Techno-Economic Analysis of ZnO Nanoparticles Pretreatments for Biogas Production from Barley Straw

Abstract

The aim of this study was to analyze the effect of ZnO nanoparticles (ZnO NPs) on the biogas production from mechanically treated barley straw and to perform a techno-economic analysis based on the costs assessment and on the results of biogas production. The structural changes of mechanically pretreated barley straw were observed using FTIR, XRD, TGA, and SEM. Additionally, both green ZnO NPs prepared from red alga (Antithamnion plumula) extract and chemically prepared ZnO NPs were characterized by FTIR, XRD, SEM, and TEM, surface area, and EDX. The results revealed that the biogas production was slightly improved by 14.9 and 13.2% when the barley straw of 0.4 mm was mechanically pretreated with 10 mg/L of both green and chemical ZnO NPs and produced 390.5 mL biogas/g VS and 385 mL biogas/g VS, respectively. On the other hand, the higher concentrations of ZnO NPs equal to 20 mg/L had an inhibitory effect on biogas production and decreased the biogas yield to 173 mL biogas/g VS, which was less than the half of previous values. It was also clear that the mechanically treated barley straw of 0.4 mm size presented a higher biogas yield of about 340 mL/g VS, in comparison to 279 mL biogas/g VS of untreated biomass. The kinetic study showed that the first order, modified Gompertz and logistic function models had the best fit with the experimental data. The results showed that the nanoparticles (NPs) of the mechanically treated barely straw are a suitable source of biomass for biogas production, and its yields are higher than the untreated barley straw. The results of the cost-benefit analysis showed that the average levelized cost of energy (LCOE), adopting the best treatments (0.4 mm + 10 mg/L ZnO), is 0.21 €/kWh, which is not competitive with the other renewable energy systems in the Egyptian energy market.

1. Introduction

Biomass is a well-known renewable source to offer energy demand in terms of heat and electricity [1]. The Renewable Energy Directive 2018/2001/EU (RED II) forces the European Union (EU) to increase renewable energy consumption to 32% by 2030 [2]. Anaerobic digestion (AD) is extremely motivating in this framework as it can function in the production of renewable energy in the form of methane (CH₄)-enriched biogas; AD is a reasonably slow microbial centered method that is reliant on pH, temperature, C:N ratio, and hydraulic retention time [3]. Various kinds of organic materials can be used as substrates for biogas production, such as manure, sewage sludge, and agriculture residues. Among these agricultural residues, straw is a motivating feedstock for the production of biogas [3,4]. For example, barley straw is one of the most worldwide plentiful crop residues. Barley straw is an agricultural residual of barley production with high total solids (TS) content [5,6]. However, the composition and character of lignocellulose, with cellulose fibers intertwined with hemicelluloses and lignin, make it hard to degrade [6]. The biogas revenue from barley straw can be improved by harvesting techniques [7], pretreatment, making the material more reachable to microbial degradation [3,6,8,9]. Feng et al. [5] examined the impact of co-ensiling of cover crops and barley straw on biogas production. The results demonstrate that cover crops are practicable for producing biogas with considerable CH₄ yield (330 mL CH₄/g VS) and has good storability. Mechanical pretreatment typically provides a big surface area in the organic materials, permitting a greater area of contact for the microorganisms that degrade the materials, leading to increased biogas yields, the overall conversion of lignocellulosic biomass into biogas without producing any toxic side streams [10]. The biogas yield of some substrates increases nearly linearly with the increase of the outer surface [11]. Compared with alternative approaches (i.e., sonication, chemical, biological), mechanical pretreatments are typically considered more suitable for industrial applications [12,13,14,15]. Balsari et al. [16] studied the effect of mechanically pretreated barley straw (sizes at 5.0, 2.0, and 0.5 cm) on the biogas and methane yield. The pretreated barley straw size 0.5 cm gives the highest yields 784 and 370 (NL kg⁻¹ VS) for biogas and methane production, respectively. Nanoparticles (NPs) are currently extensively used in commercialized products. With the exceptional physiochemical characteristics (e.g., optical, magnetic, and electrical features), the usage of NPs is considered perfect in manufacturing trades. Adding additives has become a widely used approach to improve AD performance [17]. Numerous researches have examined the addition of different kinds of NPs to improve the biogas production and enhance AD [16,17,18,19]. Mu et al. [20] stated that only chemical ZnO NPs have a reducing impact on methane production. Additionally, the lowering of ZnO NPs dosage (less than 6 mg/g TS) has little or no impact on methane production. Hassaan et al. [21] verified that NPs might enhance the AD process and stimulate the slurry digestion, which improves the biogas generation, but it depends on a certain dosage. The improvement of the biogas production from durum wheat is attained by adding lower concentrations of 5 and 10 μg/mL of ZnO NPs, while the addition of a higher concentration of 20 μg/mL of ZnO NPs has an inhibitory effect.

In this work, the novel designed synthesis procedure based on macroalgae extract for the green synthesis of ZnO NPs with small particle sizes is environmentally friendly, economically affordable, and predominant, preventing using unsafe materials, which are common in conventional chemical methods [22,23]. Four kinetic models—the first-order kinetic, the modified Gompertz, the logistic function, and Cone models—were reported to represent and reproduce the experimental data. However, to the best of our knowledge, the study on the kinetics of biogas production under the influence of NPs stress does not exist. The accuracy of these models was compared by statistical analysis of correlation coefficients (R²).

