1 Introduction

Today, environmental pollution caused by plastics and other non-degradable chemical-based polymers has become a serious global concern. So, numerous studies conducted on the use of biodegradable materials in different industries such as packaging, detergents, pharmaceutical, food, and others. In this subject, natural biopolymers, by having advantages such as lower cost, more accessibility, and higher biodegradability, attracted vast attention [1,2,3,4].

Lignocellulosic material, as the central part of plant structure, can be categorized as one of the significant promising renewable resources and their complex structure, consists of three primary polymeric materials, including cellulose, hemicellulose, and lignin. Cellulose, as the significant part of the plant cell walls, comprises 33% w/w of all vegetable matter. Cellulosic materials, as leading categories of carbohydrate macro-polymers composed of beta (1–4)-glycosidic linked chains of glucose monomers, are the most essential component of lignocellulosic material. Cellulose is the most abundant biopolymer in the world,with 1011–1012 tons per year of production by having particular characteristics like crystallinity, low solubility, and hydroxyl group’s reactivity attracted vast attention [5,6,7,8].

The distributed cellulose throughout nature is usually along with hemicellulose and lignin. Lignin, as a three-dimensional aromatic polymer bonded by a series of C–O, and C–C linkages, has a vital role in giving plants their structural integrity. Separation of different components of biomass before processing is necessary for better and more use of biomasses [8,9,10,11].

Among different resources which were studied for cellulose isolation, industrial residuals such as bagasse, sugar beet pulp, apple and carrot pulps, and orange peel, have been the subject of numerous investigations. Globally, cotton fiber is the dominant fiber that serves as a raw material for the textile industry, contributing at least $600 billion annually to the economy and about 25 million tonnes of cotton are produced worldwide each year. The cotton production in our region reaches 60 thousand tons. In this investigation, cotton pod, due to geographical abundance and the presence of different cotton cultivation lands, was selected as proper resources for cellulose isolation. The amount of cellulose in cotton pods is about 42% of the total dried mass, which can be used as a suitable source for cellulose separation and cellulose nanofiber preparation [12,13,14,15,16,17].

Also, various methods were investigated for cellulose isolation. Conventional separation methods require using organic solvents, a high amount of chlorinated materials, and highly acidic or alkaline solutions. Therefore, it is necessary to develop new and environmentally friendly methods to minimize the pollution caused by the use of conventional materials [16, 18,19,20,21,22,23].

Ionic liquids (ILs) have recently been considered green solvents due to their unique properties such as low vapor pressure and high thermal stability. Low vapor pressure is an essential characteristic of a solvent because it causes ease of outdoor application without worry about evaporation. Ionic liquids are formed from a combination of different anions and cations compounds whose properties such as hydrophobicity, polarity, and dissolution are adjustable [24,25,26,27,28,29,30,31].

Despite the promising properties of ILs, such as their excellent ability for the crystalline structure of cellulose degradation or removal of lignin/ hemicellulose, the disadvantages of these liquids, such as toxicity, low biodegradability, and high cost, have limited their use. On the other hand, the synthesis of most ionic liquids is not “green” and is not possible in industrial quantities [32,33,34,35,36].

Like as ILs, eutectic solvents (ESs), due to their unique properties, have attracted vast attention. ESs usually have low vapor pressure due to their low volatility, and low melting point and also they don’t have flammability, and toxicity, which provides the promising potential for biocatalysis, biodegradation, and treatment. Also, these types of solvents are well-known for their biocompatibility, biodegradability, and recyclability that make possible use of them more economical. Compared with ILs, ESs are easy to prepare from cheaper materials with high purities. Moreover, they have less moisture sensitivity, and this advantage makes it possible to use them in the case of wet biomass without drying and so the cost of drying significantly decreases [24, 37,38,39,40,41].

ESs preparation needs mild mixing of two or more solid-phase or liquid–phase or solid–liquid phase chemicals for the preparation of joint structure at 130 °C or less. These solvents prepared by hydrogen bonding comprise at least one hydrogen bond donor (HBD) such as carboxylic acids and one hydrogen bond acceptor (HBA) like quaternary ammonium salts, resulting in a clear solution with a low freezing point. It is necessary to mention as Abottt and his coworkers declared only a special ratio of HBD and HBA called deep eutectic solvents(DESs) and other prepared solvents which have similar properties to DESs but have more melting temperatures should be called ESs [32, 36, 37, 42,43,44,45,46].

DES has the potential to disrupt the resistive properties of biomass and hydrogen bond interactions within the chains and also crystallinity structure in mild conditions by hemicellulose and lignin removal while sugar loss due to the structure destruction is reduced. With this regard eutectic solvents (ESs) green treatment of lignocellulose biomasses attracted vast attention [47, 48].

