Bactericidal Properties of Plants-Derived Metal and Metal Oxide Nanoparticles (NPs).

Nanoparticles (NPs) are nano-sized particles (generally 1⁻100 nm) that can be synthesized through various methods. The wide range of physicochemical characteristics of NPs permit them to have diverse biological functions. These particles are versatile and can be adopted into various applications, particularly in biomedical field. In the past five years, NPs’ roles in biomedical applications have drawn considerable attentions, and novel NPs with improved functions and reduced toxicity are continuously increasing. Extensive studies have been carried out in evaluating antibacterial potentials of NPs. The promising antibacterial effects exhibited by NPs highlight the potential of developing them into future generation of antimicrobial agents. There are various methods to synthesize NPs, and each of the method has significant implication on the biological action of NPs. Among all synthetic methods, green technology is the least toxic biological route, which is particularly suitable for biomedical applications. This mini-review provides current update on the antibacterial effects of NPs synthesized by green technology using plants. Underlying challenges in developing NPs into future antibacterials in clinics are also discussed at the present review.

1. Nanoparticles and Green Technology

Nanoparticles (NPs), also known as nanomaterials, are small on a molecular scale and have various physical and chemical properties. Some examples of NPs are made up of metals ions such as Au, Ag, Pd, Pt, Zn, Fe, and Cu, and metal oxides such as Ag2O, NiO, ZnO, CuO, FeO, and CeO2. The advancement of nanotechnology has also given rise to the development of various nanocomposites, which are multiphase solid materials consisting of multiple types of NPs and polymers to improve single-metal biological effects and overcome structure-function related issues [1]. Multiple biological actions of the NPs such as antibacterial [2,3], antioxidant [2,4], anticancer [5,6], antifungal [5,7], antiviral [8,9], antiparasitic [10,11] and anti-inflammatory activities [12,13] have been associated with their highly diverse chemistry-rich characteristics [14]. There are myriad ways of synthesizing NPs, including physical (e.g., vapor deposition [15], sputter deposition [16], electric arc deposition [17], ion beam technique [18], molecular beam epitaxy [19], melt mixing [20]), chemical (e.g., co-precipitation [21], sol-gel [22], microemulsions [23], sonochemical synthesis [24] UV-initiated photoreduction [25]), and biological (e.g., synthesis using plant extracts [26], microorganisms [26], algae [27], fungi [28], animals [29] or agricultural waste [30], enzymes [31]) methods as well as hybrid methods [32] (Figure 1). There are advantages and limitations for each synthetic method, and the choice of method is selected based on the downstream applications.

Figure 1

Current synthetic methods of nanoparticles (NPs) and their biomedical applications. The core methods used for NPs construction are divided into physical, chemical, and biological methods. The generated NPs can be utilized in various biomedical applications including imaging, biosensors, diagnostics, biological therapies, drug delivery, bioconjugation, and hyperthermia.

Due to their small size and improved cell-penetrating features, NPs are extremely useful in various biomedical applications including sensing [33,34], imaging [33,35], diagnostics [36,37], drug/compound delivery system [38,39], bioconjugation [40,41], hyperthermia [42,43], and biological therapies [33,36] (Figure 1). In the past few years, NPs have been widely used to improve sensing and imaging techniques mainly due to their remarkable localization capability [33]. Some other additional advantages using NPs include flexibility in surface modification of NP [44], easy control of size [45], and production of highly degradable NPs in vivo [46]. Moreover, NPs are widely used in bio-conjugation and combination with drugs and compounds as well as in facilitating their delivery to target [47]. In a review paper authored by Werengowska-Ciećwier et al., bioconjugation of various drugs and NPs as well as their detailed chemistry have been described [47]. The same review also discussed the application of NPs in drug delivery system for targeted therapies. The potential use of NPs in hyperthermia mainly in cancer cell killing have been extensively explored [48,49]. The use of magnetic hyperthermia is one of the hot approaches [48,50]. Using magnetic NPs, heat generation can be controlled and specifically target and kill cancer cells while limiting damage to the surrounding normal tissue [50,51]. As chemo- and radiotherapies are standard cancer treatments, it is anticipated that the use of NPs in combination with current treatments could enhance the treatment outcome while reducing side effects of chemo-and radio-therapies [51]. Similarly, NPs have also been shown to be effective against other infectious diseases such as Pseudomonas aeruginosa [52] and Escherichia coli [53] infections in addition to cancers. This highlights the important role of NPs in medical and biomedical application in short future.