As in other biomass usage devoted to energy production [24,25], it is important from an economical point of view to perform a cost-benefit analysis of using mechanical and ZnO NPs treatment for barley straw to produce biogas as the profitability may ultimately determine if farmers and landowners start to utilize this practice. The economic feasibility of biogas production depends on the possible income from the biogas produced versus the total cost of production. These parameters are affected by local and site-specific conditions; for the case of this study, the assumptions made are presented in the results. The cost-benefit analysis performed in this study was based on the whole chain of crop cultivation (land preparation, planting, pesticide, and fertilizer application), harvesting, transport, mechanical (chipping and milling), ZnO NPs treatment, investment, and conversion of the crops at a biogas plant. The costs for these variables are based on prices in Egypt. The levelized cost of energy (LCOE) is an estimation of the economic lifetime energy production and cost [26]. The LCOE is sensitive to minor alterations in the input variables and expectations. Therefore, the precise values of these input statistics are crucial for consistent results. Furthermore, the sensitivity of the LCOE to different input data must also be estimated. This work aimed to study the impact of green ZnO NPs synthesized from the extract of red algae Antithamnion plumula (A. plumula) on the mechanically pretreated barley straw with particles size 0.4 mm and untreated (raw) barley straw 4 mm. Based on the literature survey, A. plumula alga extract was employed for the first time as a natural nano-factory for ZnO NPs biosynthesis. The content of innovation of this research was to perform an assessment of cost-benefit analysis and LCOE for using nanotechnology in biogas production. According to the knowledge of the author, this was the first work that describes in detail the effect of a combination of mechanical and ZnO NPs treatment based on various cost calculation approaches, such as cost-benefit analysis and LCOE. Moreover, this paper delivered new insights for NPs technologies and its availability of usage on a large scale.

2. Materials and Methods

Figure 1 shows a flow chart of the experimental examinations with the steps of the procedures proposed. Zinc acetate dihydrate and NaOH were obtained from Aldrich Chemicals, Milwaukee, WI, USA.

2.1. Inoculum and Substrates Preparation

Barely straw was used as a substrate for this work. An inoculum was obtained from a mesophilic biogas plant in Puglia, Italy. Red algae (A. plumula) for ZnO NPs synthesis was collected from the coast of Baltic Sea, Kiel City, Germany (54°453789″ N, 10°197037″ E) in the spring of 2018. The barley cultivation (Hordeum vulgare L.) was done at the experimental farm in Puglia, Italy, and was harvested in the spring of 2018. Standard methods [27] were applied for the determination of total solids (TS), volatile solids (VS), and ash content. The C, H, and N measurement was done by the elemental analyzer (LECO Model CHN 628).

2.2. Biogas Experiment

All experiments were implemented with 1 g TS of barley straw (dry weight) and 20 g of inoculum (wet weight) with a concentration of 8.40% TS, followed by mixing for 20 min for homogenization. The inoculum/substrate ratio was set based on the previous works [18,21]. The mechanically pretreated barley straw was dried, milled, and separated to attain a particle size of 0.4 mm using Fritsch, Pulverisette 2, and Filtra vibracion S.L. Other raw samples of barley straw were only dried and cut into small pieces of about 4 mm to fit the digesters. Laboratory tests were implemented in equivalent cylindrical syringes digesters [28,29,30,31]. The syringes are reversed straight forward into the lid of the reactor [21,31]. The gas was sampled with a plastic syringe, which was fitted with a 3-way valve and reinjected into the waste. The liquid part of each digester was heated by incubation and monitored with a thermometer situated at the mid-depth and adjusted to 37 ± 1 °C.

Chemical and green ZnO NPs were synthesized from A. plumula extract used to study their effects on biogas production from raw and mechanically pretreated barley straw. A stock solution of ZnO NPs at a concentration of 1 g/L was prepared by dispersing nanopowder into MiliQ water (conductivity of 18.2 MU/cm at 25 °C). The stock was then sonicated for 30 min to break aggregates and disperse NPs into the solution. The NPs dispersion was kept in the dark condition in order to prevent the photocatalytic reaction. The stock dispersion of ZnO NPs was stored at 4 °C and used within 24 h of preparation. Then, the prepared ZnO NPs solution was diluted to 5, 10, and 20 mg/L for shock loading in the present study. All the experiments were done in duplicates. The significant difference among the experiments was calculated using the T-test in Microsoft Excel.

2.3. Preparation of Extract from A. plumula

Ten grams of A. plumula were washed with sterile double distilled deionized water (SDDW), dried at air temperature (28 °C), and powdered with mortar; then, the powder was added to the Erlenmeyer containing 1000 mL of SDDW. The final mixture was refluxed (70–80 °C) for 3 h, and the mixture was cooled at room temperature (Figure 2). The mixture was centrifuged at 5000 rpm for 5 min, and the small portion of the extract was subjected to GC-MS analysis of the ethanol extract [32].

2.4. Synthesis of ZnO Nanoparticles (ZnO NPs)

The 10 mL of A. plumula extract was mixed with 90 mL of 0.1 M of zinc acetate dihydrate solution drop-wise under continuous agitation at ambient temperature for 4–5 h. After that, 50 mL of 2.0 M NaOH solution was added drop-wise into the above mixture, and the stirring was allowed for 2 h. The resulting white precipitate was filtered and washed repeatedly with distilled water and ethanol to remove the impurities. Finally, the white powder obtained was dried at 60 °C in an oven for 24 h [21,32]. Then, the dried powder was calcinated at 550 °C for 2 h to obtain a pure pale white powder of ZnO NPs (Figure 2), which was carefully collected and used for further investigation. For the chemical synthesis, the same procedures were followed without adding an aqueous extract of the plant [18,21,33].