The properties of ES had great impacts on the treatment effect and different types of ESs were prepared by mixing different kinds of HBAs and different kinds of HBDs with different molar ratios. The final properties of DES were affected by all characteristics of components in DES [49, 50].

Choline chloride–lactic acid was used as a reference in delignification trials and the results show that a high selectivity for the separation of lignin from lignocellulosic biomass and cellulose isolation which is related to the prepared ES ratio. They showed that very different solubility values were obtained for the different combinations of choline chloride and carboxylic acid. Choline chloride–lactic acid mixtures show high solubility for lignin, while cellulose was found to be immiscible with the whole series [11, 24, 49, 51].

Also, the investigation showed in the mild condition of ES treatment the temperatures and duration hours have a great impact on lignin removal and cellulose isolation (Procentese et al. 2018; Xu et al. 2016 [52].

Concerning the subject, this paper aims to apply choline chloride-lactic acid-based ESs for delignification and cellulose isolation optimization from extractive-free natural cotton pods (EFNCPs) to achieve the highest delignification and cellulose isolation and purity and consumption of ES-treated cotton pods (ESTCPs) for nano-fibrillation.

To optimize the lignin removal and cellulose extraction, modeling of influencing parameters is required. Different methods such as central composite design, Box-Behnken, Mixture Design, and Taguchi can be applied for experimental design and optimization of process or production [53,54,55,56].

The Response Surface Method (RSM) as a well-known method, is applied in experimental design, model building, and optimization. RSM as a multivariate method, which is the combination of statistical and mathematical techniques, can help analyze problems, where a relationship between the response and the independent parameters through a response function exists. Influencing parameters on cellulose extraction have complex interactions with each other, and due to this fact, data interpretation unless with the application of the suitable method is so difficult [23, 54].

In this research in the novel experimental and mathematical investigation, choline chloride-lactic acid-based ESs with different molar ratios were prepared and used for biomass treatment at different temperatures and times for the delignification and cellulose isolation from extractive free natural cotton pods (EFNCPs) in a mild condition. The delignification and cellulose isolation amount was studied as a response to the treatment process. The cellulose-enriched biomass at the optimum condition was applied for the post-treatment process and pure cellulose isolation. For this purpose, choline chloride lactic acid pretreated biomass was used for high purity cellulosic structure achievement by using biocompatible materials with less impact on the thermal and mechanical resistance. Prepared chemically purified celluloses (CPC) was applied for nano-fibrilization by ESs which has less influence on the mechanical and thermal strength of prepared cellulose nano fibers (CNFs). Totally, the viewpoint of this research was achieving the best cellulose isolation and nano-fibrilization alongside with special mechanical and chemical properties by using green- less environmental impact method.

2 Experimental, modeling, and optimization

2.1 Experimental

2.1.1 Material

Natural cotton pods (NCPs) were prepared from Moghan agro-industrial zone. Ethanol (98% v/v), toluene (98% v/v), acetic acid (98% v/v), H2O2 (35%v/v), and sodium hydroxide (98% w/w), all in the analytical grade obtained from Dr. Mojallali chemical company. The other materials, including choline chloride (98%w/w), lactic acid (98%v/v), peracetic acid (40% v/v), and sodium chlorite (98%w/w) purchased from Sigma-Aldrich company. Deionized water was used in all experiments.

2.1.2 Sample preparations

To prepare NCPs powder as a by-product for the isolation process, the sample was powdered with the aid of a laboratory grinder into fine particles and sieved properly through 150 µm screen size meshes. The prepared powder oven dried for 72 h at 50 °C and was used for the extractive-free biomass preparation (after this time the moisture first content of 5% reached 0.1%).

2.1.3 Extractive-free biomass preparation

To prepare EFNCPs for the extraction process, firstly, they were milled into fine particles and extracted in the Soxhlet extraction system, according to the ASTM D-1105–96 (American Society for Testing and Materials) standard. The extraction process was conducted in three steps and each step respectively with 50 ml of ethanol-toluene (1:2) solution, ethanol, and water as a solvent with 3 g of the sample placed in the thimble filter of the Soxhlet apparatus and incubated for 4 h.

The sample after each step was transferred to a Büchner funnel for filtration and the excess solvent was eliminated. After the last step, the filtrated biomass was dried at 90 °C for 8 h and used for the treatment process [57, 58].

2.1.4 Proximate chemical analyses

Moisture, ash content determination

Moisture content was determined by 44–40 of the AACC method (American Association for Clinical Chemistry- 1983). The ash content was measured by remaining residue determination after the combustion process at 550 °C for 16 h [23].