Generally, synthesis of NPs by biological routes has several advantages over both physical and chemical methods. First, the process is relatively simple, easy to scale up, efficient, and it consumes lesser energy [. Second, green technology is environmentally friendly as it uses lesser toxic chemicals and generating safer products and byproducts [54]. The green method is suitable and applicable for production of food, pharmaceuticals, and cosmetics [55]. Comparatively, the green method produces NPs that are generally less toxic, the end products being more suitable for a wide range of biomedical applications [26,56]. Chemical and physical methods can be very costly and usually involve the use of toxic and hazardous chemicals which tend to be more toxic to human cells. Additionally, the green method does not strictly require high temperature, pressure, and energy [57]. However, several parameters such as pH, chemical concentration, reaction time and reaction temperature are critical to consistently produce biologically functional NPs [58,59,60,61]. Furthermore, unlike using microbial system, generation of NPs from plants do not have to maintain microbial culture hence reducing the costs for microorganism isolation and culture media preparation [54]. The NPs generated from plants generally have size ranging from 1 to 100 nm (Table 1). There are also relatively large NPs that have sizes ranging from 100 to 500 nm [2,62,63]. NPs generated by all type of methods result in impurities that mainly cause toxicity to the human cells. These impurities can be removed by dialysis and filtering. The plant-derived NPs are less toxic, as shown from toxicity testing across several mammalian cell lines such as NIH3T3 [4], HEK293 [3], and primary cells such as peripheral blood mononuclear cells (PBMCs) [63] and rat aortic vascular smooth muscle cells (VSMCs) [4].

Table 1

Antibacterial effects of nanoparticles synthesized by green method using plants against various bacteria reported in PubMed-indexed publications from 2016 to 2017.