2.5. Characterization and Measurement

The samples of both ZnO NPs and barley straw (Hordeum vulgare L.) before and after mechanical pretreatment were characterized by the following techniques: Fourier transform infrared (FTIR) spectroscopy (platinum ATR) model V-100 VERTEX70, Germany, in the wavenumber range (400–4000 cm⁻¹), X-ray diffractograms (XRD) using a Bruker Meas Srv (D2-208219)/D2-2082019 diffractometer that operates at 30 kV, 10 mA with Cu tube (λ = 1.54 Å), with a range from 0 to 100°. The crystallite sizes were calculated for ZnO NPs using the Scherrer formula. The surface structure was also investigated by Joel 6360LA scan electron microscope (SEM) for both ZnO NPs and barley straw. Thermogravimetric analysis (TGA) of the impregnated sample was carried out by TERIOS Universal V4.5A TA Instruments (New Castle, DE, USA) for barley straw before and after mechanical pretreatment. The prepared nanostructure ZnO (chemically and green) was characterized individually by transmission electron microscope (TEM) (JEOL, Model JSM 6360LA, Tokyo, Japan). The mean pore diameter and specific surface area (BET (Brunauer–Emmett–Teller)) were measured on BELSORP (Mini II, BEL Japan, Inc., Osaka, Japan).

2.6. Kinetics Study and Statistical Analysis

Four kinetic models, i.e., the first-order kinetic model Equation (1), the modified Gompertz model Equation (2), the logistic function model Equation (3), and Cone model Equation (4), were selected to fit the cumulative biogas production obtained from the experimental data [33]. The most suitable kinetic model should be selected not only to predict the efficiency of particular reactors but also to correctly analyze the metabolic pathways [34,35,36].

All four kinetic models were used in this study to determine the cumulative biogas production potential, hydrolysis kinetics, lag phase duration, and maximum biogas production.

$$\mathrm{M}=\mathrm{Pb}\times \left(1-e^{\left(-kt\right)}\right)$$

(1)

$$\mathrm{M}=\mathrm{Pb}\times \exp \left\{-\exp \left[\frac{\mathrm{Rm}.\mathrm{e}}{\mathrm{Pb}}\left(\mathsf{\lambda }-\mathrm{t}\right)+1\right]\right\}$$

(2)

$$\mathrm{M}=\frac{\mathrm{Pb}}{\frac{1+\exp \{4.\mathrm{Rm}.\left(\mathsf{\lambda }-\mathrm{t}\right)\}}{\mathrm{pb}}+2}$$

(3)

$$\mathrm{M}=\mathrm{Pb}/\left[1+{\left(\mathrm{kt}\right)}^{-\mathrm{n}}\right]$$

(4)

where M is the biogas yield (L/g VS added) with respect to time t (days), Pb is the maximum biogas potential of the substrate (L/g VS added), k is the hydrolysis rate constant (1/day), t is the time (day), Rm is the maximum biogas production rate (L/g VS added), λ is the lag phase time (days), e is Euler’s function equal to 2.7183. The coefficient of determination (R²) and root mean square error (RMSE) were calculated for all models to compare the accuracy of the studied models, which was determined using SPSS 15, Origin 2020b, and Excel 2010 software. RMSE, given by Equation (5), was taken as the standard deviation between the measured and predicted values with a lower RMSE, indicating a well fit [37].

$$\mathrm{RMSE}=\sqrt{{\displaystyle \sum }_{\mathrm{i}=1}^{\mathrm{n}}(\frac{\mathrm{PVi}-\mathrm{MVi}{)}^2}{\mathrm{n}}}$$

(5)

where PVi is the predicted value, MVi is the measured value of the biogas volume, and n is the number of the measurements.

2.7. Cost Analysis and Economic Indices

The economic feasibility of biogas production depends on the possible income from the produced biogas versus the total cost of production. Both investment and operational costs, including biomass supply, are affected by several site-specific conditions. The cost-benefit analysis performed in this study was based on the whole chain of crop cultivation (preparation of land, planting, pesticide, and fertilizer), harvesting, transport, and conversion of the crops at a biogas plant. The costs for these variables are based on prices in Egypt, where the electricity price is 0.064 €/kWh, the total cultivated area for barley crop is 101,172 ha, with an average yield of 2.96 t DW/ha (personal communications with Egyptian agriculture ministry). Biogas contains roughly 50–70 percent methane, 30–40 percent carbon dioxide, and trace amounts of other gases. In this work, we used 60% for methane production as the proposed percentage to calculate the cost benefits analysis [38]. Moreover, in this work, we used a 40% efficiency of conversion from the literature [39]. Scheme 1 shows the different parameters considered for the cost-benefit analysis.

The LCOE technique has a slighter emphasis as it assesses the producing energy costs from a sole technology. In this technique, elements and resources of the energy system conversion are encompassed, thereby excluding storage, exchange, and final demand effects. This technique usually estimates costs as €/KWh or a dissimilar unit that characterizes the cost of energy generation. The LCOE costs are calculated according to these inputs present in the following Equation (6):

LCOE = Σ_t [(I_t + M_t + F_t)/(1 + r)^t]/Σ_t [(E_t/(1 + r)^t]

(6)

• The initial cost of investment expenditures in year t (I)
• Maintenance and operations expenditures in year t (M) = 10% of the initial investment cost [38,39]
• Fuel expenditures (if applicable) in year t (F)
• The sum of all electricity generated in year t (E)
• The discount rate of the project (r)
• The life of the system (n)

Calculating the LCOE is linked to the conception of assessing a project’s net present value (NPV). NPV is the value of all future cash flows over the entire life of an investment discounted to the present. The formula for the net present value is (Equation (7)):

NPV = Σ_i (Cash flows)/(1 + r)i − Initial Investment

(7)

Cash flows = cash flows in the time period, r = discount rate, and i = time period.

The calculations for cost-benefit analysis and LCOE were made only for green ZnO NPs due to their higher biogas yield than chemical ZnO NPs. The investment costs used to calculate LCOE and NPV are shown in

Table 1. Investment costs for mechanical and chemical treatment of barley straw to calculate LCOE and NPV.

Parameters	Cost
Initial Investment Cost (€)	1000
Annual Operations and Maintenance (O&M) Costs (€)	100
O&M Growth Rate (%)	2%
Annual Fuel Costs (€)	-
Project Lifespan (years)	10
Discount Rate (%)	8%

.