Determination of lignin content

A modified standard assay (TAPPIT222 om-06) was used to measure the amount of acid-insoluble lignin (AIL). This procedure, as the general method, can be used to determine acid-insoluble lignin in all unbleached pulps [45].

Approximately 150 mg of the extractive-free sample and 3 ml of 72% (v/v) cold sulfuric acid (about 10 °C) were poured into a 100-ml Erlenmeyer flask. The sulfuric acid addition was slowly along with mixing with a glass rod.

The sample was placed in a water bath at a temperature of 20 ± 1 °C and allowed to disperse for 2 h. After 2 h, 84 ml of deionized water was poured into a 100-ml Erlenmeyer flask, and the suspension was autoclaved at 123 °C for 45 min. At the end of this period, the autoclaved sample rapidly cooled and filtered using No. 1 Whatman filter paper. The filtrate solution was collected, and the residual was washed twice with boiling water to eliminate soluble lignin and residual acid.

The residue in the filter was oven-dried at 105 ± 3 °C until a constant weight was recorded.

After drying in the oven and cooling in the desiccator for 30 min, the dried AIL was weighed and the lignin percentage was determined according to the following Eq. (1):

$$Lignin \%=\frac{W\;DL}{W\;EFS}\times 100$$
(1)

W DL and W EFS were the weight of dried lignin and the extractive-free sample.

In this procedure, acid-insoluble lignin remained constant whereas the other carbohydrates hydrolyzed[23, 57].

Holocellulose preparation and determination

Measurement of holocellulose amount (total cellulose and hemicellulose) was conducted by the Waise method with some modifications [23, 51, 57].

Holocellulose was separated from extractive-free biomass by the delignification process with sodium chlorite usage. For this purpose, approximately 500 mg of the EFNCPs was dispersed in 30 ml of 6 mg/ml sodium chlorite solution containing 0.04 mL of 10% v/v peracetic acid (pH 3.5).

The prepared mixture was incubated in the 85 °C water bath for 30 min. After this time, 200 mg of sodium chlorite powder and 0.04 mL of peracetic acid solution were added and this process was repeated seven times.

The sample was filtered through a No. 1 Whatman filter paper and washed 3 times with 50 ml of hot deionized water. The filtrate was discarded, and the residual dried at 105 ± 3 °C for 2 h.

The holocellulose percentage calculation is presented in the following Eq. (2):

$$\mathrm{Holocellulose} \%=\frac{W\;DHC}{W\;EFS}\times 100$$
(2)

W DHC and W PS were the weight of dried holocellulose and the extractive free samples respectively.

Determination of cellulose and hemicellulose

Cellulose content determination was done based on the KS M 7044 (Korean Standards Association method for alpha, beta, and gamma cellulose measurement (. In this way, approximately 300 mg of prepared holocellulose was poured into a 50 ml Erlenmeyer flask and 3 mL of 17.5% w/v sodium hydroxide was added to it and mashed with a glass rod for 5 min and incubated in a 20 °C water bath for 20 min. In the next step, 3 mL of distilled water was added, and the sample and mixed well. The resulting sample was filtrated through No. 1 Whatman filter paper and washed 3 times with deionized water. Subsequently, 2.4 mL of 10% v/v acetic acid was applied for naturalization. In the last step, the sample again was washed with hot deionized water and oven-dried at 100 °C until a constant weight was reached. The final weight is considered as cellulose weight according to Eq. (3) [57].

$$\mathrm{Cellulose} \%=\frac{W\;DC}{W\;DHC}\times 100$$
(3)

W DC and W DHC are the weight of dried cellulose and dried holocellulose samples.

2.2 Preparation of eutectic solution

For this purpose, the proper molar ratio of choline chloride and lactic acid carboxylic acid (respectively as acceptors and donors of hydrogen bonds molar ratios: 1:3,1:5,1:7,1:9, 1:11) were mixed in the incubator shaker at 60 °C until achieving a clear solution. The prepared solution due to dehydration of ammonium salts absorbs moisture and therefore, after preparation should be dried (60 °C for 12 h) and stored in closed containers [23, 32].

2.2.1 Investigation of lignin removal and cellulose separation by ES solvents

Eutectic solvents have a limited ability to dissolve the lignin and separate cellulose. To investigate the degree of dissolution, different molar ratios of lactic acid–choline chloride ES were prepared.

Next, to investigate the lignin removal, the appropriate weight ratio of EFNCPs completely dried powder and pre-dried ES (1:20 w/w) mixed together. The delignification process was investigated at different temperatures and times along with stirring in the oil bath.

After pretreatment with the ES at different temperatures and times, the same initial volume of boiling deionized water and ethanol solution was added and the separation process was performed for 30 min. This process was repeated two more times with boiling deionized water. The pretreated sample was put into a vial and placed in an orbital shaker incubator and mixed at 200 RPM at 50 °C for 20 min. The water was decanted and the water-washing process was repeated 3 times to ensure that the ES was removed totally (the product of this step is called ES-treated cotton pods (ESTCPs) powder).