Nanoparticles	Size (nm)	Source	Scientific Name	Common Name	Target Bacteria	References
Ag-NPs	26–28	Leaves	Coleus aromaticus	Cuban oregano	Escherichia coli (E. coli), Staphylococcus aureus (S. aureus)	[78]
	12.46	Leaves	Salvinia molesta	Kariba weed	E. coli, S. aureus	[79]
	70.7–192.02	Leaves	Aloe vera	Aloe	Pseudomonas aeruginosa (P. aeruginosa), Streptococcus epidermidis (S. epidermidis)	[63]
	5–50	Leaves	Mentha pulegium	Pennyroyal	E. coli, S. aureus, Streptococcus pyogenes (S. pyogenes)	[5]
	5–40	Leaves	Cucurbita pepo	Summer squash	E. coli, S. aureus, Bacillus cereus (B. cereus), Listeria monocytogenes (L. monocytogenes), Salmonella typhi (S. typhi), Salmonella enterica (S. enterica)	[80]
	112.6	Crude	Ammania baccifera	Monarch redstem	S. aureus, P. aeruginosa, MRSA	[81]
	10–70	Oil cake	Cocos nucifera	Coconut	Aeromonas sp., Acinetobacter sp., Citrobacter sp.	[82]
	3.2–16	Seeds	Pimpinella anisum	Aniseed	K. pneumonia, P. aeruginosa, S. typhiStreptococcus pyogenes (S. pyogenes), Acinetobacter baumannii (A. baumannii)	[69]
	2–25	Crude	Matricaria camomilia	Camomile	E. coli, S. aureus, Bacillus subtilis (B. subtilis), P. aeruginosa	[83]
	50	Crude	Salvadora persica L.	Toothbrush tree	E. coli, S. aureus	[84]
	25	Rhizomes	Zingiber officinale	Ginger	E. coli, S. aureus, K. pneumonia	[71]
	20	Leaves	Gloriosa superba	Flame lily	E. coli, B. subtilis	[85]
	5–25	Leaves	Parkia roxburghii	Tree bean	E. coli, S. aureus	[86]
	10–20	Tubers	Dioscorea alata	Yams	E. coli, Staphylococcus auricularis (S. auricularis)	[87]
	35–42.5	Powder	Theobroma cacao	Cacao	E. coli, S. aureus, Staphylococcus epidermidis (S. epidermidis), P. aeruginosa	[88]
	10–50	Leaves	Adathoda vasica Linn	Vasaka	Vibrio parahaemolyticus (V. parahaemolyticus)	[89]
	14.63	Crude	Eleutherococcus senticosus	Siberian ginseng	E. coli, S. aureus, V. parahaemolyticusBacillus anthracis (B. anthracis)	[90]
	20–30	Seeds	Coffea arabica	Arabian coffee	E. coli, S. aureus	[70]
	20–100	Leaves	Sonneratia apetala	Sonneratia mangrove	Shigella flexneri (S. flexneri), E. coli, S. aureus, Vibrio cholera (V. cholera), S. epidermidis, B. subtilis	[62]
	50–400	Bark	Heritiera fomes	Sundari	E. coli, S. aureus, V. cholera, S. epidermidis, B. subtilis	[62]
	3–6	Crude	Allium sativum L.	Garlic	E. coli, E. faecalis, Bacillus cereus (B. cereus), S. flexneri	[91]
	3–22	Crude	Zingiber officinale Rosc.	Ginger	E. coli, E. faecalis, B. cereus, S. flexneri	[91]
	3–18	Crude	Capsicum frutescens L.	Cayenne pepper	E. coli, E. faecalis, B. cereus, S. flexneri	[91]
	10–20	Roots	Salvadora persica L.	Toothbrush tree	E. coli, S. aureus, P. aeruginosa, Micrococcus luteus (M. luteus)	[92]
	5–30	Crude	Rumex dentatus	Toothed dock	P. aeruginosa, Bacillus thuringiensis (B. thuringiensis)	[93]
	49	Flowers	Millettia pinnata	Karanja	E. coli, P. aeruginosa, Proteus vulgaris (P. vulgaris), S. aureus, K. pneumonia	[75]
	16.4	Seeds	Pongamia pinnata	Seashore Mempari	E. coli	[94]
	5–60	Rhizomes	Dryopteris crassirhizoma	Japanese fern	P. aeruginosa, B. cereus	[72]
	1–69	Leaves	Ficus religiosa	Peepul tree	E. coli, B. subtilis, S. typhi, Pseudomonas fluorescens (P. fluorescens)	[95]
	12–38	Powder	Styrax benzoin	Benzoin gum	E. coli, P. aeruginosa, S. aureus	[96]
	6.4–27.2	Callus	Taxus yunnanensis	Himalayan yew	E. coli, S. aureus, S. paratyphi, B. subtilis	[76]
	10–20	Ginseng berry	Panax ginseng	Meyer berries	E. coli, S. aureus	[4]
	45.26	Corn leaves	Zea mays L.	Maize	E. coli, S. aureus, S. typhimurium, L. monocytogenes, B. cereus	[97]
	20–80	Shoot tip	Caesalpinia mimosoides Lam.	Mimosa thorn	E. coli, L. monocytogenes	[98]
	37	Leaves	Coriandrum sativum	Coriander	Propionibacterium acnes (P. acnes)	[99]
	22.89	Aerial parts	Artemisia tournefortiana	-	E. coli, B. subtilis, S. pyogenes, P. aeruginosa	[100]
	20	Leaves	Derris trifoliata	Common derris	E. coli, S. aureus, S. enterica, Vibrio parahaemolyticus (V. parahaemolyticus)	[101]
	121	Roots	Rheum palmatum	Chinese Rhubarb	S. aureus, P. aeruginosa	[66]
	12.46	Leaves	Salvinia molesta	Giant salvinia	E. coli, S. aureus	[102]
	32.5	Roots	Decalepis hamiltonii	Indian Sarsaparilla	E. coli, S. aureus, P. aeruginosa, B. cereus, B. licheniformis	[67]
	16	Crude	Heterotheca inuloides	Mexican arnica	E. coli, S. aureus	[103]
	10–30	Fruit juices	Vitis vinifera and Solanum lycopersicum	Grape and tomato	Pseudomonas septica (P. septica), S. aureus, M. luteus, Enterobacter aerogenes (E. aerogenes), B. subtilis, S. typhi	[104]
	2.1–45.2	Callus	Artemisia annua	Sweet wormwood	Arthrobacter arilaitensis (A. arilaitensis), Staphylococcus equorum (S. equorum), Microbacterium oxydans (M. oxydans)	[77]
	15.2	Bark	Crataeva nurvala	Ayurveda	P. aeruginosa	[105]
	6–8	Fruit	Tamarindus indica	Tamarind	P. aeruginosa, S. aureus, M. luteus,Enterobacter aerogenes (E. aerogenes)B. subtilis, B. cereus, S. typhi	[106]
