement of protein’s amide-I bond (CO)
3	1617.22	Connection of alkenes due to the stretching vibration of hydrocarbons (CC)	Kora and Rastogi (2016)
4	1485.32	C-H symmetric vibrations
5	720.32	Alkene bending vibration

Figure 4. Analysis of morphology and elements distribution. (A) the FESEM analysis of synthesized SeNPs revealed uniformly distributed round shaped particles with an average size of around 50 ± 1.23 nm. (B) EDX-synthesised SeNPs’ elemental distribution.

Figure 5. DLS and ZP tests of green synthesized SeNPs. (A) Determination of Size (nm) using DLS. (B) Detection of zeta potential average (mV).

Table 2. Estimation of the MIC and MBC by resazurin microdilution method of green synthesized SeNPs and ciprofloxacin antibiotic against Staphylococcus aureus, Escherichia coli, and Corynebacterium diphtheriae

Se NPs	OD (570 nm) Resazurin Dye (Mean±SD)
Concentration (mg/mL)	S. aureus	E. coli	C. diphtheriae
0.5	0.71 ± 0.05	1.03 ± 0.08	0.72 ± 0.03
0.25	0.90 ± 0.07	1.13 ± 0.04	0.86 ± 0.09
0.125	1.47 ± 0.07	1.59 ± 0.06	1.37 ± 0.03
0.0625	1.86 ± 0.05	2.16 ± 0.04	1.86 ± 0.04
0.03125	2.19 ± 0.03	2.28 ± 0.02	2.18 ± 0.02
0.015625	2.23 ± 0.03	2.39 ± 0.01	2.24 ± 0.04
Ciprofloxacin (^+Ve C)*	0.47 ± 0.04	0.70 ± 0.02	0.49 ± 0.04
Zero (^−Ve C)*	2.17 ± 0.03	2.29 ± 0.01	1.87 ± 0.02
MIC (mean)	0.03125 ± 0.04	0.03125 ± 0.04	0.0625 ± 0.04
MBC (mean)	0.015625 ± 0.04	0.015625 ± 0.04	0.03125 ± 0.04

Notes: *+Ve C=Positive control; -Ve C= Negative control; MIC= minimum inhibitory concentration; MBC= Minimum Bactericidal Concentration.

Table 3. Estimation of the MIC and MBC by resazurin microdilution method of extract and ciprofloxacin antibiotic against Staphylococcus aureus, Escherichia coli, and Corynebacterium diphtheriae

Extract	OD (570 nm) Resazurin Dye (Mean±SD)
Concentration (mg/mL)	S. aureus	E. coli	C. diphtheriae
0.5	1.53 ± 0.04	1.77 ± 0.04	1.49 ± 0.01
0.25	1.74 ± 0.04	1.98 ± 0.09	1.68 ± 0.05
0.125	2.02 ± 0.07	2.35 ± 0.01	2.02 ± 0.05
0.0625	2.29 ± 0.02	2.47 ± 0.05	2.27 ± 0.04
0.03125	2.29 ± 0.02	2.32 ± 0.04	2.28 ± 0.02
0.015625	2.19 ± 0.06	2.25 ± 0.04	2.21 ± 0.03
Ciprofloxacin (^+Ve C)*	0.47 ± 0.05	0.70 ± 0.02	0.49 ± 0.04
Zero (^−Ve C)*	2.17 ± 0.04	2.28 ± 0.02	1.87 ± 0.03
MIC	0.125 ± 0.04	0.25 ± 0.02	0.125 ± 0.04
MBC	0.0625 ± 0.02	0.125 ± 0.04	0.0625 ± 0.02

Notes: *+Ve C=Positive control; -Ve C= Negative control; MIC= minimum inhibitory concentration; MBC= Minimum Bactericidal Concentration.

Figure 6. The zone inhibition effect of different concentrations of the green synthesized nanoparticles and the fruit extract on the Staphylococcus aureus, Escherichia coli, and Corynebacterium diphtheriae.

Table 4. The zone inhibition effect of different concentrations of the green synthesized nanoparticles and the Vaccinium arctostaphylos (L.) fruit extract on the Staphylococcus aureus, Escherichia coli, and Corynebacterium diphtheriae

Se NPs	Zone Inhibition (mm) (Mean±SD)
Concentration (mg/mL)	S. aureus	E. coli	C. diphtheriae
0.5	36 ± 1.63	29.66 ± 1.25	33 ± 1.63
0.25	30.33 ± 0.47	25 ± 0.82	25.67 ± 1.25
0.125	26.66 ± 0.47	20.33 ± 1.25	22.67 ± 0.47
0.0625	18 ± 1.69	17.33 ± 0.47	18 ± 0.82
0.3125	16 ± 0.82	12 ± 0.87	13.67 ± 1.25
Ciprofloxacin (^+Ve C)*	34 ± 0.94	22 ± 0.82	39 ± 0.85

Notes: *+Ve C=Positive control.