3. Results and Discussion

3.1. GC-MS Analysis

A broad range of compounds, such as phenols, alcohol, esters, and ethers, can be observed in the GC-MS chromatogram. The GC-MS analysis of ethanol extract of A. plumula shows 11 chemical compounds in Figure 3. Among these compounds, 7.11 min is dodecamethylcyclohexasiloxane, 7.92 min is 5-octadecenal, 9.39 min is 2,4-bis (1,1-dimethylethyl) phenol, 10.26 min is 3-hydroxyspirost-8-en-11-one, 13.41 min is estriol 16-glucuronide, and 14.82 min is 9-desoxy-9x-chloroingol 3,7,8,12-tetraacetate. An assumption of how bioreduction is promoted by these bioactive molecules is as follows: at the initial stage, the metal ions undergo the activation phase, where the growth rate of particles is usually slow as the metal ions are reduced from their salt precursors by the action of plant biomolecule metabolites with reduction capabilities, as shown in Scheme 2. In this work, the reduction of metal ions occurs as a result of biomolecules (2,4-bis(1,1-dimethylethyl)phenol) present in A. plumula.

3.2. Characterization for Barley Straw

3.2.1. Fourier Transform Infrared Spectra (FTIR)

The FTIR shows the overlapping spectra of raw and mechanically pretreated barley straw in Figure 4. The decrease in intensity spectrum of pretreated barley in comparison with raw barley straw in the bands at 1060–1645 cm⁻¹ and the broadband at 3000–3820 cm⁻¹ suggests the occurrence of deformation in the chemical structure of pretreated barley straw as a result of applying mechanical degradation of lignocellulose [40].

3.2.2. X-ray Diffractometry (XRD)

X-ray diffraction analysis is carried out to evaluate the crystallinity degree of the raw and pretreated barley straw, as shown in Figure 5. The crystallography exhibits that the peak intensity of the raw barley sample is at 15.5 and shifts to 22.4 after mechanical pretreatment. A small peak appears post-pretreatment, observed at 15.5, and this corresponds to crystalline cellulose II. The composition of the biomass is highly influenced by crystallinity. The raw material has the lowest relative crystallinity as it has a higher amorphous content of hemicellulose and lignin, and the barley with a high content of lignin and hemicellulose is the barley that presents lower crystallinity, and it could be evidenced that the crystallinity of the pretreated barley increases when treated mechanically [41].

3.2.3. Scanning Electron Microscopy (SEM)

The SEM images presented in Figure 6 reveal a significant difference between the raw and mechanically treated barley. The results suggest that the raw barley is a sheet structure consisting of fibers connected to each other by a wide pinhole, whereas the pretreated barley straw is broken with tiny pores, and its cell wall has become quite vulnerable due to small particle size to promote the exposure of the cell wall to AD and enhancing biogas production [42].

3.2.4. Thermogravimetric Analysis (TGA)

The biomass thermal stability is examined by means of TGA, the extensively approved method to define the biomass thermal degradation [43,44]. One reflects the evaporation of extractives’ desorption of moisture at 100–200 °C, while the others stand for degradation of cellulose and lignin at 300–350 and 300–500 °C, respectively, as shown in Figure 7.

3.3. ZnO NPs Characterization

3.3.1. Fourier Transform Infrared Spectra (FTIR)

The FTIR spectra are verified in a frequency range of 400–4000 cm⁻¹ to detect the structure of both chemically and green prepared ZnO NPs, as shown in Figure 8. The peak at 557.44 cm⁻¹ in Figure 8 corresponds to ZnO, confirming the formation of ZnO NPs [45]. The peaks at 887.21, 1408.08, and 1627.9 cm⁻¹ correspond to C–H, C–C, H–O–H, respectively, and are related to an organic compound. The broad peak around 3452.05 cm⁻¹ is corresponding to the OH, which represents the presence of water molecules on the surface of ZnO NPs [46,47,48,49,50,51].

3.3.2. X-ray Diffractograms

The XRD diffraction pattern of chemically and green synthesized ZnO NPs has been shown in Figure 9. The peaks 2Ө are indexed as 31.94 (100), 34.54 (002), 37.08 (101), 47.74 (102), 56.78 (110), 63.02 (103), 66.58 (200), 69.14 (201), 69.36 (201), and 77.28 (202). The obtained peaks demonstrate that the powder is highly crystalline, and all peaks agree with the hexagonal structure that agrees, as stated in the literature [33,45,52,53]. Great purity and crystallinity of the ZnO NPs are shown by the presence of a strong, sharp peak, and the nonappearance of peaks from other zinc oxide and impurity phases. The Scherrer formula is used to calculate the particle sizes and is found to be in the range of 2.7–3.7 and 2.5–3.5 nm, for the chemical and green synthesized ZnO NPs, respectively. The crystallite sizes of the ZnO NPs in

Table 2. The calculated crystal size of green and chemically synthesized ZnO NPs.

Position	Crystal Size (nm) Chemical ZnO NPs	Crystal Size (nm) Green ZnO NPs
31.94°	3.5	3.3
34.54°	3.7	3.5
37.08°	3.4	3.2
47.76°	3.2	3.0
56.78°	3.1	2.9
63.02°	2.7	2.5
66.58°	3.3	3.1
69.14°	2.6	2.4
69.36°	2.7	2.6
77.04°	3.4	3.2

are estimated by the full width at half maximum (FWHM) of the 101 anatase peak by the Debye–Sherrer Equation (8) [54,55].

C_s = 0.89λ/βcosθ

(8)

where C_s, λ, θ, and β indicate the crystallite size, the X-ray wavelength (1.5406 Å), the Braggs’ XRD diffraction angle, and the FWHM in radians, respectively.