	12–80	Callus	Nicotiana tabacum	Tobacco	E. coli, Agrobacterium rhizogenes (A. rhizogenes)	[107]
	410–450	Leaves	Lantana camara	Verbanaceae	E. coli, S. aureus, P. aeruginosa	[2]
	25–40	Crude	Actinidia deliciosa	Kiwi fruit	P. aeruginosa	[6]
	15–28	Stem bark	Ficus krishnae	Krishna fig	E. coli, S. aureus, S. typhimurium	[68]
	2–15	Callus	Catharanthhus roseus	Madagascar periwinkle	E. coli	[108]
	28	Leaves	Convolvulus arvensis	Field bindweed	E. coli	[109]
	25	Leaves	Artemisia vulgaris	Common wormwood	E. coli, S. aureus, P. aeruginosa, K. pneumonia, Haemophilus influenza (H. influenza)	[110]
	20	Leaves	Costus afer	-	E. coli, S. aureus, P. aeruginosa, K. pneumonia, B. subtilis	[111]
	23–42	Leaves	Exocoecaria agallocha	Blinding tree	P. aeruginosa, S. aureus, S typhi, B. cereus	[112]
	10–80	Aerial parts	Anthemis atropatana	-	E. coli, S. aureus, P. aeruginosa, S. pyogenes	[113]
	40–60	Leaves	Arbutus unedo	Strawberry tree	E. coli, S. epidermis, B. subtilis, P. aeruginosa	[114]
	88.8	Leaves	Cicer arietinum	Chickpea	E. coli, P. aeruginosa	[115]
	5–30	Leaves	Taraxacum officinale	Dandelion	Xanthomonas axonopodis (X. axonopodis), Pseudomonas syringae (P. syringae)	[116]
	20–44.49	Leaves	Prosopis cinerraria	Khejri tree	E. coli, K. pneumonia, S. epidermidis	[117]
	15–25	Leaves	Croton bonplandianum	Bantulasi	E. coli, S. aureus	[118]
Au-NPs	5–10	Ginseng berry	Panax ginseng	Meyer berries	E. coli, S. aureus	[4]
	5–25	Leaves	Parkia roxburghii	Tree bean	E. coli, S. aureus	[71]
	10–75	Leaves	Ginkgo biloba Linn	Ginkgo tree	Brevibacterium linens (B. linens)	[119]
	3–37	Leaves	Nigella arvensis	Love-in-a-mist	E. coli, S. aureus, P. aeruginosa, Serratia marcescens (S. marcescens), B. subtilis, S. epidermidis	[120]
	25	Fruit	Dimocarpus longan	Longan	S. aureus, B. subtilis, E. coli	[64]
	5–25	Leaves	Cerasus serrulata	Japanese cherry	E. coli, S. aureus	[121]
	7–20	Crude	Actinidia deliciosa	Kiwi fruit	P. aeruginosa	[6]
	8–25	Peel	Citrus maxima	Pomelo	E. coli, S. aureus	[122]
	20–30	Crude	Coptis chinensis	Gold thread	Drug-resistant E. coli	[123]
Ag2O-NPs	42.7	Roots	Ficus benghalensis	Banyan	Streptococcus mutans (S. mutans), Lactobacilli sp.	[124]
NiO-NPs	9.69	Crude	Moringa oleifera	Drumstick tree	S. aureus, S. pneumonia, Escherichia hermannii (E. hermannii), E. coli	[125]
	10–20	Leaves	Eucalyptus globulus	Blue glum	E. coli, S. aureus, MRSA, P. aeruginosa	[126]
ZnO-NPs	20.06	Leaves	Prunus x yedoensis Matsumura	Yoshino cherry	B. linens, S. epidermidis	[127]
	400–500	Leaves	Sonneratia apetala	Sonneratia mangrove	S. flexneri	[62]
	47.27	Leaves	Laurus nobilis	Bay tree	S. aureus, P. aeruginosa	[128]
	50	Fruit	Rosa canina	Dog rose	E. coli, L. monocytogenes, P. aeruginosa	[65]
	-	Leaves	Lobelia leschenaultiana	Lobelia	P. aeruginosa, Shigella sonnei (S. sonnei), P. vulgaris, V. parahaemolyticus	[129]
	27–85	Fruit, seed, and pulp	Citrullus colocynthis L.	Schrad	MRSA, P. aeruginosa, E. coli, B. subtilis	[130]
Cu-NPs	21–30	Leaves	Terminalia catappa	Tropical almond	E. coli	[131]
	18.9–32.09	Leaves	Prosopis cineraria	Khejri tree	E. coli, K. pneumonia, S. epidermidis	[117]
CuO-NPs	30 –222.5	Leaves	Seidlitzia rosmarinus	Keliab	E. coli, S. aureus	[132]
Pt-NPs	2–7	Crude	Taraxacum laevigatum	Red-seeded dandelion	P. aeruginosa, B. subtilis	[133]
FeO-NPs	-	Peel	Punica granatum	Pomegranate	P. aeruginosa	[134]
Pd-NPs	5	Leaves	Sapium sebiferum	Chinese tallow tree	S. aureus, P. aeruginosaBacillus subtilis	[135]
	30	Seeds	Phyllanthus emblica	Indian Gooseberry	S. aureus, P. aeruginosa, B subtilisProteus mirabilis	[136]
	27	Peel	Moringa oleifera	Horseradish tree	E. coli, S. aureus	[73]
CeO2-NPs	45	Peel	Moringa oleifera	Horseradish tree	E. coli, S. aureus	[74]
	24	Leaves	Olea europaea	Olive	E. coli, S. aureus, K. pneumonia, P. aeruginosa	[137]
Ce2O3-NPs	8.6–10.5	Crude	Euphorbia amygdaloides	Wood spurge	Pediococcus acidilactici (P. acidilactici)	[138]
Pectin/Ag-NPs	20–80	Shoot tip	Caesalpinia mimosoides Lam.	Mimosa thorn	E. coli, L. monocytogenes	[98]
Ag/Ag2O-NPs	8.2–20.5	Leaves	Eupatorium odoratum	Christmas bush	E. coli, S. typhi, S. aureus, B. subtilis	[139]
Ag/Au-NPs	10	Leaves	Gloriosa superba	Flame lily	E. coli, B. subtilis	[85]
Chitosan/Ag-NPs	378–402	Crude	Rumex dentatus	Toothed dock	P. aeruginosa, B. thuringiensis	[140]
Chitosan/CeO2-NPs	3.61–24.4	Leaves	Sida acuta	Common wireweed	E. coli, B. subtilis	[141]
PCL/Cur/GLE-Ag-NPs	200	Leaves	Vitis vinifera	Grape	E. coli, S. aureus, P. aeruginosa, B. subtilis, S. enterica	[93]
GLE-Ag-NPs	30	Leaves	Vitis vinifera	Grape	E. coli, S. aureus, P. aeruginosa, B. subtilis, S. enterica	[93]
Cellulose/Cu-NPs	20–40	Leaves	Terminalia catappa	Tropical almond	E. coli	[131]
Ag-MnO2-NPs	5–40	Leaves	Cucurbita pepo	Summer squash	E. coli, S. aureus, B. cereus, L. monocytogenes, S. typhi, S. enterica	[142]

