Nanobiopesticides: Sustainability Aspects and Safety Concerns

Abstract

The use of chemical pesticides has significantly improved crop yields and global food security but poses risks to environment and human health. To address this, nanobiopesticides, combining nanomaterials and biopesticide, have emerged as a potential alternative. Therefore, this article evaluates their sustainability and safety through a literature review using Scopus. The results indicate that nanobiopesticides offer advantages over conventional pesticides, including greater precision, controlled release, and reduced dosage requirements. An illustrative Life Cycle Assessment conducted in this study confirmed that they potentially offer more sustainability than commercial pesticides, showing reductions in environmental impacts from −6% to −99%. However, several gaps remain related to the effect of nanoparticles on non-target organisms and biodiversity, bioaccumulation, and environmental persistence in ecosystems, and their ecotoxicological safety. Additionally, regulatory frameworks in major agricultural markets are complex and fragmented, potentially hindering large-scale adoption. Currently, nanobiopesticides are commercially available in countries such as the U.S., India, and Brazil, primarily for pest control in crops like rice, maize, and vegetables. Their market presence is growing, yet widespread implementation will depend on clearer regulations and further research on long-term environmental impacts.

1. Introduction

Pesticides have greatly enhanced food security and production since they were first exploited over one hundred years ago [1]. The Green Revolution principles and, lately, the rapid rise in world population, urbanization, and environmental issues, have further posed the protection of crops and the improvement of agriculture yield as primary goals at the global level. These aspects, among others, prompted the use of pesticides worldwide, as witnessed by consumption amounts that passed from 1.23 kg per hectare in 1990 to over 2.37 kg in 2022. Central and South America are the leaders, accounting for about one-third of the total market (20.29 billion U.S. dollars by value) [2]. The Asia Pacific region ranked second, with 25.05 billion U.S. dollars in 2022, whereas Europe consumed about 473,000 metric tons of pesticides of lower agricultural land and population [3]. At the country level, the same trend can be observed, and Brazil, the United States, Indonesia, Argentina, China, and Vietnam are the nations most involved in pesticide consumption. Among the different classes of pesticides (i.e., herbicides, fungicides, bactericides, insecticides, plant growth regulators, and rodenticides), herbicides are the most frequently used, representing about 55% of the total products, whereas all the other classes stop below one million metric tons [2].

Although they are very useful in agriculture land management, the massive use of pesticides at the same time poses serious environmental hazards. Ecological burdens include harmful effects on non-target wildlife, thus altering species’ distribution and biodiversity [4]. Other alarming aspects are linked to soil, water, and food contamination [5]. Finally, several pesticides have been proven to be dangerous for human health, being related to both short- (headaches, nausea, etc.) and long-term adverse effects (cancer, reproductive harm, endocrine disruption, etc.) [6]. It follows that it is urgent to find different strategies for pest control, by replacing synthetic chemical pesticides with effective, affordable, and more sustainable products [7]. In this sense, organic agriculture could not be considered a permanent solution as it is more demanding, in comparison to conventional agriculture, from the soil requirement perspective. Other drawbacks that limit organic agriculture’s effectiveness against chemical pesticide dependence relate to social, economic, and infrastructural issues [8].

In recent decades, an appealing approach focusing on biopesticides came to light to overcome chemical pesticides. The basic concept aimed to take advantage of the great number of natural antagonists of pathogens and pests that can have different sources (microbial, bacteria, fungal, viral, etc.). They can also be plant-incorporate-protectants (active substances produced by plants after genetic modification) or biochemical pesticides (plant extracts, pheromones, insect growth regulators, etc.) [9]. Biopesticides have showed many useful traits compared to their synthetic counterparts, including higher specificity, lower environmental impact, integrated pest management compatibility, suitability for organic practices, and less harmful effects on wildlife and humans [10]. Nonetheless, several disadvantages still limit their widespread application in agriculture as they are thermally degradable, highly volatile, poorly persistent, less efficient, and more subject to environmental changes than chemical products. Moreover, their production process is rather expensive and their application in the field is quite uneven [11].

To overcome the abovementioned drawbacks without affecting all the positive features of biopesticides, the application of nanotechnologies offers promising possibilities. Owing to their inherent dimension, nanomaterials show advantageous features in comparison to bulk materials, mostly related to surface-to-volume area, size, solubility, permeability, and overall reactivity [12]. It follows that advanced nanomaterials can serve as carriers of biopesticides to ensure higher stability and bioavailability, higher effectiveness, adhesion and targeting properties, enhanced environmental safety, and controlled release of the active ingredient (AI) [13]. Therefore, in light of the above, this article aimed to identify and analyze the most recent applications of nanobiopesticides, describing the current state of the art and evaluating them in terms of both sustainability and safety. In more detail, the focus was on identifying the main biobased precursors used, the environmental benefits, safety requirements, and the main challenges that nanobiopesticides will and must still face, both in the present and in the future. To this end, a literature review was conducted, using the Scopus database, and articles that were available and relevant from 2015 to 2024 were examined. The results of this evaluation could be useful in providing knowledge to the scientific community, practitioners, and other stakeholders involved in integrated pest management and agriculture in general, contributing to a broader understanding of the potential and limitations of nano-biopesticides, and promoting informed decision-making for more sustainable cultivation practices.

2. Pesticides, Biopesticides, and Nanobiopesticides

The use of chemical pesticides, which have been used in agriculture for decades to control pests and diseases and regulate plant growth [14], has been instrumental in improving crop quality and yields, significantly enhancing food security in a rapidly growing global population.

However, the use of synthetic pesticides, despite their benefits in terms of increased productivity, poses significant risks to the environment and human health, two aspects strongly related by a cause-effect chain, mainly due to their persistent nature and potential for bioaccumulation [15]. From an environmental point of view, it is useful to mention some of the problems of chemical pesticides. For example, these substances, designed to eliminate pests, often also harm non-target species. Suffice it to say that, of all the pesticides applied to soil, only 2% affect target organisms, while about 98% affect other non-target organisms [16]. This could lead to contamination of ecosystems and degradation of air, water, and soil quality, leading to negative effects on wildlife, humans, and food security. For example, in Europe, pesticide use is estimated to reduce soil respiration by −35%, insect biomass by −70%, and farmland bird populations by −50% [16]. Furthermore, in both Europe and America, pesticides are responsible for a −30% decrease in bee populations [17]. However, a large proportion of pesticides are non-biodegradable, and thus penetrate the soil, altering its chemical and biological characteristics [18], and into groundwater and surface water [19], leading to biodiversity loss and pollution. Consequently, pesticide residues can be ingested by humans either through direct consumption of crops or indirectly through animals feeding on accidentally contaminated fodder [20], and, thus, enter the food chain. Consequently, the consumption of foods such as vegetables or meat from animals that contain pesticide residues can lead to diseases in humans, including cancer, diabetes, respiratory disorders, and neurological disorders, as shown by Zhou et al. (2025) [15]. Additionally, in this regard, Otorkpa et al. (2024) [21] estimate that approximately 385 million cases of acute pesticide poisoning occur globally each year, resulting in approximately 11,000 deaths. However, synthetic pesticides also face additional drawbacks, such as pest resistance and high purchase costs. In the first case, these are mutations that lead to a decrease in the ability of the pesticide to bind or block the activity of the target [22]. This is an issue that is widely known in the literature, which must nevertheless be addressed, since the spread of resistance can compromise the long-term effectiveness of pesticides and require ever-increasing dosages, resulting in increased environmental impacts and economic costs. In the second case, however, reference is made to prices that can reach as high as $900/gallon (about $237.75/L) [23], in some cases making their adoption economically unsustainable for many farmers, especially in developing countries. These high costs could limit access to them, further exacerbating inequalities in the agricultural sector. Therefore, due to the challenges and negative impacts of synthetic pesticides, alternative means of pest control have become necessary, leading to the development of biopesticides. This is a new category of pesticides, created and named by the US Environment Protection Agency (USEPA), which originates not only from natural precursors such as microorganisms, plants, animals, and minerals [24], but also antagonistic organisms, plant chemicals, viruses, fungi [25], etc. In recent years, mainly due to public health and environmental safety concerns and stringent regulations on pesticide residues in agricultural products, there has been a −2% annual decline in the use of synthetic pesticides, compared to a +10% increase in the use of biopesticides [26]. Biopesticides differ from synthetic pesticides in their natural origin, specific mode of action, precision in targeting a small number of pests, high selectivity, and the need for smaller quantities. However, one of their great advantages lies in their ability to preserve the well-being of ecosystems [27], which is why biopesticides are generally safer for the environment, people, and animals than chemical pesticides, since, for example, they can be easily removed from fruit and vegetables with a simple wash. More recently, research has evolved further through the development of nanobiopesticides, which combine nanomaterial and biopesticide technologies. In more detail, nanotechnology, as broadly defined in 1999 by the US National Science and Technology Council [28], is “the ability to work at the molecular level, atom by atom, to create large structures with fundamentally new molecular organization. The aim is to exploit these properties by gaining control of structures and devices at atomic, molecular, and supramolecular levels and to learn to efficiently manufacture and use these devices”. As a consequence, nanomaterials show intrinsic features such as: (i) dimensions between 1 and 100 nm in either X, Y, or Z direction; (ii) synthesis by chemical or physical processes with control at the molecular-scale level; and (iii) material building involving a construction set-like process. More recently, the European Commission stated in Recommendation 2011/696/EU [29] that a nanomaterial can be considered “a natural, incidental, or manufactured material containing particles, in an unbound state or as an aggregate or as an agglomerate and where, for 50% or more of the particles in the number size distribution, one or more external dimensions is in the size range 1 nm–100 nm”. Widely exploited in many fields (i.e., material science, medicine, transportation, energy, and the environment, among others) [30], nanotechnology has also proved its versatility in farming, with many applications mainly related to nanofertilizers, nanobiosensors, precision and/or sustainable agriculture, pollution management, agriculture diagnostics, and nanopesticides. In particular, by combining nanotechnology and biopesticides, new formulations were proposed to effectively control pests, at the same time reducing chemical pesticides risks and offering solutions to biopesticide limiting factors. More in detail, the formulation of nanobiopesticides involves the coupling of the natural-derived active ingredient (AI) with nanocarriers [11]. The latter can be solid (nanocrystals, nanospheres, polymeric nanoparticles, etc.) or liquid (emulsion, liposomes, etc.), with organic (lipids, biopolymers, carbon, etc.), inorganic (metal nanoparticles, silica nanoparticles, nanocrystals), or hybrid structure biopolymers conjugated with biomaterials) [31]. They can also show different forms (vesicular, particulate, or miscellaneous) and origins (plant, animal, microbial, agricultural wastes, mineral, etc.) [32]. Finally, the AI can be linked to the nanocarrier’s surface, loaded in its core using ligands, or trapped into the nanocarrier’s polymeric network [33]. Properly modulating the dimensions, shapes, and surface characteristics of the various nanocarriers can achieve different features and action properties. On the other hand, the stability, effectiveness, safety, cost, and commercial success of nanobiopesticides largely depends on the overall design and formulation of the final product [34].