3.3.3. SEM Microscopy and EDX

Figure 10 shows the morphologies of ZnO NPs structures with spherical morphology grains like and smooth, but clustered grains are composed of many tiny crystallites and clearly illustrate particle loose aggregation. The aggregation of particles (or the creation of larger particles) would have resulted from the large specific surface area of ZnO NPs and the high surface energy, and the synthesized ZnO NPs have a diameter of approximately 5 nm. The energy-dispersive X-ray spectroscopy (EDX) shows the presence of el ts Zn and O in the synthesized ZnO sample.

Table 3. EDX spectra of chemical and green synthesized ZnO NPs to determine the elemental contents of typical points on the surface.

Elements	Chemical	Green
Zn	85.01	89.4
O	5.72	5.18
Na	9.27	5.42
Total	100%	100%

indicates the elementary composition together with their weight percentage of chemically and green synthesized ZnO samples, where the contents of Zn on the chemical and green surfaces are 85.01 and 89.4 mass %, respectively, which specifies the greater purity of the green ZnO NPs.

3.3.4. Transmission Electron Microscope (TEM)

TEM photograph for morphology and distribution of NPs is giving information about the inner core and also gives a more precise finding of particle sizes, as shown in Figure 11. It is evident that the spread nanospheres mixed with little nanorods and incomplete growth of nanorods are formed for both chemically and green synthesized ZnO NPs. The particle size, on average, ranges between 6 and 55 nm.

3.3.5. BET Analysis Surface Area

Table 4. BET analysis surface area and the porosity of chemical and green synthesized ZnO NPs.

Sample	BET Surface Area (m2/g)	Mean Pore Diameter (nm)	Total Pore Volume (cm3/g(
Chemical ZnO NPs	24.33	10.370	0.0631
Green ZnO NPs	46.47	12.728	0.1479

demonstrates using the Brunauer–Emmett–Teller (BET) surface area, and the porous nature of the ZnO NPs is studied. The surface area of ZnO NPs is ranged from 24.33 to 46.47 m²/g, and the pore volume is ranged from 0.06310 to 0.1479 cm³/g for both chemical and green synthesized ZnO NPs at the temperature of 550 °C. The average pores diameter is less than 50 nm, which is consistent with the mesoporous material characteristics.

3.4. Chemical Compositions of Barley Straw

The TS content of the studied barley straw is about 52%, as shown in

Table 5. The chemical compositions of barley straw.

Component	Barley Straw	Inoculum
TS%	51.9	8.40
Ash%	5.05	28.30
VS%	94.05	71.70
N	1.56	4.30
C	51.15	49.90
H	6.42	5.50
C:N	33.04	9.07

. The operated barley straw has no S content. On the other hand, the determination of C and N content is detected by using an elemental analyzer, and the measurement method is carried out following [56]. The C:N ratio is about 33%, as presented in Table 5. Most of the literature works recommend an operating C:N ratio between 20 and 30, with the optimum ratio of 25 for anaerobic bacterial growth in an AD system [57]. The AD of the substrate at an improper C:N ratio will release high total ammonia nitrogen and/or volatile fatty acids (VFA) accumulation in the digester [5]. However, for better methanogenic performance, the optimum C/N ratio is 16–19% when considering hardly degradable complexes like lignin [58,59,60].

3.5. Effect of Both Mechanical Pretreatment and ZnO NPs on Biogas Production

The biogas production yield is gathered throughout a period of 30 days and charted in Figure 12. The higher biogas production outcomes during the 1st week also agree with the previous literature works [17,61], followed by inactivity, which is probably due to the methanogens undergoing a metamorphic growth process [62].

The startup of mean biogas production yield is slightly enhanced when the mechanically treated barley straw is treated with 5 and 10 mg/L ZnO NPs compared with the biogas production yield without using both chemical and green synthesized ZnO NPs, as shown in Figure 12. The low doses of both chemical and green synthesized ZnO NPs (5 mg/L) have a significantly positive impact on biogas production (p < 0.05). Using 10 mg/L ZnO NPs produces higher biogas yield than 5 mg/L. It is also clear to notice that ZnO NPs concentration of 20 mg/L of both chemical and green synthesized ZnO NPs has negative effects on the biogas production. The biogas tests end when the daily production is less than 1% of the overall production for most of the operated tests, as noticed in Figure 13. On the other hand, it is clear to see that the mechanically treated barley straw of size 0.4 mm yields a biogas production of about 340 mL/g VS, which is higher than that of the untreated barley straw of size 4 mm, which yields about 279 mL/g VS.

Mechanical pretreatment is the first step in splitting the constituents of lignocellulose, which are cellulose, lignin, hemicellulose, and other extracted components, and are related to the hydrogen bonds that bind the cellulose chain strongly in crystal form, which hinders the degradation of cellulose to glucose [5,14,15,16]. To solve the problems of biogas production based on lignocellulose and to enhance the efficiency of biogas production, it is essential to mechanically pretreat the substrates to increase feedstock utilization towards AD, which are to be fermented and NPs supplied with the substrate to enhance the enzymatic activity.

Besides, this study is the first study that examines the impact of mechanically treated barley straw in combination with the green synthesis of ZnO NPs on biogas production, which has resulted in a greater biogas yield in comparison with chemical ZnO NPs. The mechanical pretreatment enhances the surface area of reaction and biogas production, as confirmed by FTIR, XRD, SEM, and TGA. It is clear that mechanically pretreated barley straw (0.4 mm) has a good biogas yield, more than the untreated raw barley straw (4 mm) and with the same dosage of ZnO NPs due to the mechanical treatment, which increases the surface area of the reaction and enhances the AD, as shown in Figure 12. On the other hand, for 5 and 10 mg/L ZnO NPs, the mechanically pretreated barley straw has higher biogas yield due to the combined effect of both mechanical and chemical treatment.