Finally, the sample was drained with a 20-µm nylon filter and dried at 60 ± 2 °C oven for 12 h. The refined sample was used to measure lignin, hemicellulose, and cellulose in the structure.

As mentioned, ES has a limited ability to dissolve the lignin and separate soluble, and therefore the post-treatment process is necessary. This research aims to develope a green method for cellulose isolation, using biocompatible materials such as acetic acid, peracitic acid, water, and hydrogen peroxides with low concentration. After this treatment, cellulose remains with high purity [24].

In this stage, unlike the previous stage, due to the opening of the lignocellulose network, the time and conditions of treatment are much milder and there was no need to use a high concentration of materials. The post-treatment was done by the addition of 1 g of pretreated sample to 10 ml of H2O2 6% v/v which the pH of that was pre-adjusted at 3 by acetic acid. For this purpose, the mixture was completely stirred about 10 min at room temperature (this process was done for remaining lignin elimination). The treated sample at the last step was treated with 10 ml of 6% w/v NaOH solution containing 3% v/v H2O2 (to remain hemicellulose removal). The sample was washed three times by boiling deionized water and filtrated with No.1 Whatman paper and used for the last analyses and structure determination. The resulting material is called chemically purified cellulose (CPC).

2.2.2 Preparation of cellulose nanofibers

CPC has been used for cellulose nanofibers (CNFs) production with ES solvent. In this step, about 2 g of completely dried extracted CPC was treated with 30 ml of previously prepared and dried ES (with the 1:10 molar ratio of choline chloride-lactic acid, dried at 80 °C) for 30 min.

To complete ES removal, approximately 100 ml of hot deionized water was poured into the mixture and centrifuged at 4000 rpm and the solvent was eliminated. The process of washing was repeated 3 times. The prepared CNF dispersed well by ultra turrax homogenizer (t-25-IKA) and was freeze-dried at − 85 °C for 2 h and used for complementary investigations.

2.2.3 Cellulose characterization

Functional groups of EFNCPs, ESTCPs, CPC, and CNF were identified by Fourier transform infrared (FT-IR) spectra (The Tensor 27 spectrometers-Bruker, Germany).

In the sample preparation step, 2 mg of each sample was mixed with 200 mg of potassium bromide (KBr) and transmittance data (%) were recorded from 500 to 4000 cm−1.

Cellulose crystallization determination was done based on X-ray diffraction (XRD) method results. The XRD pattern data was collected on a SIEMENS D500 diffractometer (Germany) and Cu Kα radiation (λ = 1.54 Å) from 10 to 80° 2θ with the 0.25° step size per minute.

The cellulose Crystallinity Index (CrI) was calculated using the following Eq. (4):

$$\mathrm{CrI \%}=\frac{\mathrm{A\;cr}}{\mathrm{A\;sample}}\times 100$$
(4)

A in the presented equation is the area under the sample intensity curve associated with the total crystalline phase (for all calculations the background should be eliminated).

Thermogravimetric (TG) analysis and derivative thermogravimetric (DTG) analysis were performed in Mettler Toledo instrument. The experimental setup consisted of a 10◦C/min linear heating program from 30 to 600 °C, with 30 mL/min N2 low rates.

Field emission scanning electron microscopy (FE-SEM) (MIRA3 FEG-SEM, Czechia) and transmission electron microscopy (TEM) (LEO 906 E- Germany) microscope at an acceleration voltage of 120 kV were applied for surface morphology investigation of NCP, ESPC, CPC, and CNFs.

2.3 Computational details

Optimizing cellulose extraction conditions for achieving the maximum extraction amount is aimed to be done by application of the Response Surface Methodology-Central Composite Design (RSM-CCD) system. To develop an experimental database RSM method is applied and experimental results are used to achieve optimum isolation conditions.

2.3.1 DOE and statistical analysis

There are several ways to experimental design. The design of the experiment (DOE) method, as a strong tool was used for factor optimization in different processes and products to achieve the best condition. The RSM-CCD as a suitable design method can be used for polynomial modeling establishment and involved parameter efficiency determination to reach optimal conditions[23, 54].

A Central composite design (CCD) method with α =  ± 2 (to avoid decimal values and make easy control of preparation and process parameters) was used for studying the effect of three main parameters on cellulose isolation and delignification yield. The Software of Design Expert (8.0.7.1 version) was applied for experimental design. Three main parameters which have been selected for this purpose are the process temperature (°C), ES solvents ratios (mol acceptor: mol donor), and process time (h). The lower and upper levels and center points of these parameters were selected according to the previous studies[24, 52, 59].