3. Materials and Methods

For this analysis, a literature review was performed, using Scopus as the database. The search was carried out considering two searches in parallel characterized by the keywords ‘Nanobiopesticide’ (Search 1 or S1) in one search and ‘Nanobiopesticides’ (Search 2 or S2) in another search, resulting in 24 and 52 papers, respectively. The scientific production for S1 starts from 2015, while that for S2 starts from 2017. The inclusion and exclusion criteria for both searches are shown in Figure 1. Specifically, for S1, these were as follows. Of the initial 24 articles, 2 were discarded because, after reading them in their entirety, they did not deal with nanobiopesticides but only with biopesticides, without nanometric application. Of the 22 remaining articles, 7 were then discarded because they were not accessible.

These are chapters belonging to books (including, but not limited to, ‘Nano-biopesticides. Today and Future Perspectives’ and ‘Handbook of Agricultural Biotechnology: Volume I Nanopesticides‘) not available in open access. Similarly, of the 15 remaining articles, five were excluded because they again belonged to journals not available in open-access mode. Thus, Search 1 returned 10 articles. The same was carried out for Search 2. Of the initial 52 documents, 7 were discarded because they were not directly related to the topic of nanobiopesticides, but again, only to the topic of biopesticides, without nanometric application. Of the 45 papers identified, 34 were excluded for the same reason expressed above, i.e., because they were part of chapters belonging to books and journals that were, in all cases, not available in open-access mode.

Search 2 therefore ultimately returned 11 articles. Considering S1 and S2, the two searches returned 3 articles in common. Therefore, considering the 10 articles of S1 and the 11 of S2, and considering the 3 common articles, the analysis was conducted on a total of seventeen documents, which were read in full and analyzed. This number might seem small for a literature review, but it is in any case necessary to emphasize the small base of papers found on international databases, in addition to their limited accessibility. Nonetheless, given this, the results of this study could nevertheless provide a basis for some preliminary considerations on the current applications of nanobiopesticides, and in any case, could offer useful insights into initial evaluations in this field of research.

4. Results and Discussions

4.1. Literature Overview: Temporal and Spatial Distribution, Type of Source

The topic shows itself to be a relatively unexplored and relatively new field of research. As shown in Figure 2, the scientific production seems to start from 2015 (Search 1) with the article by Lallawmawma et al. (2015) [35], and is distributed as follows: For S1, there was 1 article in 2015, 2 in 2017, 6 in 2019, 2 in 2020, 3 in 2022 and 2023, and 7 in 2024. For S2, on the other hand, there was 1 article in 2017, 9 in 2019, 3 in 2020, 2 in 2021, 5 in 2022, 12 in 2023, and 20 in 2024, showing in both cases an increasing interest in recent years and denoting a rise in research.

Analyzing the entire scientific production since 2015 (S1 + S2), and excluding overlaps and joint articles, out of a total of 61 articles in the database, as shown in Figure 3, the Scopus publications are divided as follows: 27 journal articles (44%), 23 book chapters (38%), 6 reviews (10%), 3 conference papers (5%), and 2 books (3%).

This distribution may reflect an application interest in nanobiopesticides as much as academic and advanced research, as well as efforts to synthesize existing knowledge and disseminate it in conference settings. From a spatial perspective, however, the two most prolific countries seem to be Nigeria and India. For example, in S2, the search that returned the most articles, as shown in Figure 4, had 14 and 12 papers, respectively, which accounts for 36% of the scientific production for that search.

Similarly, in S1, India returned 6 articles and Nigeria 3, but in this case, it was not possible to merge the data of the two searches to get the total. As an example, considering S2, the reasons behind these numbers could be multiple. For instance, in the case of Nigeria, agriculture constitutes about 25% of its gross domestic product [36], representing a particularly important sector for the national economy. Thus, a large portion of the rural population is engaged in agriculture and depends on inorganic fertilizers and pesticides for agricultural production, showing a potential interest in the subject. Furthermore, some typical crops in Nigeria, such as maize [37], rice [38], cassava [39], groundnut [40] and cocoa [41], are highly susceptible to pest and disease attacks, which could cause major economic losses, and this vulnerability may have stimulated the exploration of more effective and sustainable alternatives, such as nanobiopesticides. However, another key factor could also be the country’s rich biodiversity. Indeed, Nigeria is rich in plants with bioactive properties that are high candidates for the production of nanobiopesticides, such as Moringa oleifera [42] and Azadirachta indica (neem) [43]. These plant extracts could represent an abundant and accessible source of precursors for the synthesis of nanobiopesticides, in fact offering an important advantage for their local development.

4.2. Analysis of Typology and Effectiveness of NPs

The literature, an overview of which is shown in

Table 1. Overview of studies analyzed.