There is an extensive list of NPs possessing various biological actions as mentioned above. However, the focus of this review emphasizes the NPs’ antibacterial activities. This short review provides updates on the recent NPs with promising antibacterial activity synthesized by green technology using whole plant or other extracts of plants such as leaves [5,63], fruits [64,65], roots [66,67], barks [62,68], seeds [69,70], rhizomes [71,72], peels [73,74], flowers [75], and callus [76,77]. Here, we also discuss the challenges of adopting NPs for clinical applications.

2. Bactericidal Properties and Synergistic Enhancement of Common Antibiotics

NPs derived from plants that show promising antibacterial activities have high potential to be developed into future antibacterials mainly due to their low toxic effects [143]. NPs have been previously reported to inhibit gram-positive bacteria such as Staphylococcus spp. [4,5], Streptococcus spp. [113,124], and Bacillus spp. [114,120], and gram-negative bacteria such as Escherichia spp. [97,122], Pseudomonas spp. [6,68], Salmonella spp. [68,104], Shigella spp. [97,129], Proteus spp. [75,136], and Vibrio spp. [101,129]. More promisingly, NPs have also been shown to inhibit antibiotic-resistant bacteria such as Methicillin-resistant S. aureus (MRSA) [81,130] and drug-resistant E. coli [123]. Table 1 summaries the antibacterial action of NPs reported in PubMed-indexed journals in the past two years (2016–2017). A total of 107 articles was obtained from Pub-Med search engine through National Center for Biotechnology Information (NCBI) website using four keywords (nanoparticles, green synthesis, plant, and antibacterial) [144]. Out of the 107, 17 articles have been excluded as they do not contain relevant information for this review. These articles included retracted papers, review papers, and other non-plant-derived NP research papers. The remaining 90 articles are reviewed, and the information is tabulated in Table 1 and categorized based on the type of NPs. Since there are various synthetic methods to generate NPs as mentioned in Section 1, the protocol of constructing the NPs tabulated in Table 1 is different from one to another even though they are derived from the same part of extracts. Overall, the most reported NPs are Ag-NPs, Au-NPs, followed by other metal/metal oxide-based NPs and nanocomposites. Most of the NPs have size of less than 100 nm except for a few NPs as shown in Table 1 [2,62,63,81]. From Table 1, it is also shown that most of the NPs were produced from the plant’s leaves rather than other parts of the plants, regardless of type of NPs.