Table 1 represents the actual and coded values of independent variables. The following Eq. (5) was used for encoding actual parameters:

Table 1 The levels of the independent operation variables in actual and coded value [24, 52, 59]
Full size table
$${X}_{i}=\frac{{X}_{i}-{X*}_{i}}{{\Delta X}_{i}}$$
(5)

Xi and xi respectively show the coded and uncoded value of the i’th parameter. xi* is the xi regarding value at the Central point in the studied area, and Δxi is the step size [54].

Due to the possibility of interaction between parameters the linear and zero-order models cannot correlate response and input variables correctly and have more lack of fit and also lower R2 values and so a second-order polynomial equation was used for correlation between response and involving parameters. The modeling was carried out by adjusting the second-order polynomial equation to the experimental results of the response. Analysis of variance (ANOVA) was performed to confirm the adequacy of the predicted model by assessing the lack of fit, regression coefficient (R2), and the p and F-values obtained from Analysis of variance (ANOVA). The validated model can be plotted in form of 2 and 3-D graphs to generate surface response for the determination of the best operation conditions.

The R2 value near unity shows the fitness quality of the model. The p values of parameters and models lower than 0.05 show the model terms are significant. Also, the p-value of more than 0.05 for lack of fit shows the model's ability to contribute to response and influencing parameters.

The response assessment process can be done by. The perturbation, and 2-dimensional (2D) contour plots based on the established model can clarify the effect of each parameter and the interaction between them. While flat lines in the perturbation plot indicate that the response is not sensitive to the change of a factor, a steep slope or curvature shape shows sensitivity to that [54].

To evaluate the differences between all parameters at a particular point (e.g., Central point) in the design space, the perturbation plots were applied. A steep slope or curvature form in a parameter shows response sensitivity to that. When the lines are relatively flat represent that response is not sensitive to change in specific factor [54].

Contour plots represent the relationship between the independent and dependent variables. These plots show the correlation between two variables on delignification and cellulose amount when the third parameter is kept constant. While the elliptical or saddle shape of contour plots represents the interaction considerability, the plot’s circular mode displays that the interaction between parameters can be neglected [54].

To obtain the optimum conditions for achieving the highest delignification and cellulose in the structure, the optimization tool of Design Expert software has been applied. This procedure has been done for maximization of delignification and cellulose isolation. Numerical optimization uses the models to search the factor space for the best trade-offs to achieve multiple goals. A verification experiment should be conducted under the predicted conditions and empirical responses that were close to the estimated amount shows the validity and adequacy of the obtained models.

3 Results and discussion

In this work, the effect of different choline chloride lactic acid ES pretreatment conditions on the efficiency of NCP delignification and cellulose isolation was investigated and optimized using the RSM-CCD method. The treated sample at the optimum condition was used for more process and cellulose Nano-fiber preparation.

Having an outlook of the initial structure and composition of NCP is necessary. For this purpose, chemical determinations of the main material in the NCP structure are processed according to declared methods. The experimental results of moisture, ash, lignin, hemicellulose, and cellulose determination in NCP are presented in Table 2.

Table 2 Percent of different materials in extractive-free natural cotton pods
Full size table

3.1 Mathematical results and modeling

3.1.1 RSM-CCD and ANOVA results

EFNCP with different amounts of lignin and cellulose in structure was investigated as raw material for delignification and cellulose isolation process with choline chloride ES at the different conditions of temperature and time according to the experimental design. The matrix regarding experimental design conditions by the RSM-CCD method and corresponding lignin, holocellulose, and cellulose amounts is shown in Table 3.

Table 3 Design matrix and correlated values
Full size table

The following second-order polynomial equations model the lignin and cellulose amount (Y1: lignin percentage, Y2: cellulose percentage) regarding the influencing parameters variation (the values are coded- insignificant terms were eliminated).

$$Y1=-0.6686{X2}^{2}-0.2602{X3}^{2}-1.27X1X2-0.9398X1X3-0.7761X1-0.8263X2- 1.12X3+25.82$$
(6)
$$Y2=+1.04X1X2+0.9721X1X3++1.34X1+1.62X2+0.7217X3+42.43$$
(7)

In these equations as mentioned in Table 1., X1 and X2, and X3 are respectively choline chloride: lactic acid molar ratio, process temperature, and process time in coded values. The coefficient of X1, X2, and X3 show a variable direct effect on the response. As well as a direct effect of parameters on the response they can interact with each other and strengthen the parameter’s effect on the response which can be shown by X1X2, X1X3, and X2X3 coefficients. In the coded equation the coefficient of each variable shows the importance of that parameter on the response as the dependent variable.