Ref.	Year	Focus	Precursor/ Reducing Agent	NPs Size	Larvicidal Efficacy			Lethal Time	Target
					LC50	LC90	LC95	LT50
[35]	2015	AgNPs Biosynthesis	Jasminum nervosum	4–22 nm	57.40 µg/mL	N.S.*	144.35 µg/mL	2.24 h × 150 µg/L	Culex quinquefasciatus (Southern house mosquito)
				2–20 nm	82.62 µg/mL	N.S.	254.68 µg/mL	4.51 h × 150 µg/L
[44]	2017	AgNPs Biosynthesis	Artemisia vulgaris L.	89.76 nm	third-stage larvae: 156.55 ppm/12 h, 62.47 ppm/24 h fourth-stage larvae: 97.90 ppm/12 h, 43.01 ppm/24 h	third-stage larvae: 2506.21 ppm/12 h, 430.16 ppm/24 h. fourth-stage larvae: 1677.36 ppm/12 h, 376.70 ppm/24 h	N.S.	5.5 h × 150 µg/L	Aedes aegypti (Yellow fever mosquito)
[45]	2017	AgNPs Biosynthesis	Bacillus thuringiensis kurstaki (Btk).	2–100 nm	0.81 mg/mL (supernatant), 0.46 mg/mL (pellet)	N.S.	N.S.	N.S.	Trichoplusia ni (Cabbage looper)
					5.20 mg/mL (supernatant), 1.95 mg/mL (pellet)	N.S.	N.S.	N.S.	Agrotis ipsilon (Seedling Noctule)
[46]	2017	ZnNPs Biosynthesis	Bacillus thuringiensis	20 nm	10.71 μg/mL	N.S.	N.S.	N.S.	Callosobruchus maculatus (cowpea weevil)
[47]	2019	NPs encapsulated in succinic anhydride cross-linked chitosan	Azadirachta indica (neem)	271.6 nm	100% mortality at 0.3% (nanobiopesticide) vs. 81.67% (simple neem extract)				Spodoptera litura (tobacco cutworm)
[48]	2020	Lemongrass essential oil based nanobiopesticides	Cymbopogon flexuosus (lemongrass)	N.S.	N.S.	N.S.	N.S.	N.S.	N.S.
[49]	2020	NBP production from neem oil encapsulated in zein NPs	Azadirachta indica (neem)	198 ± 16 nm	0.375 µg/mL	0.859 µg/mL	N.S.	N.S.	Bemisia tabac (Silverleaf whitefly)
					0.210 µg/mL	0.715 µg/mL	N.S.	N.S.	Tetranychus urticae (Red spider mite)
					0.455 µg/mL	0.940 µg/mL	N.S.	N.S.	Acanthoscelides obtectus (Bean weevil)
[50]	2020	Nanospinosad preparation by using spiky silica hollow NPs to load spinosad	Biopesticide spinosad, derived from Saccharopolyspora spinosa	330–350 nm	100% mortality at 0.1 mg/cattle hair and 74% at 0.01 mg/cattle hair. More effective than smooth nanoparticles (mortality of 63% at 0.1 mg/hair and 27% at 0.01 mg/hair), commercial spinosad product (mortality of 35% at 0.1 mg/hair), and pure spinosad (mortality of 25% at 0.01 mg/hair)			N.S.	Rhipicephalus microplus (Asian blue tick)
[51]	2021	Lemongrass essential emulsion-based nanobiopesticides	Essential oils of Cymbopogon flexuosus (lemongrass)	70–125 nm	N.S.	N.S.	N.S.	N.S. but LT100 is 30 min. a 50 μL/mL	Lucilia cuprina (Australian sheep blowfly)
[52]	2022	Production of nano-encapsulated formulations of abamectin	Abamectin in polymeric nanoparticles	175–200 nm	Precise values are N.S., but 65–75% mortality of larvae is achieved within 24 h, which is higher than the commercial formulation.				Tuta absoluta (Tomato pinworm)
[53]	2022	Silicon dioxide (SiO2) nanoparticles modified with epoxy, silane, and amide groups	SiO2	10–20 nm	SiO2 combined with Bt shows an increase in mortality of +10–15% compared with Bt alone.			37% within 3 days when treated with a combination of SiO2 nanoparticles and Bt	Leptinotarsa decemlineata (Colorado potato beetle)
[54]	2022	Nanoemulsion production from essential oils of anise, fennel, and mint	Pimpinella anisum (Anise), Foeniculum vulgare (fennel) and Mentha piperita.	100–150 nm	No LC values are specified, but RC50, which is 3.25% for anise, 3.47% for fennel, and 7.95% for mint.				Bactrocera oleae (Olive fruit fly)
[55]	2023	ZnO-NPs synthesis	Azadirachta indica, Emblica officinalis (emblica), Allium sativum (garlic)	14–27 nm	Average mortality in nymphs of 36.69% on day 1 and 54.15% on day 3, higher than the average adult mortality of 29.09% on day 1 and 40.20% on day 3.				Myzus persicae (Green peach aphid)
[56]	2023	ZnO-NPs synthesis	Silybum marianum (milk thistle) seeds	5180 nm	Average mortality of 78 ± 0.57% after 72 h				Tribolium castaneum (Red flour beetle)
					Average mortality 74 ± 0.57% after 72 h.				Sitophilus oryzae (Rice weevil)
					Inhibition zone of 18 ± 0.4 mm.				Clavibacter michiganensis (Ring Rot)
					Inhibition zone of 25 ± 0.4 mm.				Pseudomonas syringae
					Inhibition zone of 21 ± 0.57 mm.				Fusarium oxysporum
					Inhibition zone of 19 ± 0.4 mm.				Aspergillus niger
[57]	2024	Moringa oleifera leaf extract-based nanobiopesticides synthesis	Extract of Moringa oleifera leaves stabilized with Polyvinylpyrrolidone (PVP).	174 nm	The nanobiopesticide shows a mortality of 83.00% ± 0.56 (after 72 h), +3.7% compared with the mortality of 79.30% ± 2.64 of the crude extract of Moringa oleifera.				Tribolium castaneum (Red flour beetle)
					The nanobiopesticide shows a mortality of 92.48% ± 3.12 (after 72 h), +11.3% compared with the mortality of 81.15% ± 2.97 of the crude extract of Moringa oleifera.				Rhyzopertha dominica (Lesser grain borer)
[58]	2024	Mentha piperita-based nanobiopesticides synthesis	Mentha piperita	259,8 nm	Maximum mortality of 84.4% at 1.17% after 72 h, +22% compared with maximum mortality of 62.22% in crude extract.				Tribolium castaneum (Red flour beetle).
					Maximum mortality of 77.7% at 1.17% after 72 h, +20% compared with maximum mortality of 57.7% in crude extract.				Sitophilus oryzae (Rice weevil).
[59]	2024	Nanostructures of magnesium hydroxide as nanocarriers for Cry1Ac protein from Bt	Bt	N.S.	1.93 μg/mL	N.S.	N.S.	N.S.	Ectropis obliqua
[60]	2025	AgNPs Synthesis	Ocimum sanctum (Holy basil)	20 nm	LC50: 93.21 ppm at 24 h, 23.38 ppm at 48 h, 5.96 ppm at 72 h.				Spilosoma obliqua (hairy jute caterpillar)

* N.S. = Not specificed

, has shown that research has focused on three types of nanoparticles: metallic, inorganic, encapsulated and derived from different essential oils or plant extracts, whose efficacy and characteristics are described in the following sections.

4.2.1. Metal Nanoparticles

Metal nanoparticles, such as silver (AgNPs) and gold (AuNPs), were among the first to be explored for pest control. These NPs showed a high speed of action and strong larvicidal efficacy, although concerns arose related to their sustainability and potential toxicity. In more detail, Jasminum nervosum extract was used for the synthesis of AgNPs and AuNPs for the control of Culex quinquefasciatus [35] and Aedes aegypti larvae [44]. The AgNPs showed a Lethal Concentration 50 (LC50) and a Lethal Concentration 95 (LC95) (the concentrations that result in the death of 50 and 95% of the insects) of 57.40 μg/mL and 144.36 μg/mL, respectively, as well as a lethality time (LT50) of 2.24 h/150 μg/mL, showing superior efficacy compared to aqueous crude extracts (500 μg/mL to achieve the same effect, with a LT50 of 9.44 h). This effectively demonstrated how NPs can have the advantage of reducing both doses and the time required for pesticide action. Similarly, AuNPs showed an LC50 of 82.62 μg/mL and an LC95 of 254.68 μg/mL, with a LT50 of 4.51 h at 150 μg/mL. Again, the efficacy of AuNPs exceeds that of the crude extract, reducing both the concentration required and the time of action required to achieve lethal results. Likewise, AuNPs showed greater efficacy after 24 h against third- and fourth-stage Aedes aegypti larvae [44], with lower LC50 and LC90 than essential oil, indicating a need for lower concentrations to be effective. For example, for third-stage larvae, LC50 and LC90 for AuNPs were 62.47 ppm and 430.16 ppm, while for essential oil they were 111.15 ppm and 1441.51 ppm. Similar results were observed for fourth-stage larvae. However, AgNPs were also synthesized by Bacillus thuringiensis kurstaki (Btk-AgNP) [45] to control Trichoplusia ni and Agrotis ipsilon, proving to be more toxic than commercial AgNPs, with lower LC50s, highlighting the greater efficacy of biological synthesis (specifically, 0.46 mg/mL for Btk-AgNP vs. 0.81 mg/mL for commercial AgNPs for T. ni and 1.95 mg/mL for Btk-AgNP vs. 5.20 mg/mL commercial AgNPs for A. ipsilon).