Table 2 shows both gram-positive and gram-negative bacterial species that have been targeted by various NPs, and the frequency of bacterial species being studied is tabulated. Gram-negative species that have been targeted the most are E. coli followed by P. aeruginosa, whereas gram-positive species that are mostly targeted are S. aureus followed by B. subtilis and B. cereus. Comparatively, the type and total number of gram-negative bacterial species that are being targeted are more than gram-positive species. This highlights the potential of NPs as antibacterial agents since they could effectively permeate and kill gram-negative bacterial species which are notorious of their difficult-to-penetrate multilayer membranes [145]. This notion can be further supported by recent finding by Acharya et al. which showed a potent killing of AgNPs towards K. pneumonia [146]. FE-SEM analysis demonstrated that the NPs overlaid with K. pneumonia with damaged cell surfaces and disrupted cells due to the interaction with NPs. Similarly, it has also been shown by SEM that AgNPs damaged E. coli by causing a large leakage on cell membrane and the bacteria were disorganized to several parts [147]. The potent antibacterial killings of NPs demonstrate their potential to be developed into antibacterials against gut-related bacteria (e.g., E. coli, P. aeruginosa, B. cereus, K. pneumoniae, S. flexneri, and S. typhi) and skin infection–related bacteria (S. aureus). The collective findings from Table 2 also show that Ag-NPs are the most bactericidal NPs against the gut bacteria such as E. coli, P. aeruginosa, B. cereus, K. pneumoniae, S. flexneri, S. pyogenes, S. typhi) and skin-related bacteria such as S. aureus and S. epidermidis. Followed by Ag-NPs, Au-NPs were reported to be effective in killing E. coli and S. aureus. Interestingly, Table 2 shows that some specific types of NPs were more effective against a particular bacterial type compared to others. For example, ZnO-NPs was effective against P. aeruginosa while Pd-NPs was effective against S. aureus.

Table 2

Gram-negative and gram-positive bacterial species targeted by NPs synthesized from plants via green technology.

Gram-Negative Species	Ag	Au	Cu	Pt	Pd	Ag2O	NiO	ZnO	CuO	FeO	CeO2	Ce2O3
E. coli	46	6	2	-	1	-	2	2	1	-	2	-
Drug-resistant E. coli	-	1	-	-	-	-	-	-	-	-	-	-
E. hermannii	-	-	-	-	-	-	1	-	-	-	-	-
P. fluorescens	1	-	-	-	-	-	-	-	-	-	-	-
P. aeruginosa	23	2	-	1	2	-	1	4	-	1	1	-
P. syringae	1	-	-	-	-	-	-	-	-	-	-	-
P. septica	1	-	-	-	-	-	-	-	-	-	-	-
K. pneumoniae	6	-	1	-	-	-	-	-	-	-	1	-
P. vulgaris	1	-	-	-	-	-	-	1	-	-	-	-
P. mirabilis	-	-	-	-	1	-	-	-	-	-	-	-
S. flexneri	4	-	-	-	-	-	-	1	-	-	-	-
S. sonnei	-	-	-	-	-	-	-	1	-	-	-	-
S. paratyphi	1	-	-	-	-	-	-	-	-	-	-	-
S. typhi	6	-	-	-	-	-	-	-	-	-	-	-
S. typhimurium	2	-	-	-	-	-	-	-	-	-	-	-
S. enterica	2	-	-	-	-	-	-	-	-	-	-	-
V. parahaemolyticus	3	-	-	-	-	-	-	1	-	-	-	-
V. cholera	2	-	-	-	-	-	-	-	-	-	-	-
Aeromonas sp.	1	-	-	-	-	-	-	-	-	-	-	-
Acinetobacter sp.	1	-	-	-	-	-	-	-	-	-	-	-
A. baumannii	1	-	-	-	-	-	-	-	-	-	-	-
Citrobacter sp.	1	-	-	-	-	-	-	-	-	-	-	-
E. aerogenes	2	-	-	-	-	-	-	-	-	-	-	-
A. rhizogenes	1	-	-	-	-	-	-	-	-	-	-	-
H. influenza	1	-	-	-	-	-	-	-	-	-	-	-
X. axonopodis	1	-	-	-	-	-	-	-	-	-	-	-
S. marcescens	-	1	-	-	-	-	-	-	-	-	-	-
Lactobacilli sp.	-	-	-	-	-	1	-	-	-	-	-	-
Gram-Positive Species	Ag	Au	Cu	Pt	Pd	Ag2O	NiO	ZnO	CuO	FeO	CeO2	Ce2O3
S. aureus	35	6	-	-	3	-	2	1	1	-	2	-
S. epidermidis	5	1	1	-	-	-	-	1	-	-	-	-
MRSA	1	-	-	-	-	-	1	1	-	-	-	-
S. pyogenes	4	-	-	-	-	-	-	-	-	-	-	-
S. mutans	-	-	-	-	-	1	-	-	-	-	-	-
S. pneumonia	-	-	-	-	-	-	1	-	-	-	-	-
B. thuringiensis	1	-	-	-	-	-	-	-	-	-	-	-
B. cereus	9	-	-	-	-	-	-	-	-	-	-	-
B. subtilis	11	2	-	1	2	-	-	1	-	-	-	-
B. licheniformis	1	-	-	-	-	-	-	-	-	-	-	-
B. anthracis	1	-	-	-	-	-	-	-	-	-	-	-
E. faecalis	3	-	-	-	-	-	-	-	-	-	-	-
L. monocytogenes	3	-	-	-	-	-	-	1	-	-	-	-
M. luteus	3	-	-	-	-	-	-	-	-	-	-	-
P. acnes	1	-	-	-	-	-	-	-	-	-	-	-
A. arilaitensis	1	-	-	-	-	-	-	-	-	-	-	-
M. oxydans	1	-	-	-	-	-	-	-	-	-	-	-
B. linens	-	1	-	-	-	-	-	1	-	-	-	-
P. acidilactici	-	-	-	-	-	-	-	-	-	-	-	1