Established models variance analyses in presented in Table 4. As the R2 results show the presented models have a good ability to correlate the response and involving parameters (Fig. 1). The p-values lower than 0.0001 for both models (less than 0.05) and a high amount of F-values (43.81and 67.95 for lignin and cellulose percentage) show this fact. Also, the lack of fit p-values which were 0.6923 and 0.3349 respectively, validate these sentences.

Table 4 ANOVA for cellulose isolation
Full size table
Fig. 1
figure 1

Predicted versus actual plot for a) lignin percentage and b) cellulose percentage in the structure after treatment with ESs

Full size image

The adequate precision for lignin and cellulose amount in the structure were 26.9431 and 30.7995 which are more than 4 (signal-to-noise ratio) show that models can navigate the design space.

The coefficient of variation (CV) of less than 10%, shows the model’s reproducibility. The CV value of 1.74% and 1.25% for lignin and cellulose amounts demonstrate good precision of experiments.

3.1.2 Variable effect on perturbation, response surface, and counter plots

The perturbation plot regarding the lignin and cellulose percentage in the ESTCPs is presented in Fig. 2. As this figure shows all three influencing factors, including lactic acid: choline chloride ratio, process time, and temperature have direct effects on lignin elimination and cellulose isolation percentage in the pretreated sample, so that by lignin amount decreases in the structure cellulose amount increase.

Fig. 2
figure 2

The perturbation plot regarding the a) lignin percentage b) cellulose percentage in the structure after treatment with ES solvents (A: choline chloride: lactic acid molar ratio, B: process temp (°C), C: time (h))

Full size image

As previously mentioned, by using contour plots the relationship between different parameters (including lactic acid: choline chloride ratio, process time, and temperature) on lignin and cellulose percentage after treatment with ES can be illustrated (as shown in Fig. 3). Figure 3a and 3c respectively demonstrates the contour plots of lignin and cellulose percentage regarding the different amounts of ES ratios and temperature at the constant time (3 h). According to the saddle shapes of the contour plots, there is effective interaction between temperature and ES ratio. It is clear from the figure that the lignin and cellulose percentage amount respectively decreases and increases by increasing the ES ratio or temperature. The lowest lignin percentage and highest cellulose amount in the structure are achieved when the amount of ES ratio is about 7 to 11 and the process temperature is between the 901–20 °C. It was suggested that by increasing the choline chloride lactic acid molar ratio and temperature increase the viscosity decreased and the ability of ES for delignification enhances. It should be mentioned higher temperatures cause ES destruction and decrease its ability in the process. Also, it was reported in the more than 1:12 choline chloride lactic acid ratio the acidic property of ES solvent is dominant and can change the structure. For this purpose, most of the investigation has been conducted in the range of 1:1 to 1:12 [24, 52, 60, 61].

Fig. 3
figure 3

Contour plot image of the effect of different parameters on lignin and cellulose percentage after treatment with ESs

Full size image

Figure 3b and d shows the lignin and cellulose percentage as a function of the ES ratio and the processing time at the constant temperature (100 °C). According to the contour plot’s saddle shapes, the interaction between variables is significant. As it can be understood from this figure, the delignification and cellulose isolation percentage increases by ES ratio or process time values. The most delignification and cellulose isolation percentages were achieved when the ES ratio and the temperature were at. 7–11 ratio and 3–5 h. This result was validated by other investigations that by increasing the time of treatment the delignification amount increases [61].

It should be mentioned the lignin amount in the structure is consistent with cellulose percentage (as the lignin amount during treatment by ES in the structure decreases cellulose percentage increases which are due to the high ability of ES in lignin solubilization) [62].

3.1.3 Optimization and validation of responses

To achieve the maximum delignification and obtain the most cellulose extraction amount, the Design Expert software optimization tool was applied. This procedure is used for lignin removal and cellulose isolation maximization (between zero and one). The optimization condition by desirability method and best conditions for pretreatment are represented in Table 5. Under predicted conditions, the verification experiment was repeated and experimental responses were close to the estimated amount.

Table 5 Optimization condition and optimum values of ES treatment for achieving maximum delignification and cellulose isolation
Full size table

The optimum condition which suggested by the design expert numerical optimization tool applied for the post-treatment process. The amount of lignin and cellulose in different samples is presented in Fig. 4.

Fig. 4
figure 4

The amount of lignin and cellulose in different samples with the standard deviation ± 0.2

Full size image

3.2 Experimental results and characterization

3.2.1 FTIR analysis

Typical FT-IR spectra related to the functional group of EFNCPs, ESTCPs, CPC, and CNF are presented in Fig. 5.

Fig. 5
figure 5

FTIR Analysis of different forms

Full size image

This figure shows that all spectra have similarities that are related to the base’s similar chemical compositions.