Finally, AgNPs have also been synthesized from the leaf extract of Ocimum sanctum [60], showing significant efficacy in controlling the hairy jute caterpillar (Spilosoma obliqua), with higher efficacy than pure leaf extract. It is therefore reasonable to assume that biological synthesis could improve specificity and efficacy against parasites. However, the use of zinc-based nanoparticles (ZnO-NPs) has also been explored. For example, when coated with Bt [46] for the control of Callosobruchus maculatus, synthesized from Silybum marianum seeds for the control of Tribolium castaneum and Sitophilus oryzae [56] and from Azadirachta indica for the control of Myzus persicae [55], it achieved mortality rates of more than 75% within 72 h, with an efficacy of 20–22% more than crude extracts.

4.2.2. Inorganic Nanoparticles

Inorganic NPs, such as silicon dioxide (SiO₂) NPs, have also been explored and studied, and have shown some stability and ability to ensure the controlled release of the active ingredient, improving long-term efficacy and reducing environmental dispersion. For example, rough-surface silica nanoparticles (RS-SP) [50] have been developed for the release of spinosad against Rhipicephalus microplus. These nanoparticles achieved 100% mortality at low dosages (0.1 mg), compared to 63% of the commercial product and 35% of pure spinosad at the same dose, due to their ability to better adhere to pest surfaces and slowly release the active ingredient. However, the high cost of silica may limit its large-scale adoption. Modified SiO₂ nanoparticles [53] with Bt were then used to control Leptinotarsa decemlineata and Phyllotreta spp. The combination with Bt increased mortality by 10–15% compared to the use of Bt alone, demonstrating the synergistic potential between inorganic nanoparticles and biological agents.

4.2.3. Encapsulated Nanoparticles

NPs encapsulated in biological materials, such as chitosan [47], zein [49] and polylactic acid (PLA) [52], were then also considered in the literature. The use of these materials has actually improved both the efficacy and bioavailability of the active ingredient as well as providing better protection of the bioactive compounds from environmental degradation. For instance, the encapsulation of a nanobiopesticide based on neem extract within chitosan ensured 100% mortality of Spodoptera litura at a concentration of 0.3%, higher than the 81.67% of the crude extract. This encapsulation ensured controlled release and increased resistance to UV exposure, as it remains effective for up to 12 h under UV exposure, in contrast to the crude extract, which degrades rapidly. The use of zein nanoparticles to encapsulate neem oil [49] against Acanthoscelides obtectus, Bemisia tabaci, and Tetranychus urticae resulted in a reduction of LC50 and LC90 by 40–60% compared to unencapsulated oil. In fact, this result could further emphasize the efficacy of biological nanoparticles in reducing the required doses as well as an improvement in bioavailability and larvicidal efficacy by the nanobiopesticide. The encapsulation of abamectin in polylactic acid (PLA) then demonstrated significant results in terms of efficacy and safety [52], ensuring a gradual release of the active ingredient, achieving more than 90% mortality against Tuta absoluta, exceeding the performance of commercial formulations. A particularly relevant aspect was the reduction in toxicity towards natural predators: mortality for Nesidiocoris tenuis was less than 25–41%, while for Macrolophus pygmaeus it was significantly reduced compared to the 100% observed with conventional pesticides. These results, therefore, suggest that nanoformulations could be effectively integrated into integrated pest management programs, limiting the impact on beneficial organisms. Similarly, Bt toxins have been encapsulated in biodegradable nanoparticles [49] for the control of Ectropis obliqua (tea pests), achieving 95% mortality within 72 h at 2 mg/mL, a reduction in tea infestation of −70% in one week, with less frequent applications than conventional pesticides. Furthermore, the encapsulated toxins showed −70% reduced UV degradation and 90% efficacy at 40 °C for 48 h, compared to 50% for the free toxins. Finally, the nanobiopesticide was also shown to be safe for non-target organisms, such as bees and earthworms, with mortality of <5%, and the nanoparticles completely degraded within 30 days.

4.2.4. Plant Extracts and Essential Oils

Finally, the literature focused on the use of nanobiopesticides based on plant extracts and essential oils, which showed improved efficacy compared to the corresponding crude extracts. In particular, all the studies show that nanotechnology increases the bioavailability, stability, and efficacy of bioactive compounds, reducing the necessary doses and prolonging the duration of the pesticide effect. Furthermore, several studies point to additional beneficial effects, such as improved plant health and soil fertility [48], or the protection of compounds from environmental degradation [57]. For example, citronella-based nanoemulsions have been studied both against mosaic disease in Pogostemon cablin (patchouli) [48] and against Musca domestica and Lucilia cuprina [51]. In the former case, it was shown that the combined application of a 1% citronella-based nanobiopesticide (Nano-BC) and biofertilizers reduced the incidence (20–50%) and intensity (9.37–12.47%) of the disease, as well as improving plant height, dry biomass, and increasing the P₂O₅ content in the soil by up to 39 ppm, showing benefits for both crop health and soil fertility. In the second case, nanoemulsions proved particularly useful in environmental applications due to their larger active surface area, which improves adhesion and distribution, making them a promising solution for indirect insecticide treatments (nanoemulsions reached LT100 in 30 min compared to 45 min for untreated oil). However, nanoemulsions of anise, fennel, and mint have also been developed, with effective deterrence of Bactrocera oleae oviposition [54], with Median repellent concentrations (RC50) of 3.25% (aniseed) and 3.47% (fennel), significantly lower than mint, thus requiring lower concentrations to achieve the same repellent effect. These results suggest that anise and fennel oils are ideal candidates for eco-friendly ovideterrents against B. oleae. Iqbal et al. (2024) [57] showed that a nanobiopesticide based on Moringa oleifera was more effective than the crude extract, with +3.7% higher mortality against T. castaneum (83% ± 0.56 vs. 79.30% ± 2.64) and 11.3% higher mortality against R. dominica, and prolonged protection of the bioactive compounds. Finally, Jahan et al. (2024) [58] developed a nanobiopesticide based on Mentha piperita (259.8 nm), demonstrating superior efficacy compared to the crude extract both as an antimicrobial and as a pesticide. The nanosuspensions showed high antimicrobial activity against Clavibacter michiganensis, Pseudomonas syringae, and Fusarium oxysporum due to improved penetration into microbial tissues. At the insecticide level, the nanobiopesticide achieved 84.4% mortality on Tribolium castaneum and 77.77% mortality on Sitophilus oryzae at a concentration of 1.17% after 72 h, exceeding the efficacy of the crude extract by 22% and 20%. Even at low concentrations (0.60%), it maintained superior efficacy due to improved cuticular penetration, greater stability, and gradual release of the active ingredients, making it a more effective and sustainable solution.

Therefore, the literature shows that all types of nanoparticles significantly reduce the required doses and time of action compared to traditional pesticides and crude extracts. Encapsulation and the use of inorganic and biological nanoparticles could increase the stability of active ingredients, prolonging their efficacy even under adverse environmental conditions (UV, heat). Finally, encapsulated formulations in biodegradable materials and combined approaches with biofertilizers could show great potential for integration into integrated pest management programs, reducing the impact on beneficial organisms.