In addition to exhibiting antibacterial effects, NPs derived from plants can also serve as carriers to deliver antibacterial molecules or drugs to the target cells either via conjugation or nanoemulsions, hence synergistically enhance the antibacterial effect [97,139]. For example, Kalita and coworkers demonstrated that a gold NP was able to enhance the bacterial killing effects of Amoxicillin against both gram-positive (Staphylococcus spp. and Bacillus spp.) and gram-negative bacteria (E. coli) [139]. Patra and colleagues reported the potential of silver NP synthesized from corn leaves of Zea mays in foodborne pathogenic bacterial killings when used in combination with Kanamycin and Rifampicin [97]. More interestingly, the NP was able to reverse the development of antibiotics resistance by killing MRSA clinical isolates in vitro and in vivo using murine MRSA infection models [148]. The exact mechanism of NPs’ synergism with antibiotics remains exploratory. It could be due to (a) generation of additional bactericidal Ag+ ions by NPs [149], (b) generation of bactericidal hydroxyl radicals by NPs [150], and (c) effective blocking of the efflux pump for drug-resistant bacterial killing [143]. This synergistic effect could ultimately help in reducing the dosage of antibacterials that may potentially toxic to host system.

The exact mechanisms of NPs against various bacteria remain unknown. There have been several studies supporting the possible mechanisms of bactericidal effects including (a) attachment of large number of NPs on bacterial surface that interrupts respiration and other permeability-dependent functions [97], (b) generation of electrostatic attraction between negatively charged bacterial cells and the positively charged NPs [151], (c) inactivation and degradation of bacterial essential proteins [152], and (d) breakage or damage of bacterial genes following the efficient penetration of NPs [153]. For example, it has been shown that silver NPs permeated into bacterial cells and resulted in significant DNA damages by interacting with sulphur- and phosphorus-containing compounds [154,155]. Similarly, it has also been shown that silver NPs released highly reactive Ag+ ions and radicals for the antibacterial effects [156]. These ions have also been reported to interact with sulphur-containing proteins in the bacterial cell wall that caused multiple functionality impairs [152]. Raffi and colleagues also showed that the silver NPs could inactivate the bacterial enzymes and generate toxic hydrogen peroxide leading to bacterial cell death [157].

3. Plant-derived Nanoparticles as Future Antibacterials

To date, numerous NPs have been approved by either Food and Drug Administration (FDA) in United States, or European Medicines Agency (EMA) in the European Union for various clinical applications including imaging (e.g., Resovist), iron-replacement therapy for anaemia treatment (e.g., Vifor), delivery of anticancer drugs (e.g., Onivyde and MEPACT) [158,159], vaccines for viral diseases (e.g., Epaval against hepatitis A and Inflexal V against influenza) [160,161], fungal infection (e.g., AmBisome) [162], and so on. However, none of the currently approved NPs are used for controlling bacterial infection [163]. The only liposomal NP formulations that is undergoing clinical trial is CAL02, which has been designed for bacterial pneumonia management [163]. In this section, we discuss the potential challenges of developing NPs into clinically approved antimicrobial agents. Table 3 summaries some of the limitations and challenges of using plant-derived NPs for antimicrobials development. Undoubtedly, the nano-scale size of NPs has facilitated the cell-penetrating capacity including crossing blood-brain barrier (BBB) [164,165], hence improving the target specificity and biological activity. However, the small size may be one of the challenges in the clinical trials. It has been reported that NPs have poor stability and bioavailability under physiological conditions when they are administered into host system [166]. Studies have shown that NPs have been targeted and degraded by various enzymes and proteins from human blood before reaching to target sites [167,168]. These significantly reduced the biological functions of NPs. To overcome this limitation, several modification methods have been adopted such as conjugation with stabilizer (e.g., serum albumin) [169,170], synthesis of stable nanoemulsions [169,170], modification of surface chemistry and functionalization [171], and development of composite NPs [172]. In addition to size, it has also been reported that the NPs’ shape and morphology could determine their biological actions and toxicity profile [172,173,174]. Some NPs have high tendency to form aggregates as a result of the particle surface chemistry. This may cause unwanted toxicity and drastically limit the access of NPs into the target cells [175]. Toxicity is also attributed to several characteristics of NPs (e.g., chemistry, retention rate, biodistribution, stability, and specificity), mode of administration, and target sites [176].