As it is clear in a structure of NCP, EPC, and CPC a broad absorption band at 3400–3500 cm−1 related to the tensile vibrations of -OH groups in the cellulose structure is observed. Also, it was mentioned there is a negligible effect on hydrogen bonding in the cellulose after ES treatment [63,64,65].

Regarding the obtained results, in a structure of EFNCP, ESTCPs, CPC, and CNF in a range of 2860 cm−1, and 2940 cm−1, an absorption peak was observed, which shows tensile vibrations aliphatic saturated C-H associated with methylene groups in the structure of the desired materials [62, 66].

In the spectrum of EFNCP, an absorption peak around 1740 cm−1 is related to the C = O group vibration in the acetyl or ureic ester group of hemicellulose or steric bonds in the lignin structure. In the process of ES treatment and after the cellulose complete isolation process in the ESTCP, CPC, and CNF structures, this peak strength decreased and was eliminated which shows the lignin and hemicellulose removal. Also, the FTIR vibration at near 1515 cm−1 indicates the lignin aromatic ring skeletal stretch. As it is clear during the ES treatment and isolation process the intensity of this peak decrees [67,68,69,70].

In the FTIR spectrum of EFNCP, ESTCP, and CPC, an absorption peak around 1630 cm−1 was observed which is related to the hydroxyl group vibration and water absorption [71].

Several peaks were observed in the EFNCP, ESTCP, CPC, and CNF structures in the 1210–1490 cm−1 and 1055 cm−1 regions. These are associated with symmetric and asymmetric vibrations of carboxylic acids, CH, and CO in the cellulose and hemicellulose structure [72]. Also, the peak around 1030 cm−1 is related to the vibration of C–O–C in the pyranose ring in the cellulose structure. More intense vibrations at these regions show increasing the cellulose amount during the treatment and isolation process [62]. Also, the increase of band at 890 cm−1 in the ESTCP, CPC, and CNF in comparison to EFNCPs is related to the glycosidic linkages of the glucose ring of cellulose in the cellulose typical structure which shows increasing the amount of cellulose in the structure along the isolation process [73].

Absorption peaks indicating the bending vibrations of C–OH groups in the structure have been reported for EFNCP, ESTCP, and CPC structures in the 650 cm−1 region. As reported in some literature, the strength of this peak is related to prepared cellulose nanostructures [74, 75].

3.3 XRD analysis

The XRD pattern of EFNCP, ESTCP, CPC, and CNF samples is demonstrated in Fig. 6. With regards to the existing results, the primary peaks appear at about 16° and 22.5° respectively, which shows the cellulose phase in the structure.

Fig. 6
figure 6

XRD Analysis of EFNCP, ESTCP, CPC, and CNF

Full size image

As it is clear, the XRD result shows that the EFNCP has different impurities that are obvious in the XRD patterns by small peaks. In the exaction and nano-fibrillation process, the small peaks were eliminated, and the crystallinity increased [76, 77].

As it is clear from this figure the diffraction patterns of all materials showed characteristic peaks of cellulose at the same range which shows that the crystal morphology of cellulose did not change during the treatment but that the crystallinity was increased during the separation and nanofabrication processes [78].

The cellulose Crystallinity Index (CrI) was calculated using the 4 equation and the CrI results are reported in Table 6. It is obvious from these results in the extraction process by structure cleavage and impurities like lignin elimination the fibrillation increases and crystallinity enhances [22].

Table 6 Crystallinity Index
Full size table

3.3.1 TG and DTG analysis

Figure 7a indicates the TG curves of EFNCP, ESTCP, CPC, and CNF. All TG curves show a small preliminary drop between 50 and 150 °C, which corresponds to a mass loss of about 5% of absorbed moisture. The decomposition of the original untreated plant fibers occurs in several stages, indicating the presence of different components that decompose at different temperatures. Due to the low decomposition temperature of hemicelluloses and lignin, EFNCPs begin to degrade from about 200 °C, and the glycosidic bonds of cellulose are broken. A small shoulder appeared in the EFNCP DTG curve at 250 °C, which corresponds to the hemicellulose decomposition, which is removed during the isolation process. The peaks on the DTG curve (Fig. 7b) correspond to the temperature of the maximum decomposition rate. The wide peak in the 200–400 °C region is related to cellulose degradation. As mentioned in previous articles, the peaks in EFNCPs around 450–650 °C are related to the lignin remaining in the structure. During isolation by the removal of lignin and hemicellulose, the narrowness of the peaks increased. As it is clear during isolation by ES treatment, impurity-related peaks are removed, which is consistent with FTIR and XRD analysis. Furthermore, as it is clear in the nano-fibrillation process by ES, the maximum thermal decomposition temperature of cellulose is only 12 °C lower than that of EFNCPs (and 6 °C lower than isolated CPC), which indicates the structure of cellulose fibers is almost unchanged. This suggests that the CNF prepared by this procedure is highly heat resistant and can be used for specific subjects [79,80,81,82,83].