4.3. Nanoparticle Sizes

The size of nanoparticles could be a particularly significant parameter in the design and application of nanobiopesticides, as it influences their physical, chemical, and biological properties. The literature review revealed significant differences in the size of the NPs used, as shown in Table 1, with direct effects on the bioavailability, efficacy, and environmental sustainability of formulations. In particular, nanometric particle sizes (typically < 200 nm) provide a large specific surface area, which improves target interaction, controlled release, dispersion, and adhesion. Indeed, smaller particles can easily penetrate insect tissues or microbial membranes, increasing the efficacy of the active ingredient, adhering better to plant or pest surfaces, improving product distribution, and limiting losses by run-off. For example, AgNPs and AuNPs synthesized from Jasminum nervosum [35], Artemisia vulgaris [44] and Ocimum sanctum [60] have sizes of 2–22 nm. These extremely small sizes have an advantage because they allow rapid penetration into parasite tissues, ensuring immediate lethal action, but may increase the risk of environmental bioaccumulation and long-term toxicity. Studies such as those by Jahan et al. (2023) [56] and Khaleel et al. (2023) [55] produced ZnO-NPs with an average size of 14–51 nm. These particles showed high efficacy against pest insects, fungi, and phytopathogenic bacteria. Zhang et al. (2020) [50] and Shatalova et al. (2022) [53] developed silica nanoparticles with smooth or rough surfaces, with sizes of 10–350 nm. Smaller particles (10–20 nm) improved efficacy when combined with biological agents (Bt), while larger ones (330–350 nm) ensured a controlled release, reducing the amount of pesticide required and the environmental impact. Biodegradable matrices, such as those made of polylactide acid, used to encapsulate abamectin [52], are between 175 and 200 nm in size. These sizes offer a balance between sustained release and stability, minimizing toxicity to non-target organisms, but may be less effective against pests that require faster penetration. Joeniarti et al. (2019) [47] and Pascoli et al. (2020) [49] used chitosan and zein matrices to encapsulate plant extracts such as neem and citronella, producing particles of 271–350 nm. Although this size is larger than in other formulations, the use of biological materials ensures optimal degradability and reduces ecotoxicological risk. What also emerged is that the size of NPs directly influences the stability of formulations. For example, larger nanoparticles, such as PLA and silica nanoparticles (175–350 nm), offer greater protection to active ingredients, reducing degradation by up to 70% compared to free formulations [50,59]. However, some issues may arise. For instance, as also pointed out by Machado et al. (2023) [11], very small nanoparticles (<20 nm) have a high potential for bioaccumulation and may negatively affect ecosystems, requiring more in-depth assessments. In addition, the large-scale production of NPs with precise dimensions can be costly and technically complex. Therefore, nanoparticle size is a determining factor in the efficacy and sustainability of nanobiopesticides. Although smaller particles (<50 nm) offer greater bioavailability and faster penetration, medium-sized particles (150–350 nm) balance efficacy and stability, making them ideal for sustainable agricultural applications. Size optimization, combined with biodegradable materials, therefore represents a promising avenue for developing effective and environmentally friendly nanobiopesticides.

4.4. Impact on Targets and Non-Targets

Another aspect that emerged from the literature review is that a wide range of targets were studied, including insect pests, crop pests, bacteria, and phytopathogenic fungi, showing the versatility of nanobiopesticides. However, a central theme that emerges is the potential selectivity of these products and their impact on non-target organisms, such as pollinators, natural predators, and soil microorganisms, which is important in assessing the efficacy and sustainability of nanobiopesticides. From the point of view of targets, it is possible to distinguish between three macro-areas: (i) crop pests, (ii) crop-specific pests, and (iii) bacterial and fungal phytopathogens. Specifically, in the first case, ZnO-NPs [56] and nanobiopesticides derived from Moringa oleifera [57] showed high mortality against Tribolium castaneum and Sitophilus oryzae, demonstrating the efficacy of nanoparticles in improving the penetration and stability of active ingredients. Gold nanoparticles synthesized with plant extracts [44] showed significant larvicidal efficacy against Aedes aegypti larvae, while Khaleel et al. (2023) [55] used zinc oxide nanoparticles to reduce the incidence of Myzus persicae while improving plant growth. In the second case, abamectin nanoformulations [52] achieved mortalities of over 90% against Tuta absoluta, showing superior efficacy compared to commercial formulations. Zhang et al. (2020) [50] used rough silica nanoparticles to control Rhipicephalus microplus, showing reduced environmental dispersion due to improved adhesion to surfaces. Additionally, nanobiopesticides based on Bt toxins [59] reduced Ectropis obliqua infestations by −70%, improving the stability of the active ingredient and minimizing the required applications. Lastly, NPs based on Mentha piperita [58] and zinc [56] showed significant inhibition zones against Clavibacter michiganensis, Pseudomonas syringae, and Fusarium oxysporum, due to the greater bioavailability and stability of the active ingredients. However, one of the most debated aspects in the literature, as also highlighted in the reviews by Machado et al. (2023) [11] and Pan et al. (2023) [59], concerns the ability of nanobiopesticides to protect beneficial organisms and minimize collateral damage. This aspect emerged in some studies from this review, although it is important to note that it should be further investigated. For example, abamectin nanoformulations are significantly less toxic to natural predators such as Macrolophus pygmaeus and Nesidiocoris tenuis, with mortalities of less than 41% compared to 100% of commercial formulations, as shown by Cherif et al. (2022) [52]. Furthermore, Pan et al. (2024) [59] showed that Bt-based nanobiopesticides do not affect beneficial organisms such as bees and earthworms, with mortalities of less than 5%. However, silica nanoparticles [50] and zinc oxide nanoparticles [56] are also considered safe for soil microorganisms, although further investigations are needed to assess their long-term impact. In contrast, nanoparticles of very small size (<20 nm), such as AgNPs and AuNPs [35,44], while extremely effective, could present a potential risk of bioaccumulation and environmental toxicity, especially in aquatic ecosystems or soil. What has emerged, therefore, is that nanobiopesticides show high efficacy against a wide range of targets due to their ability to penetrate pest tissues and improve the stability of active ingredients. However, although many studies show a low impact on non-target organisms, such as pollinators and soil microorganisms, a gap identified in the literature is the need to more thoroughly assess the risk of bioaccumulation and biodegradability of nanoparticles, as well as to broaden the scope of studies against non-target organisms. Wide-ranging assessments on multiple biodiversity were not always seen in the studies found.

4.5. Environmental Sustainability Aspects

A further aspect that the literature review might suggest is that, despite some critical issues, nanobiopesticides could offer significant advantages over traditional pesticides, helping to mitigate some of the environmental problems associated with intensive agriculture. These benefits stem mainly from their advanced formulation, increased efficacy, and reduced waste, making them promising tools for more sustainable agriculture. In particular, thanks to their nanometric structure, nanobiopesticides theoretically require lower quantities of active ingredients to achieve the same effects as conventional pesticides, as demonstrated by the studies of Sundararajan and Kumari (2017) [44] and Pascoli et al. (2020) [49], which reduced the concentrations needed to control pests by up to 60%, sometimes minimizing the release of chemicals into the environment. However advanced formulations, such as those based on Bacillus thuringiensis [59] or rough-surface silica nanoparticles [50], offer a gradual release of the active ingredients, ensuring prolonged efficacy over time. This could reduce the number of applications required, thus decreasing the impact of repeated use of agricultural inputs and associated energy costs. Furthermore, this controlled release could reduce the risk of accidental contamination in non-target ecosystems. Furthermore, many nanostructured formulations, such as those based on chitosan [47] or zein [49], are designed to be biodegradable, which means that, once their function is exhausted, they naturally degrade without leaving persistent toxic residues in soil or watercourses, and this aspect could potentially make them more advantageous than conventional synthetic pesticides. Formulations such as PLA-based [52] and biodegradable Bt nanoparticles [59] have also demonstrated low toxicity to non-target organisms, including pollinators, earthworms, and beneficial soil microbes. This selective approach favors integrated agriculture that protects biodiversity and supports ecological balances in cultivated land. Finally, encapsulated nanoparticles [59] have superior resistance to degradation by exposure to UV light or high temperatures, which would allow for greater product stability, reducing the need for frequent reapplications and the risk of releasing inactive substances into the environment. However, to ensure that nanobiopesticides are indeed a sustainable solution, it is necessary to balance the environmental benefits with the possible critical issues. While they reduce the impact of many environmental categories, questions remain open as to their persistence and ecotoxicological safety, particularly for metal nanoparticle formulations.

