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Article

Comparative Study of Mechanical and Microstructural Properties of Biocemented Sandy Soils Enhanced with Biopolymer: Evaluation of Mixing and Injection Treatment Methods

by
Mutlu Şimşek
1,*,
Semet Çelik
2 and
Harun Akoğuz
1
1
Faculty of Engineering and Architecture, Erzincan Binali Yildirim University, Erzincan 24002, Türkiye
2
Faculty of Engineering, Atatürk University, Erzurum 25240, Türkiye
*
Author to whom correspondence should be addressed.
Appl. Sci. 2025, 15(14), 8090; https://doi.org/10.3390/app15148090
Submission received: 8 June 2025 / Revised: 10 July 2025 / Accepted: 16 July 2025 / Published: 21 July 2025
(This article belongs to the Section Civil Engineering)
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Abstract

Soil improvement is one of the fundamental practices in civil engineering, with a long-standing history. In today’s context, the rapidly increasing demand for construction driven by urbanization has further emphasized the necessity and significance of soil stabilization techniques. This study aims to determine the optimum parameters for improving sandy soils by incorporating sodium alginate (SA) as a biopolymer additive into the microbial calcium carbonate precipitation (MICP) process. Sand types S1, S2, and S3, each with distinct particle size distributions, were selected, and the specimens were prepared at medium relative density. Three distinct approaches, MICP, SA, and MICP + SA, were tested for comparison. Additionally, two different improvement methods, injection and mixing, were applied to investigate their effects on the geotechnical properties of the soils. In this context, hydraulic conductivity, unconfined compressive strength (UCS), and calcite content tests, as well as scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDS) analyses, were performed to assess the changes in soil behavior. SA contributed positively to the overall efficiency of the MICP process. The study highlights SA-assisted MICP as an alternative that enhances the microstructural integrity of treated soils and responds to the environmental limitations of conventional methods through sustainable innovation.