Fig. 7
figure 7

a)TG and b) DTG curves of EFNCP, ESTCPs, CPC, and CNF

Full size image

3.3.2 Fe-SEM and TEM

Figure 8 represents respectively the Fe-SEM image of EFNCP, ESTCP, CPC, CNF, and TEM and the size distribution of CNFs. AS Fig. 8a shows, the surface layers have some impurities (lignin, oil, etc.) and have an undisrupted, rigid, homogenous surface. Figure 8b demonstrates the ESEPCs structure after the ESs application process and lignin removal. As it is clear in the primary treatment process with ES the surface homogeneity decreases and fiber bundles appeared which is due to the lignin elimination. Figure 8c shows the CPC fibrils after the post-treatment process, including delignification, and removal of hemicellulose and other residuals. As it is shown after the post-treatment process the cellulose fibers become clear and impurities are highly eliminated.

As can be assumed from Fig. 8b and c in the process of EFNCP pretreatment with ES and post-treatment process for lignin and hemicellulose removal, surface roughness increased. The increase in surface roughness is related to the non-cellulosic layer removal (hemicelluloses and lignin) during each treatment process. The increase in surface roughness was produced by the pretreatment process, since the hemicelluloses coat the cellulose fibrils, whereas lignin fills the empty spaces [84, 85].

Fig. 8
figure 8

Fe-SEM and TEM images of different forms (a: EFNCP, b: ESTCPs and c: CPC, d: CNF, e: TEM image of CNF and f: histogram of size distribution frequency%)

Full size image

The prepared CNFs structures by ES solvent treatment are demonstrated in Fig. 8d.

It can be understood that treatment and nano-fibrillation by remaining impurities and amorphous region elimination alongside structure cleavage cause cellulose fibrils’ size and diameters decrease and their crystallinity and uniformity enhance.

The morphological structure of the prepared CNFs was studied by transmission electron microscope (Fig. 8e).

For transmission electron microscopy (TEM), the frieze-dried cellulose nanofibers were diluted by ultrapure deionized water and dispersed by ultra-sonication (30 min at 20 Hz). A small droplet of 0.005% (w/v) cellulose nanocrystal suspension was added to the carbon-coated electron microscopy grid and excess solvent was eliminated by drying at room temperature.

As TEM results show after ES treatment of CPC, the morphology and surface obviously changed. As it is shown from the TEM and size distribution histogram (Fig. 8f) the prepared CNFs with smooth surfaces have short rod shapes and high aspect ratio with diameters of 20–40 nm, and lengths of 400–800 nm. The changes in fiber size are related to the glycosidic linkage cleavage of prepared EFNCPs during the cellulose isolation and nano-fibrillation process in the presence of ES solvents.

4 Conclusion

Concerning all of the subjects mentioned in this investigation, this paper aims to apply choline chloride- lactic acid-based ESs for delignification and cellulose isolation optimization from extractive-free natural cotton pods (EFNCPs) to achieve the highest cellulose extraction and purity and consumption of ES-treated cotton pods (ESTCPs) for nano-fibrillation. The relationships between various parameters affecting delignification and cellulose separation are illustrated using contour plots. The optimum condition for cellulose isolation reaches 1:10.01 choline chloride lactic ratio, 118 °C temperature, and 4.56 h. Under optimal conditions, the experimental separation test conducted and experimental cellulose extractions performed were close to the predicted values that validated the optimization and modeling process. The primary treated biomass at the optimum condition was secondary treated in mild conditions with H2O2 and NaOH solution and high purity cellulose structure with 95% pureness obtained. The extracted cellulose was used for cellulose nano-fibrillation, and choline chloride-lactic acid-based ES treatment was applied for this purpose. In the process of treatment and nono-fibrillation, all of the samples were characterized by FTIR, XRD, TG, DTG, and Fe-SEM methods. The results show in the treatment process as impurities related to the lignin and hemicellulose presence eliminate and the surface morphology becomes smoother and the crystallinity index enhances. TG analyses alongside DTG analyses show that there is little difference in the maximum thermal decomposition temperature of cellulose and source biomass. TEM and Fe-SEM analyses which show the high length-to-width ratio of prepared nanofibers is in contestant with TG analyses show the mechanical and thermal strengths of isolated nanofibers is almost unchanged which is due to the mild isolation and nano- fibrillation process. The result of this investigation can be used for cellulose isolation optimization with special chemical and mechanical properties in a green and environmentally friendly method.