Life Cycle Assessment Application

In any case, the sustainability of nanobiopesticides should be demonstrated using quantitative tools for assessing impacts, such as the Life Cycle Assessment. A further aspect that has emerged from the literature is the total absence of sustainability studies assessing the environmental impact of nanobiopesticides, both on an absolute level and in comparison. Therefore, within this study, an attempt was made to fill, at least partially, this gap by preliminarily assessing the sustainability of nanobiopesticides through the LCA methodology. In particular, an effort was made to compare the production of nanobiopesticides with that of a traditional pesticide, analyzing the consumption of resources, the energy required, and the associated emissions. Two nanobiopesticides were considered: a plant-based one, based on Mentha piperita [58] and without metal nanoparticles, and a Silver Nitrate (AgNO₃)-based one [60], to assess the differences between two different formulations (one completely natural and one with metal nanomaterials). For nanobiopesticides, the articles by Jahan et al. (2024) (plant-based) [58] and Ghosh et al. (2025) (AgNO₃-based) [60] were chosen because, among others, they provide a clear description of the composition and production parameters of the nanobiopesticides, including details on the quantities used, the synthesis process, and the energy required, a level of detail that was therefore satisfactory for setting up an accurate life cycle assessment. Specifically, the formulation of the Mentha piperita-based biopesticide was made using the plant’s essential oil, a non-ionic surfactant (Tween 80) and deionized water, following established protocols. The process involved the preparation of a crude emulsion, which was then sonicated to obtain a nanoemulsion with droplet sizes in the nanometre range. No metal nanoparticles were used in the formulation. For the production of the extract, 10 mg, 1 g leaves), 3 mg polyvinylpyrrolidone, 0.00083 kWh ultrasonication, and 0.3 mL deionized water were used [58]. The AgNO₃-based nanobiopesticide formulation, on the other hand, was obtained from a 1 M silver nitrate solution, prepared by dissolving a small amount of the compound in double-distilled water. In parallel, a plant extract was made from the dried leaves of Ocimum sanctum, ground and infused with methanol for 48 h, then filtered and concentrated by controlled heating. The extract obtained was then slowly added to the AgNO₃ solution under continuous stirring while the system was heated between 50 and 60 °C for about 10 to 15 min, promoting the reduction of silver ions to AgNP. Subsequently, the mixture was left to incubate at 25 °C for 24 h under dark conditions. Finally, to obtain an applicable formulation, the plant residue was dissolved in acetone, creating a 1% stock solution from which various concentrations of between 50 and 650 ppm were prepared [60]. For the traditional pesticide, an average generic pesticide was considered within the Ecoinvent v3.11 database (process name: ‘Pesticide, unspecified [GLO], market for, APOS’). This process is modeled as a weighted average of various commonly used pesticides, considering the inputs and outputs associated with their production. The objective is to provide a representative dataset that reflects the average environmental characteristics of pesticides without referring to a specific substance. This approach is commonly used when detailed data on a specific pesticide are not available or a general assessment of the environmental impact of pesticides is desired.

However, it is important to emphasize that the selection of the two nanobiopesticides represents a relatively simple and illustrative result, leading to preliminary and generic results, as well as not being representative of the entire category. Indeed, the production process described in the study by Jahan et al. (2024) [58] and Ghosh et al. (2025) [60] uses readily available resources and standardized technologies, such as ultrasonication, which, while demonstrating high energy efficiency and replicability, is rather basic compared to more complex formulations that could integrate innovative materials or multifunctional approaches. While this methodological simplicity favors replicability and facilitates life cycle analysis, it also limits the comparison to a preliminary scenario that is unrepresentative of the complexity that characterizes many other nanobiopesticides. Therefore, the results obtained from the sustainability analysis on this product should be interpreted as a simplified example, useful to build a methodological starting point, but not necessarily generalizable to all types of nanobiopesticides available on the market or under development. The LCA was therefore conducted following the three mandatory steps required by ISO 14040 [61] and 14044 [62]: (i) goal and scope definition; (ii) life cycle inventory (LCI); and (iii) life cycle impact assessment (LCIA). Concerning of goal and scope definition, this evaluation aimed to assess the environmental sustainability of two nanobiopesticides, one with and one without metal nanoparticles, by comparing them with a conventional pesticide. The system’s boundaries included a ‘from cradle to gate’ assessment, thus considering the production and application of the nanobiopesticides and the traditional pesticide. The treatment of 1 m² of soil was chosen as the functional unit to represent the agronomic efficacy. The literature review showed that nanobiopesticides generally require lower doses to achieve the same effect as conventional pesticides, due to better bioavailability and controlled release of the active ingredients. Therefore, this saving should be reflected in the analysis, which is why, in the case of nanobiopesticides, the application of 10 mg/m² was theorized against 30 mg/m² for the traditional pesticide. The LCI data were taken from the literature, particularly from Jahan et al. (2024) [58] and Ghosh et al. (2025) [60], modeled according to the Ecoinvent v3.11 database [63] and available in the Appendix A (

Table A1. Life Cycle Inventory of Mentha piperita-based nanobiopesticide (×10 mg/m²).

Input	Quantity
Mentha piperita extract	1 g of leaves
Polyvinylpyrrolidone	3 mg
Distilled water	0.3 mL
Ultrasonication energy	0.00083 kWh
Output
Mentha piperita-based nanobiopesticide	10 mg

and

Table A2. Life Cycle Inventory of AgNO₃-based nanobiopesticide (×10 mg/m²).

Input	Quantity
AgNO3	0.001690 g
Double-distilled water	10.00 mL
Ocimum sanctum extract	2.00 g
Methanol	5.00 mL
Output
AgNO3-based nanobiopesticide	10 mg

). Concerning the LCIA, on the other hand, data were processed using SimaPro 9.6 software, and the ReCiPe 2016 Midpoint method was chosen [64], which considers 18 impact categories and follows an individualistic perspective (20 years), focusing on short-term impacts, which are shown in

Table A3. Results of the LCA comparison between a nanobiopesticide and a commercial pesticide. (GWP = Global Warming Potential; SOD = Stratospheric Ozone Depletion; IR = Ionizing radiation; OFHH = Ozone Formation, Human Health; FPMP = Fine Particulate Matter Formation; OFTE = Ozone formation, Terrestrial ecosystems; TAP = Terrestrial acidification Potential (TAP); FEP = Freshwater Eutrophication Potential; MEP = Marine Eutrophication Potential; TEC = Terrestrial Ecotoxicity; FEC = Freshwater Ecotoxicity; MEC = Marine Ecotoxicity; HCT = Human Carcinogenic Toxicity; HNCT = Human Non-Carcinogenic Toxicity; LU = Land Use; MRS = Mineral Resources Scarcity; FRS = Fossil Resources Scarcity; WC = Water Consumption).

Impact Categories	Unit	AgNO3-Based Nanobiopesticide	Plant-Based Nanobiopesticide	Commercial Pesticide
GWP	kg CO2 eq	9.98 × 10−4	6.56 × 10−4	1.23 × 10−2
SOD	kg CFC11 eq	5.30 × 10−10	2.02 × 10−9	3.61 × 10−8
IR	kBq Co-60 eq	7.84 × 10−5	5.99 × 10−5	6.57 × 10−4
OFHH	kg NOx eq	6.68 × 10−6	2.12 × 10−6	2.74 × 10−5
FPMP	kg PM2.5 eq	1.07 × 10−6	2.62 × 10−7	6.89 × 10−6
OFTE	kg NOx eq	6.68 × 10−6	2.12 × 10−6	2.77 × 10−5
TAP	kg SO2 eq	6.32 × 10−6	6.78 × 10−6	7.58 × 10−5
FEP	kg P eq	2.36 × 10−6	1.96 × 10−7	6.06 × 10−6
MEP	kg N eq	5.64 × 10−8	9.30 × 10−7	3.19 × 10−6
TEC	kg 1,4-DCB	8.49 × 10−5	4.79 × 10−5	1.15 × 10−2
FEC		3.72 × 10−6	4.98 × 10−6	1.18 × 10−4
MEC		1.33 × 10−6	9.36 × 10−7	9.69 × 10−6
HCT		8.78 × 10−8	5.52 × 10−8	1.83 × 10−5
HNCT		1.57 × 10−5	1.24 × 10−6	2.13 × 10−4
LU	m2a crop eq	1.04 × 10−4	6.22 × 10−4	6.84 × 10−4
MRS	kg Cu eq	1.75 × 10−4	2.55 × 10−6	2.66 × 10−4
FRS	kg oil eq	2.40 × 10−4	1.54 × 10−4	3.65 × 10−3
WC	m3	9.49 × 10−6	1.14 × 10−4	3.99 × 10−5

and Figure 5. This initial analysis shows how both nanobiopesticides, for the same treated soil, reduce environmental impacts compared to the commercial pesticide.