Finally, for the 20 mg/L ZnO NPs, even if they are mechanically treated, the higher concentrations of ZnO NPs have inhibitory effects on the methanogenesis bacteria. From the above results, the order of biogas production can be arranged as mechanically and chemically treated barley straw with lower concentrations (5 and 10 mg/L) > mechanically treated barley straw 0.4 mm > raw untreated barley straw 4 mm > mechanically and chemically treated barley straw with higher concentrations (20 mg/L).

3.6. The Proposed Mechanism of ZnO NPs in Biogas Production

The specific effects of ZnO NPs on microorganisms in the AD system are hard to explain because the species of bacteria are not identified in the current study. In comparison, the exposure to ZnO NPs at various concentrations has different effects on the yield of biogas. ZnO NPs cause biogas inhibition at a large concentration (20 mg/L), about 50% for both green and chemical ZnO NPs, and the severity of the effect is directly related to the exposed concentration, which agrees with the reported data [63,64]. At a tolerable exposure of low concentrations of green (ZnO at 5 and 10 mg/L), there is a slight increase in the biogas production of about 10.3 and 14.9% and about 9.7 and 13.2% of chemical (ZnO at 5 and 10 mg/L), which agrees with the reported data [21,61], which have similar results with durum wheat.

When NPs are applied to the digester, the bacterial toxicity of NPs is reduced. This might have been due to agglomeration and adsorption into biomass, and this might be the reason for low toxicity in small concentrations of ZnO NPs, but this needs to be studied in depth for a clearer understanding, as previously mentioned [64]. ZnO NPs are actually dispersed in the solvent, not dissolved, and therefore, they cannot release Zn²⁺ ions [64,65]. This may also explain why in our work, the low concentrations of ZnO NPs (5, 10 mg/L) do not release enough amount of Zn²⁺ ions that could inhibit bacterial activity. Moreover, the enhancement of biogas yield is due to the stress of anaerobic bacteria by introducing low dosages ZnO NPs, which enhance its resistance and may affect the bacterial activities, resulting in this slight increase in the biogas yield than control (mechanically treated barley straw with 0 mg/L ZnO NPs), but this also still needs further studies. These explanations may agree with previous work [66], who has studied the aerobic denitrifying bacteria and mentioned that to prevent ZnO NPs entering cells by adsorption, the production of extracellular polymeric substances (EPS) of two studied strains is increased by 13.2% and 43.9%, respectively. The up-regulation of amino sugar and carbohydrate-related metabolism has contributed to the increase of EPS production, and the increased nitrogen metabolism has contributed to higher activities of nitrate and nitrite reductases. The same study should be done in the future for anaerobic bacteria to study its metabolism pathways.

However, adding ZnO NPs 20 mg/L has made negative impacts on the biogas yield. This could be explained by a large amount of toxic Zn²⁺ ions, which are released from ZnO NPs that may damage the cell membrane of anaerobic bacteria, and then the anaerobic bacterial activity is reduced, even causing some anaerobic microbial death [65]. In our work, the amount of Zn²⁺ ions released by a high dose of ZnO NPs 20 mg/L is sufficient to reduce biogas yield by half, inhibiting the anaerobic bacteria. Since ZnO is amphoteric, it reacts with acids as well as alkalis, giving Zn²⁺ ions as in Equations (9)–(11) [64].

ZnO + HCl    acidic medium  ZnCl₂ + H₂O

(9)

ZnO + 2NaOH  acidic medium Na₂ZnO₂ + H₂O

(10)

ZnCl₂ + H₂O    aqueous medium  Zn²⁺ + 2Cl⁻

(11)

The free Zn²⁺ ions are instantly bound to the biomolecules due to their positive charge and negative charge on the biomolecules [64] like proteins and carbohydrates, and they no longer serve any essential role in the bacteria, as in Equation (12). Zn²⁺ ions are not always 100% bioavailable and may change invariably with physiological, redox potential, and pH [64,67], which needs more future studies.

Zn²⁺ + Biomolecule   Zn-biomolecule complex

(12)

Mu and Chen [20] found that the methane yield was reduced by 18.3 and 75.1% from the control in the presence of 30 and 150 mg ZnO NPs/g TS, respectively. A high part of the reduction was usually related to the release of Zn²⁺ from the ZnO NPs. The reduction due to the release of Zn²⁺ ions was 9.4 and 63.8% for the equivalent of 30 and 150 mg ZnO NPs/g TS dose compared to the control, respectively. This would indicate that at higher ZnO NPs doses, the reduction is mainly caused by the release of Zn²⁺ rather than the NPs [68].

Wang et al. [63] mentioned that lower concentrations (1.3 and 4.6 μg/mL) of Fe²⁺ were found to enhance the AD, whereas higher concentrations (3.3 and 9.8 μg/mL) of Ag+ and Mg2+ were found to reduce the AD. Comparing the AD of sludge induced by nano zerovalent iron (nZVI), Ag NPs, or MgO NPs with that induced by the corresponding amounts of Fe²⁺, Ag⁺, and Mg²⁺, it was found that the released Fe²⁺, Ag⁺, and Mg²⁺ were primarily responsible for the enhancement and/or inhibition impacts of nZVI, Ag NPs, and MgO NPs [63]. The contact between NPs and the bacterial cell wall is enough to make toxicity [64]. If it is right, then higher quantities of metal NPs are essential so that the bacterial cells are totally enveloped and protected from its environment, leaving no chance for nutrition to be absorbed to continue the life process. This may explain why in our work, the higher concentrations of ZnO NPs cause inhibition for biogas yield and reduce it by 50%.

It has been observed that the growth inhibition of microbes increases with raising the concentration of NPs. The incubation duration is supposed to be increased; the inhibition of growth will increase without any major changes in the mechanism of action [64,69]. For most experiments, the metal ions emitted from NPs play a significant role in the biological processes of microorganism populations. Some scientists found that during the sewage sludge treatment process, higher metal ion dosages released from NPs have prevented these microorganisms from working [63].