Table 3

Challenges of developing nanoparticles into clinically used antibacterial agents.

Structural Challenge
Size	Smaller size enhances the cell penetration, but may have decreased stability or bioavailability
Shape	Certain shape of NPs may improve the functionality due to total surface exposure area
Aggregate	NPs that form aggregate increase the overall particle size, hence limiting the cell permeation and may increase toxicity
Biological Challenge
Biodistribution	Poor dispersion due to limited entry (e.g., skin barrier)
Bioavailability	Poor bioavailability results in rapid loss of function
Specificity	High specificity results in less off-target effects and more effective
Clearance	High retention rate ensures the high efficiency
Toxicity	Accumulation of toxic materials may damage the host
Technological Challenge
Heterogeneity of human disease	Variation within disease may complicate treatment
Scale-up	Optimization of NPs synthesis and production with uniform size without aggregates in controlled and consistent fashion
Throughput	Synthesis of NP is multistep and laborious which does not allow high-throughput optimization
Prediction	Prediction using computer modelling on NP efficiency is extremely challenging
Industrial Challenge
Quantity	Large scale production may result in inconsistent size and physicochemical properties of NPs
Processes	Reproducible and consistent manufacturing processes requires modern technology and instrumentation
Quality	Continuous production of high level uniformity and functionality of NPs

There are numerous limitations for NPs productions for biomedical application (summarized in Table 3). As the biological functions of NPs highly depend on their shape, size, permeability, and physicochemical properties, manufacturing NPs in industrial scale must strictly adhere to the tight-controlled, consistent, and reproducible standard operating procedure (SOP) and Good Manufacturing Practice (GMP). Another challenge is the heterogeneity of diseases in human. The clinical effect of NPs on infected humans could be very complicated as different individuals present varied profiles (mainly due to the individual’s immunity) even though they have been infected from the same source. This will require detailed and organized planning to validate the clinical value of NPs. Following the increase of potential NPs, the experimental design that requires high throughput setup for biological screening is also increasingly demanding. This is also closely linked with automation that allows cost-effective NPs production and, computing and modelling technology that would predict NPs’ efficacies or toxicities on target cells. The improvement in high throughput screening and computation will surely boost the development of nanotechnology in biomedical applications.

From an industrial point of view, the chosen synthesis method of NPs-based antibacterials must be compatible for large-scale production. As most NPs possess complex chemical make-up and arrangement of components, retaining these key characteristics in the process of scaling up remains a huge challenge in the manufacturing field. Secondly, the production method has to consistently produce high quality (e.g., size, uniformity, and physicochemical properties) of NPs. The formulation process must be recorded meticulously to ensure high-level of reproducibility. The advancement of modern technology contributed from large high-tech companies and academia will continuously support the production of high-quality NPs in a consistent and timely fashion.

4. Conclusions and Future Perspectives

This mini-review provides a recent update on the NPs derived from plants that possess promising antibacterial action. Antibacterial NPs produced from green technology have great potential to be developed into future antibacterials and are able to synergistically enhance the efficacy of antibiotics. While the exact mechanisms remain unknown, a great effort is currently underway to produce a highly potent and robust NPs for clinical use. Understanding the bactericidal mechanism of NPs is also important to control and overcome the emerging issue of bacterial resistance to NPs [177]. As highlighted above, the limitations and potential challenges need to be overcome to maximize the use of NPs to clinical applications. Uniformity, stability, specificity, and toxicity of NPs are the main biological properties in deciding the fate of NPs in clinical application. From an industrial perspective, the development of sustainable process and modern instrumentation is crucial to produce a practical amount of NPs for clinical use [178]. The advancement of nanotechnology and biotechnology are anticipated to boost the use of NPs in biomedical applications in the future.