For example, for the plant-based nanobiopesticide, the reductions cover 17 out of 18 impact categories and range from −94% for GWP (6.03 × 10⁻⁴ kg CO₂ eq/m² for the nanobiopesticide vs. 1.06 × 10⁻² kg CO₂ eq/m² for the commercial pesticide) and for FPMP (1.46 × 10⁻⁶ kg PM₂.₅ eq/m² for nanobiopesticide vs. 2.86 × 10⁻⁵ kg PM₂.₅ eq/m² for commercial pesticide), to −90% for SOD (3.10 × 10⁻⁹ kg CFC11 eq/m² for nanobiopesticide vs. 3.31 × 10⁻⁸ kg CFC11 eq/m²) and ionizing radiation (6.13 × 10⁻⁵ kBq Co-60 eq/m² for nanobiopesticide vs. 6.76 × 10⁻⁴ for commercial pesticide). Regarding AgNO₃-based nanobiopesticide, on the other hand, reductions affect all the impact categories considered, with values ranging from −34% for MRS to −100% for HCT. For example, values ranged from −92% for GWP (9.98 × 10⁻⁴ for AgNO₃-based nanobiopesticide vs. 1.23 × 10⁻² kg CO₂ eq for commercial pesticide), −99% for SOD (5.30 × 10⁻¹⁰ for AgNO₃-based nanobiopesticide vs. 3.61 × 10⁻⁸ kg CFC11 eq for commercial pesticide), to −92% for TAP (6.32 × 10⁻⁶ for AgNO₃-based nanobiopesticide vs. 7. 58 × 10⁻⁵ kg SO₂ eq for commercial pesticide), −86% for MEC (1.33 × 10⁻⁶ for AgNO₃-based nanobiopesticide vs. 9.69 × 10⁻⁶ for commercial pesticide), and −100% for HCT (8.78 × 10⁻⁸ for AgNO₃-based nanobiopesticide vs. 1.83 × 10⁻⁵ for commercial pesticide). Thus, from the LCA data, a hierarchical scale emerges among the three pesticides, where the commercial pesticide appears to be the most impactful, while the plant-based nanobiopesticide appears to be the most sustainable, and the AgNO₃-based nanobiopesticide is in an intermediate position. The traditional pesticide shows the highest values in all the impact categories, while the AgNO₃-based nanobiopesticide is a more sustainable alternative, but with a few more critical issues than the plant-based version, especially in terms of MRS and toxicity, all due to the presence of metal nanoparticles, which result in a more energy-intensive synthesis step and potential bioaccumulation problems. Mentha piperita-based nanobiopesticide, on the other hand, appears to be the most sustainable in almost all categories, compared with both AgNO₃-based nanobiopesticide and commercial pesticide, with a lower mineral resource scarcity, being free of metal nanoparticles. However, its land use (6.22 × 10⁻⁴ m²a crop eq) and water consumption (1.14 × 10⁻⁴ m³) are higher than the AgNO₃-based nanobiopesticide, probably due to higher electricity use, although there could be many reasons for this, too. In general, for both types, however, it appears that nanobiopesticides can reduce impacts by −6% to −99% compared to commercial pesticides. What is important to consider, however, is the impact on toxicity, for which pesticides are particularly responsible, as amply demonstrated by this literature review. In particular, the use of a plant-based nanobiopesticide would seem to reduce the impact categories related to toxicity, with values ranging from −94% to −99%. More specifically, the values are −99.23% for terrestrial ecotoxicity, −95.49% for freshwater ecotoxicity, −94.31% for marine ecotoxicity, −99.60% for Human Carcinogenic Toxicity, and −99.10% for Human Non-Carcinogenic Toxicity. Overall, the AgNO₃-based nanobiopesticide could be a more sustainable alternative to traditional pesticides, but less than a plant-based version. Therefore, what emerges is that nanobiopesticides, based on a literature application, appear to be more sustainable than conventional pesticides throughout their life cycle. However, despite these results, it is necessary to consider certain limitations and aspects that require further investigation. First, these results are based on a preliminary analysis considering ideal scenarios. The variability of operating conditions (climate, soil, application) could significantly influence the environmental impacts. Next, the production of nanobiopesticides could involve energy consumption and the use of unsustainable materials on a large scale, influencing the results in terms of overall sustainability. Furthermore, despite reductions in toxicity categories, not all aspects of bioaccumulation and environmental persistence of nanoparticles have been assessed. Indeed, some nanomaterials could accumulate in soil or water with as yet unknown long-term consequences. As shown by Rajput et al. (2020) [65], soils may act as sinks for NPs, where they can accumulate through atmospheric precipitation, sedimentation of dust and aerosols, and various anthropogenic activities. Additional sources of release include wastewater treatment plants and landfills, which contribute to the spread of NPs into the agricultural environment. Once in the soil, nanoparticles do not remain static but undergo chemical and physical transformations that change their bioavailability and toxicity. Parameters such as soil pH, the presence of organic matter, and water content play a crucial role in these processes, influencing, for example, the dissolution of metal NPs. This phenomenon can increase the concentration of free ions in the soil, which are more easily absorbed by plants but also potentially more toxic. Nanoparticles, in fact, can be absorbed by plant roots, translocated to aerial tissues, and accumulate in cell organelles, resulting in alterations in the plant’s own physiological processes. This process not only affects crop health but also carries the risk of NPs entering the food chain, with potential implications for human health. As shown further by Wu et al. (2021) [66], AgNPs in particular exhibit dynamic behavior in soil, as their dissolution affects silver availability to plants and can significantly alter accumulation in plant tissues. This dynamic not only favors the uptake of AgNPs by plants but also has significant effects on bacterial communities in the rhizosphere, changing their structure and biodiversity. Therefore, these results emphasize the complexity of the interactions between NPs, plants, and soil microbes, but also the need for further studies to assess the long-term effects on the agricultural ecosystem. In any case, these preliminary results, given the absence of LCA studies assessing the environmental sustainability of nanobiopesticides, could represent a starting point for assessing their potential as a sustainable alternative to traditional pesticides.

4.6. Safety and Regulatory Aspects

Although nanobiopesticides represent a particularly promising approach for sustainable agriculture, as shown by the literature review, their regulation still requires an approach that takes into account the peculiarities of nanomaterials and the potential risks associated with their production, use, and disposal. Globally, regulation of nanomaterials in agriculture is under development and varies significantly from country to country, with the European Union being an example of advanced but rather complex legislation. Specifically, in the EU, nanomaterials have been regulated by Regulation (EC) 1907/2006 [67], known as the Registration, Evaluation, Authorization and Restriction of Chemicals (REACH). Although this regulation does not explicitly mention nanomaterials, they fall under the general definition of “substance”. In addition, the European Commission’s Recommendation 2011/696/EU [29] provided a harmonized definition of a nanomaterial, literally defined as “a natural, incidental or manufactured material containing particles in a free state or as aggregates or agglomerates, where 50% or more of the particles in the number size distribution have one or more external dimensions between 1 nm and 100 nm”. After much work by the European Chemicals Agency (ECHA), which has developed numerous guidance documents since 2017 to support registrants of nanomaterials, NPs are subject to specific requirements as of 1 January 2020, thanks to Regulation (EU) 2018/1881 [68], which complements REACH. These requirements include: the characterization of nanoforms (size, shape, and physicochemical properties must be clearly defined), chemical risk assessment (bioaccumulation, toxicity, and environmental degradation), and registration requirements (with details on production methods, use, volume, and exposure potential). Before the specific updates introduced by Regulation (EU) 2018/1881, nanoforms were covered by REACH under the generic definition of “substances,” but detailed regulations for their registration and management were lacking. Although Recommendation 2011/696/EU had already provided a harmonized definition of nanomaterials, this did not automatically translate into specific requirements for the assessment and management of nanomaterials under REACH. Regulation (EU) 2018/1881 [68] thus ensured that nanoforms were treated differentially from conventional chemicals, given their unique nature and specific properties. This update introduced clear requirements for the characterization, registration, and evaluation of nanoforms, improving transparency and the ability to monitor and manage risks associated with nanomaterials. Regulation (EU) 2018/1881 aligns with the Biocidal Products Regulation (BPR), or Regulation (EU) 528/2012 [69], which requires a separate assessment of nanoforms in biocidal products, requiring dedicated dossiers and risk assessment. Recommendation 2022/692/EU [70], then, extended and further specified what was established in the previous definition of 2011, stating that to be defined as a nanomaterial, it is necessary that 50% or more of the particles in the number distribution fall into at least one of the following conditions: (1) outer dimensions between 1 nm and 100 nm; (2) elongated shape, such as fibers, tubes, or rods, where two outer dimensions are less than 1 nm and the other is greater than 100 nm; and (3) flat shape, such as plates, where one outer dimension is less than 1 nm and the other two are greater than 100 nm. In addition, materials with a specific surface area per volume of less than 6 m²/cm³ are not considered nanomaterials, even if they meet the other conditions. In fact, within this Recommendation, the concept of the nanomaterial is expanded to include particles with specific shapes (elongated or flat), and some specific exclusions are established. In agriculture, specifically, nanomaterials are also regulated by the Plant Protection Products (Regulation 1107/2009/EC) [71], which requires the authorization of active ingredients at the European level and formulated products at the national level. Nanoforms of active substances must be evaluated separately, with specific labeling requirements (e.g., the word “nano” next to the name of the substance).