Table 5. The zone inhibition effect of different concentrations of the Vaccinium arctostaphylos (L.) fruit extract on the Staphylococcus aureus, Escherichia coli, and Corynebacterium diphtheriae

Extract	Zone Inhibition (mm) (Mean±SD)
Concentration (mg/mL)	S. aureus	E. coli	C. diphtheriae
0.5	11.67 ± 0.943	7.67 ± 1.247	12 ± 1.633
0.25	7 ± 1.42	5.33 ± 1.25	8 ± 0.82
0.125	0.37 ± 0.47	NS	4.67 ± 0.47
0.0625	NS*	NS	2.67 ± 0.47
0.3125	NS	NS	NS
Ciprofloxacin (^+Ve C)*	34 ± 0.94	22 ± 0.82	39 ± 0.85

Notes: *+Ve C=Positive control; NS= Not Specified.

Table 6. Determination of the viable, injured, and dead cell percentage after treatment with different concentrations of green synthesized SeNPs using propidium iodide stain and flow cytometry

Green synthesized SeNPs
Bacteria	Concentration	Viable bacteria	Membrane Damaged bacteria	Injured bacteria	Dead bacteria
Escherichia coli	0.5 MIC	57.30%	23.70%	2.47%	16.53%
	MIC	32.60%	38.20%	11.70%	17.50%
	2 MIC	12.10%	49.20%	14.40%	24.40%
Staphylococcus aureus	0.5 MIC	44.60%	33.70%	1.89%	19.81%
	MIC	24.20%	32.60%	9.80%	33.40%
	2 MIC	7.10%	27.10%	21.20%	44.60%
Corynebacterium diphtheriae	0.5 MIC	50.12%	24.81%	4.91%	20.16%
	MIC	28.32%	30.10%	12.09%	29.49%
	2 MIC	13.80%	50.41%	15.50%	20.68%

Figure 7. Determination of the live/dead percentage of Escherichia coli after treating with different concentrations of green synthesized SeNPs and Vaccinium arctostaphylos (L.) extract using propidium iodide stain and flow cytometry.

Figure 8. Determination of the live/dead percentage of Staphylococcus aureus after treating with different concentrations of green synthesized SeNPs and Vaccinium arctostaphylos (L.) extract using propidium iodide stain and flow cytometry.

Figure 9. Determination of the live/dead percentage of C. diphtheriae after treating with different concentrations of green synthesized SeNPs and Vaccinium arctostaphylos (L.) extract using propidium iodide stain and flow cytometry.

Table 7. Determination of the viable, injured, and dead cell percentage after treatment with different concentrations of Vaccinium arctostaphylos (L.) fruit extract using propidium iodide stain and flow cytometry

	Vaccinium arctostaphylos L. Extract
Bacteria	Concentration	Viable bacteria	Membrane Damaged bacteria	Injured bacteria	Dead bacteria
Escherichia coli	0.5 MIC	97.9 %	0.64%	0.57%	0.87%
	MIC	95.77%	1.71%	1.32%	1.2%
	2 MIC	94.18%	2.1%	1.52%	2.2%
Staphylococcus aureus	0.5 MIC	97.92%	0.66%	0.94%	0.48%
	MIC	96.18%	1.32%	1.87%	0.63%
	2 MIC	95.61%	1.25%	1.91%	1.23%
Corynebacterium diphtheriae	0.5 MIC	98.11%	0.54%	0.91%	0.44%
	MIC	97.69%	0.59%	0.88%	0.84%
	2 MIC	95.61%	1.25%	1.91%	1.23%

Safdar, M., Aslam, S., Akram, M., Khaliq, A., Ahsan, S., Liaqat, A., Mirza, M., Waqas, M., & Ak, W. (2023). Bombax ceiba flower extract mediated synthesis of Se nanoparticles for antibacterial activity and urea detection. World Journal of Microbiology & Biotechnology, 39(3), 80. https://doi.org/10.1007/s11274-022-03513-z

 PubMed Web of Science ®Google Scholar

Kora, A. J., & Rastogi, L. (2016). Biomimetic synthesis of selenium nanoparticles by Pseudomonas aeruginosa ATCC 27853: An approach for conversion of selenite. Journal of Environmental Management, 181, 231–236. https://doi.org/10.1016/j.jenvman.2016.06.029

 PubMed Web of Science ®Google Scholar

Data availability statement

The data that support the findings of this study are available from the corresponding author on reasonable request