Ganzoury et al. [70] specified that the higher surface area of NPs has a positive impact on the AD course. In this study, the higher biogas yield from green ZnO NPs than chemical one may be due to its higher surface area, as presented in Table 4. Other literature works [21,71] have shown that the green synthesized NPs are more stable and less toxic than chemical NPs, and this is one of the reasons to choose the extract of A. plumula in our study.

3.7. Kinetic Study

Table 6. Results of the kinetic study using the first order and cone models.

First-Order Kinetic Model
Chemical
Samples	R2	PredictedP(mL/gVS)	Differences%	K(1/day)	RMSE
(4 mm)	0.995	282.03	1.1	0.387	5.5
(0.4 mm)	0.992	343.78	1.1	0.399	8.3
5 mg/L	0.995	373.50	0.1	0.356	7.2
10 mg/L	0.997	379.37	1.5	0.347	5.4
20 mg/L	0.977	172.08	0.6	0.44	6.9
Green
5 mg/L	0.993	380.93	1.6	0.362	8.3
10 mg/L	0.999	386.53	1.1	0.334	3.9
20 mg/L	0.967	166.72	3.6	0.537	7.6
Cone Model
Chemical
Samples	R2	PredictedP(mL/gVS)	Differences%	K(1/day)	RMSE
(4 mm)	0.760	275.96	1.1	0.329	21.62
(0.4 mm)	0.754	336.15	1.1	0.344	27.3
5 mg/L	0.758	363.90	2.4	0.316	30.83
10 mg/L	0.764	370.41	3.8	0.299	29.81
20 mg/L	0.745	168.01	1.7	0.4	13.86
Green
5 mg/L	0.757	371.05	1.1	0.324	31.4
10 mg/L	0.769	377.73	12.6	0.283	29.4
20 mg/L	0.742	163.49	5.5	0.478	11.9

and

Table 7. Results of the kinetic study using the modified Gompertz and logistic function models.

Modified Gompertz Model
Chemical
Samples	R2	PredictedP(mL/gVS)	Differences%	RmaxmL/gVS.day	λ (day)	RMSE
(4 mm)	0.991	279.03	0.01	1.46	0.736	6.24
(0.4 mm)	0.994	340.28	0.08	1.45	0.785	5.77
5 mg/L	0.995	369.73	0.87	1.60	0.651	4.58
10 mg/L	0.991	375.09	2.57	1.58	0.624	7.07
20 mg/L	0.982	171.24	0.14	1.37	0.777	5.30
Green
5 mg/L	0.994	377.47	0.66	1.58	0.653	5.11
10 mg/L	0.988	381.93	2.20	1.60	0.852	8.23
20 mg/L	0.962	166.12	3.98	1.11	0.900	7.29
Logistic Function Model
Chemical
Samples	R2	PredictedP(mL/gVS)	Differences%	RmaxmL/gVS.day	λ (day)	RMSE
(4 mm)	0.981	278.16	0.30	2.05	0.736	9.3
(0.4 mm)	0.986	339.32	0.20	2.01	1.16	8.00
5 mg/L	0.987	368.57	1.19	2.27	0.952	7.29
10 mg/L	0.981	373.69	2.94	2.27	0.917	9.98
20 mg/L	0.978	170.96	0.021	1.97	1.12	5.08
Green
5 mg/L	0.987	376.40	0.37	2.26	0.951	7.05
10 mg/L	0.976	380.14	2.65	2.31	0.863	11.52
20 mg/L	0.953	165.89	4.11	1.63	1.25	7.55

summarize the results of a kinetic study. The first-order, modified Gompertz, and logistic function models are found to have a good fit within the experimental data. In the experiment, mechanically treated barley straw (0.4 mm) with ZnO NPs concentration 20 mg/L exhibits the highest hydrolysis rates (K) of 0.537 d⁻¹ (based on the first-order model) and 0.478 d⁻¹ (based on the cone model). The maximum biogas production rate (Rm) of 1.60 L/g VS and 2.31 L/g VS is observed for the modified Gompertz model and the logistic function model, respectively. The late response and the subsequent adaptation of microorganisms to the fluctuating environment are expressed by the lag phase (λ) [37,72]. The modified Gompertz and the logistic function models have attained λ value of 0.9 days and 1.25 days, respectively. The value of λ, in this study, is relatively close to the previously reported λ of 1.2–1.8 days and 1.5–2.1 days (by Deepanraj et al. [73]) for the modified Gompertz model and logistic function model, respectively.

To evaluate the soundness of the model results in the four studied models, the predicted values for biogas production are plotted against the measured values, as presented in Figure 14. The low values—(3.9), (4.5), and (5.08)—of RMSE reflect the first order, modified Gompertz, and logistic function models and have a high ability to accurately predict the bioactivities, as opposed to the cone model (31.4). The statistical indicators (R²) are given in Table 6 and Table 7 to provide a picture of the kinetics study. Nguyen et al. [34] described that the higher value of R² (0.999, 0.995, and 0.987) and the lower values of RMSE for the first order, modified Gompertz, and logistic function models, respectively, indicated a more suitable kinetic model.

3.8. The Cost-Benefit Analysis

The data for cost-benefit analysis, such as TS and VS, are taken from the biogas experiment whenever possible. Where the extra data are needed, such as field preparation and harvesting, they have been sourced from the relevant literature and personal communications, as shown in Scheme 1.

Table 8. Cost-benefit for barley straw with and without mechanical pretreatment.

Cost	Units	Barley (4 mm)	Barley (0.4 mm)
Mechanical Treatment Cost for Barley Straw (chipping + milling)	€/ton	0	13.72
Chemicals (NPs + NaOH)	€/100g	0	0
Total	€/ha	871.11	884.83
Benefits
Electricity Production (40% eff)	KWh/ha	969	1181
Electricity Sales	€/ha	62	76
Total	€/ha	62	76
Net Benefits	€/ha	−809.11	−808.83

and

Table 9. Cost-benefit for barley straw with combined mechanical and chemical pretreatment.