In addition, biocidal products containing nanomaterials are governed by Regulation (EU) 528/2012 [69], which requires detailed risk assessments and prohibits simplified authorization processes for products containing nanomaterials. Globally, however, no country has yet developed specific legislation to cover the use of nanomaterials in the agricultural sector. In the United States, the Environmental Protection Agency (EPA) regulates pesticides, including those containing nanomaterials, through the Federal Insecticide, Fungicide, and Rodenticide Act (FIFRA) [72] and the Toxic Substance Control Act (TSCA) [73]. EPA requires detailed information on nanomaterials, including production methodologies, exposure, and safety data, and can restrict the use of nanomaterials to protect public and environmental health. In Canada, the Canadian Environmental Protection Act (CEPA) provides for the registration of nanoforms not listed on the Domestic Substances List [74,75]. Canada and the United States collaborate under the Regulatory Cooperation Council to harmonize regulations on nanomaterials [76]. In Australia, regulation is handled by the Australian Pesticides and Veterinary Medicines Authority (APVMA), which enforces the National Registration Scheme for Agricultural and Veterinary Chemicals [77]. However, there are no specific provisions in the regulations for nanomaterials, which are regulated on a case-by-case basis. The regulatory framework shows some complexity, especially in the EU, since the regulation of NMs, while advanced, is still quite fragmented and complex, with multiple regulations interacting with each other. In addition, the need for a separate evaluation for nanoforms (as required by Regulation 2018/1881 and the Plant Protection Products Regulation) could further increase the time and cost of registration, slowing down the market introduction of new products. Furthermore, although Recommendation 2022/692 [70] expanded the definition of nanomaterials by including specific forms and introducing exclusion criteria, the complexity of the classification could generate ambiguity in its practical application. The exclusion of materials with a low specific surface area could exclude some potentially toxicologically relevant nanoparticles, leaving open questions as to their regulation. Finally, at the global level, the lack of specific legislation for nanomaterials in agriculture leaves a significant regulatory gap, which is why regulation at the global level is desirable and may be effective in managing the specific risks of nanomaterials. In this context, the requirement to clearly label nanomaterials in biocides and plant protection products (e.g., by indicating “nano”) could be a step forward in providing greater transparency to consumers and farmers, although there is still a need to educate end users about the benefits and risks associated with nanomaterials. Finally, it will be critical to expand monitoring studies of the long-term effects of nanomaterials on ecosystems and human health, the absence of which was also found in the literature review.

4.7. Challenges and Opportunities

Although nanobiopesticides are generally presented as more sustainable solutions than traditional synthetic pesticides, there are nonetheless some critical issues that have emerged from the literature review. First, their application is still in its infancy, as evidenced by the few studies present and further confirmed by Machado et al. (2023) [11], in addition to the fact that they are not yet currently accessible in sufficient quantities for widespread commercial use. Additionally, for example, the synthesis of some nanoparticles, especially those based on inorganic materials such as silica [50], requires energy-intensive processes or the use of chemical reagents, which may not be sustainable on a large scale. This contrasts with biological materials such as chitosan and zein [47,49], which instead offer lower environmental impacts but are limited by the availability of natural resources. Moreover, the use of plant extracts such as neem, moringa, and peppermint [47,57,58] could increase agricultural pressure to grow plants for extraction, taking land away from food crops and compromising food security in particularly vulnerable areas.

Nanoparticles of extremely small size (<20 nm) [35,44], then, are very effective but could pose a risk of environmental bioaccumulation. Non-biodegradable particles, such as silica particles [50], could also accumulate in soils and aquatic ecosystems, with the long-term effects not yet known. Although some nanoparticles, such as biodegradable Bt-based nanoparticles [59], degrade rapidly (within 30 days), other formulations could persist longer, adversely affecting natural cycles. For example, ZnO-NPs [55,56] require further studies to understand their degradation times and potential accumulation. Many studies report reduced toxicity to pollinators and natural predators, as demonstrated by Cherif et al. (2022) [52] and Pan et al. (2024) [49]. However, the impact on soil microbes and symbiotic fungi is often overlooked, as are the long-term environmental impacts and consequences. Indeed, metal formulations, such as zinc oxide formulations, could alter the soil microbiome, adversely affecting nutrient cycling. Finally, there is a question of affordability and accessibility. In particular, some advanced technologies, such as rough-surface nanoparticles [50], despite offering greater efficacy, may be expensive to produce, limiting accessibility for farmers in developing countries, where agriculture accounts for 15–30% of GDP [78]. Cheaper solutions, such as those based on biofertilizers [48], could mitigate these limitations but require more complex logistics, especially to maintain uniform quality standards. Finally, an additional problem could be farmers’ unfamiliarity with nanotechnology and lack of specific training, especially in traditional agricultural settings. Although nanobiopesticides represent a promising solution for improving sustainability in agriculture, their large-scale use requires the careful evaluation of environmental and social implications. In particular, the need emerges to:

• Develop synthesis methods that are energy-efficient and less dependent on limited resources.
• Improve the biodegradability of nanoparticles and assess bioaccumulation risks.
• Implement long-term impact studies on ecosystems and non-target organisms.
• Ensure affordable costs to promote widespread adoption, especially in the most vulnerable settings.

By addressing these critical issues, it will be possible to effectively integrate nanobiopesticides into a truly sustainable agricultural model.

5. Conclusions

The literature review highlighted the potential of nanobiopesticides to address some significant challenges related to agriculture, such as the need to reduce the use of conventional chemical pesticides, improve the efficiency of plant protection treatments, and reduce the overall environmental impact. The results of the study showed that the literature is fairly unanimous in highlighting that nanobiopesticides offer advantages over conventional pesticides, including greater precision of action, controlled release of the active ingredient, and reduced doses needed to achieve comparable results. In addition, an illustrative LCA approach conducted in this study further confirmed that nanobiopesticides can be more sustainable than commercial pesticides, as for the same area treated, they reduce environmental impacts, with reductions as high as −99%. The characteristics mentioned, then, as shown by the 17 studies analyzed and as confirmed by the LCA, make them particularly promising. However, despite these positive prospects, several critical issues remain that require further investigation. First, the overall sustainability of nanobiopesticides needs to be further demonstrated through quantitative tools such as the Life Cycle Assessment, thus increasing the number of studies. The literature has shown a dearth of research in this area, limiting the ability to draw definitive conclusions about their long-term environmental impact. In addition, the effect of nanoparticles on non-target organisms and biodiversity remains an area of great uncertainty.

Although some studies suggest lower toxicity to non-target species than conventional pesticides, it is still unclear how bioaccumulation and environmental persistence may affect ecosystems. Therefore, it would be necessary to develop standardized and expanded evaluation protocols to consider not only the efficacy of nanobiopesticides but also their ecotoxicological safety. Another particularly significant issue is regulation. Although the European Union has developed an advanced regulatory framework with the REACH Regulation and specific additions, the complexity and length of authorization processes could hinder the large-scale adoption of nanobiopesticides. Globally, the lack of specific regulations in major agricultural markets, such as the United States and Canada, also highlights a regulatory gap that needs to be filled. Finally, it is important to consider the economic issue. The production of nanobiopesticides often requires specific resources and advanced technological processes, which may make them inaccessible to small-scale farmers, especially in developing countries. Therefore, it will be necessary to invest in strategies to reduce production costs and ensure an equitable diffusion of technologies. In conclusion, the review showed that nanobiopesticides represent a promising application area in crop protection and sustainable agriculture. However, to effectively integrate these solutions into global agricultural systems, it will be essential to address challenges related to environmental sustainability, ecotoxicological safety, regulation, and affordability. Only in this way will it be possible to harness the potential of nanobiopesticides, minimizing their risks and maximizing their benefits.