Cost	Units	5mg/L (0.4 mm)	10 mg/L (0.4 mm)	20 mg/L (0.4 mm)
Mechanical Treatment Price Needed for Barley Straw (chipping + milling)	€/ton	13.72	13.72	13.72
Chemicals (NPs + NaOH)	€/100g	5.52	5.52	5.52
Total	€/ha	890.35	890.35	890.35
Benefits
Electricity Production (40% eff)	KWh/ha	1302	1356	601
Electricity Sales	€/ha	83	87	38
Total	€/ha	83	87	38
Net Benefits	€/ha	−807.35	−803.35	−852.35

show that the use of barley straw for biogas production is a practical option, and the biogas yield is comparable to previous studies [16]. However, it may be not profitable if barley straw is used instead of the current energy crops being used, such as wheat or maize.

From the results of the cost-benefit analysis shown in Table 8 and Table 9, it is observed that with current costs and revenues, it is not profitable to use barley straw for biogas production. The significant differences in cumulative biogas production between the treated and untreated barley straw lead to higher specific methane yield. These differences translate to higher electricity production and higher revenues through electricity sales. Solely from the point of view of the revenues generated, the mechanically and chemically treated barley straw 10 mg/L (0.4 mm) gives the highest economic value. The net benefit of this cost-benefit analysis shows that the mechanically and chemically treated barely straw 10 mg/L (0.4 mm) is closest to profitability than the mechanically and chemically treated barely straw 5 mg/L (0.4 mm).

It is clear that to generate a profit in the future, it’s a must to minimize costs. The cost of field preparation, crop cultivation, and fertilizer and pesticide application equals 542.35 €/ha, and to save this cost, the use of other crops, such as perennial crops that do not compete with the food chain and aquatic plants, appears to be the best option. Along with minimizing costs, there are some options to increase biogas production and associated revenue, such as digester type, co-digestion, and pretreatment method, which are considered the major factors that influence biogas production.

3.9. Levelized Cost of Energy

The LCOE depends on the biomass processing and treatment techniques, such as field preparation, crop cultivation, fertilizer and pesticide application, and mechanical and chemical treatment. The investment costs used to calculate LCOE and NPV are shown in Table 1. Figure 15 represents the LCOE of the untreated and treated barely straw. From Figure 15, it’s clear to see that the combined mechanical and chemical treatment with 10 mg/L ZnO dosage can produce higher energy output. The LCOE decreases as the energy output (NPV) increases, as noticed in Figure 15. Specifically, the LCOE is computed based on the net present value of all costs (NPV) divided by the total amount of energy produced over the energy system’s lifetime [73]. The lowest value of LCOE is obtained for mechanically and chemically treated barley straw (0.4 mm + 10 mg/L ZnO), and the highest value is obtained for mechanically and chemically treated barley straw (0.4 mm + 20 mg/L ZnO).

The average LCOE for the best mechanical and chemical treatments is 0.21 €/kWh, which can be considered non-competitive with the other candidate renewable energy systems, such as wind energy 0.068 €/kWh [74,75]. Moreover, the LCOE from biomass energy in this study is 200% higher than the LCOE from the biomass energy in Europe (0.088 €/kWh) in 2019 [76]. The reason for this high LCOE, as mentioned before, could be eliminated by choosing another aquatic crop with a high yield of biogas and choosing the suitable treatment techniques. Besides, changing the type of NPs, for example, iron oxide NPs, might have a good impact on the treatment processes and can be a good practical solution, as mentioned [63].

4. Conclusions

From the aforementioned results, it can be concluded that the mechanical pretreatment methods have improved the biodegradation of barley substrates and contributed to biogas enhancement with biogas yield 340 mL/g VS without adding ZnO NPs. There is a successful synthesis of green synthesized ZnO NPs using A. plumula extract, which contains phytochemicals components, acting as capping and stabilizing agents. Both morphology and the size of the synthesized ZnO NPs have been confirmed by SEM, FTIR, TEM, and EDX. The use of ZnO NPs in biogas is dosage-dependent, where the improvement of the biogas production is attained by adding 5 and 10 mg/L of ZnO NPs for mechanically treated barley straw. In particular, the biogas production yield is increased from 340 mL/g VS, using the barley biomass without ZnO NPs, to 385 and 390.5 mL/g VS, using 10 mg/L of chemical and green synthesized ZnO NPs, respectively. The green synthesized ZnO NPs provide a slightly higher biogas production yield than chemical synthesized ZnO NPs by 1 and 2% for 5 and 10 mg/L, respectively. The improvement of biogas production is about 14.9 and 13.2% for 10 mg/L of chemical and green synthesized ZnO NPs, respectively, more than mechanically treated barley straw of size 0.4 mm. But this improvement is still far from other NPs, such as nano zerovalent iron (nZVI) and Fe₂O₃ NPs with concentrations of 10 and 100 mg/g TSS, yielding more cumulative methane production levels at 120 and 117%, respectively, of the total biogas yield [63]. The mechanically treated barley straw of size 0.4 mm yields a higher biogas production of about 340 mL/g VS than the untreated barley straw of size 4 mm, which yields about 279 mL/g VS. Among the four kinetic models, the first-order model (R² = 0.999), modified Gompertz model (R² = 0.995), and logistic function model (R² = 0.987) are the most suitable models for fitting the measured biogas yield, and they could be used to describe the kinetics of the AD process more reasonably. Based on the obtained results, the mechanically treated barley straw is a suitable source of biomass for biogas production, and the yields are higher than the untreated barley straw, but the cost-benefit analysis and LCOE show that it is not currently profitable to use barley straw for biogas production due to higher annual total cost. As a future development, the environmental analysis of the life cycle assessment (LCA) type could be also needed to evaluate the eco-compatibility of the bioprocess from barley straw in its entirety.