1. Introduction

In the past two decades, researchers have extensively studied biological techniques as a replacement for traditional methods [1]. As a biological technique, MICP technology offers advantages over traditional methods like cement, lime, and sintered bricks by reducing resource consumption [2]. It is also an efficient and eco-friendly approach for removing contaminants, including heavy metals, from polluted environments [3]. This technology has been used effectively in enhancing the properties of soil and concrete [4,5,6] and numerous researchers in the past have concentrated on enhancing the parameters of soils, such as erosion, strength, cohesion, shear strength, crack healing, seepage, hydraulic conductivity, and permeability [4,7,8,9,10,11,12,13], making it a viable option for various geotechnical engineering applications. While MICP has many advantages in soil improvement, implementing it at the field scale is still challenging because of the inhomogeneous distribution of calcite precipitation [14].
A variety of studies, both experimental and theoretical, have been conducted to gain insight into this effect. For example, Xiao et al. [15] examined the homogeneity and mechanical properties of sand using a one-phase temperature-controlled MICP method, demonstrating that specimens treated with this approach exhibited increased CaCO3 content with a nearly homogeneous distribution along the specimen’s height, leading to a significant enhancement in strength and the development of noticeable dilatancy, attributed to the effective bonding network created by the homogeneous CaCO3 precipitation. They also noted that a non-uniform MICP-treated specimen would form with the majority of CaCO3 precipitation near the grouting inlet and proposed improving homogeneity by inhibiting bacterial activity and uniformly distributing the bacteria and cementation solution before the MICP reaction. Xiao et al. [16] investigated microfluidic chips to examine the pore-scale behavior of MICP. The results showed that calcium carbonate first forms on the bacterial surfaces before gradually spreading more evenly across the microchip. This approach provides valuable insights into distribution patterns and can aid in refining conditions to achieve more uniform calcite precipitation. Liu and Fernø [17] focused on microfluidic micromodel design, calcite functionalization, and the investigation of biogeochemical interactions in porous media. In another study, Liu et al. [14] developed an immersion method for MICP treatment, where soil samples are submerged in cementation media, allowing the media to diffuse freely into the soil, resulting in a uniform distribution of precipitated CaCO3 and suggesting its potential for more consistent soil improvement. Kirkland et al. [18] utilized X-ray μ-CT imaging to assess the efficacy of MICP in repairing cement defects, enabling the visualization of changes over time and across different spatial regions. Their findings revealed that increasing the injection flow rate and providing more frequent nutrient solution stimulation to the bacterial community resulted in a larger reduction in void fraction and a more uniform MICP seal.
To gain insight into a quantitative and effective analysis for the MICP process, numerical models have also been investigated, in addition to experimental studies [19,20]. For instance, Martinez et al. [21] developed a transport model to predict the MICP treatment of sand in one-dimension. The results showed a strong correlation between the simulated outcomes and experimental data, effectively representing the magnitudes and trends of various MICP treatment scenarios in half-meter, one-dimensional flow columns. It is understood that ensuring a consistent distribution of calcite in the MICP process is vital for its successful application in various fields, and these experimental and numerical methods contribute to better uniformity and mechanical properties of the treated soil, making MICP a more effective option for soil enhancement. Nonetheless, while field trials have demonstrated the viability of various bio-based methods such as bio-gelation, bio-clogging, biogas desaturation, and bio-cementation, challenges still exist, such as inhomogeneity, and an effective injection strategy is crucial to ensuring the substrates are delivered properly, promoting a uniform distribution of precipitated calcium carbonate [22].
Hydrophobic gel-like biopolymers such as polyhydroxybutyrate, xanthan, chitosan, and polyglutamic acid are also used for soil improvement [23]. In recent years, studies have been conducted in the literature using biopolymer additives to increase the effectiveness of MICP. For instance, Zhang et al. [24] examined the role of a chitosan biopolymer in influencing the MICP process and observed that it facilitates nucleation and modulates the growth of MnCO3. This is achieved by providing nucleation sites for mineral formation and mitigating the toxic effects of metal ions. Their findings suggest that chitosan holds promise for enhancing the efficiency and sustainability of the MICP process. In another study, Hang et al. [25] investigated the impact of adding xanthan gum biopolymer to enhance the reinforcement effect of MICP in sandy soils. They found that incorporating 2 g/L xanthan gum into the bio-cementation process significantly enhanced the strength and structural integrity of sandy soil samples more than those treated with either MICP or xanthan gum alone. In addition, naturally occurring organic substances, such as polysaccharides, may play a role in controlling the polymorphism and morphology of MICP [26]. SA, a biosynthetic polysaccharide, can simulate the impact of polysaccharides on CaCO3 formation, offering several advantages in this process: The calcium alginate gel provides a habitat for microorganisms, gradually releasing calcium for in situ precipitation, while alginate molecules affect the CaCO3 morphology through gel collapse, with the negative charges on the alginate molecules significantly affecting the polymorphism and morphology of CaCO3, as observed with varying concentrations of SA [26]. When considering the unique contribution of sodium alginate (SA) to MICP efficiency in comparison to other biopolymers, SA stands out due to its strong gel-forming ability, effective calcium ion binding capacity, and superior performance in reducing soil permeability. According to the literature, SA has been shown to reduce permeability by approximately 20 times more than biopolymers such as guar gum [27]. Additionally, SA supports the sustained activity of MICP by supplying oxygen and nutrients to bacteria, enhances the injectability of the cementing solution, and contributes to more homogeneous calcite precipitation [28]. For these reasons, the SA + MICP combination fills an important gap in the literature and offers an original solution for simultaneously improving both soil strength and hydraulic conductivity.
It is understood that studies in the literature highlight the importance of ensuring homogeneous distribution, and while experimental and analytical approaches have been proposed, they remain limited in number; additionally, biopolymer additives have proven to be a highly effective factor in CaCO3 formation. In addition, the method of applying the bacteria-cementitious solution into the soil affects the homogeneity of the treated solution, which in turn determines the success of the method, with three possible introduction techniques: injection, surface percolation, and pre-mixing [29]. While the importance of homogeneity is well-established, most studies have primarily concentrated on either the injection or mixing method, with the immersion method potentially presenting limitations for field applications. Moreover, the impact of SA additive on the homogeneous distribution of MICP has not been thoroughly investigated or fully understood, although SA gel’s remarkable microorganism immobilization capability is demonstrated by the uniform distribution observed in microscope images [26].
Hence, this study aimed to examine the impact of SA as a natural biopolymer on the MICP process under varying SA concentrations (0.5%, 1.0%, 1.5%, 2.0%, 2.5%), particle sizes of soils (three particle sizes), and treatment procedures (injection and mixing), with a specific focus on achieving a uniform distribution of CaCO3. The effect of SA on MICP was assessed by analyzing the calcium carbonate content, unconfined compressive strength, and hydraulic conductivity. Furthermore, SEM and EDS techniques were used to analyze and compare the microstructure and mineral composition of the soils after and before treatment, offering insights into how homogeneity affects the MICP bio-cemented samples with SA additive. This insight would enhance the understanding of homogeneity and the role of SA in the MICP technique.

2. Materials and Methods

2.1. Materials

Soil

To examine the effect of various grain sizes, sandy soils with three different grain diameter ranges were used in the experiments. The soil samples used in our study generally fall within the size range of 4.75 mm (passing the No. 4 sieve) to 0.075 mm (between No. 4 and No. 200 sieves) in the grain size distribution (GSD) curve. This range represents the sand fraction. The gravel fraction is defined as particles larger than 4.75 mm, while the fine fraction (fines) refers to particles smaller than 0.075 mm. These boundaries have been taken into account when evaluating the GSD chart. The sandy soils are classified as poorly graded (SP) sandy soil based on the Unified Soil Classification System (USCS). The grading curves and physical characteristics of the soils are illustrated in Figure 1, Table 1.
The visual appearances and SEM images of the soil types S1, S2, and S3 used in this study are presented in Figure 2.

2.2. Experimental Details

Within the scope of the study, two different soil improvement methods—namely injection and mixing—were employed. These improvement techniques were applied to soil specimens composed of three different particle size distributions (S1, S2, and S3). The maximum (emax) and minimum (emin) void ratios of the S1, S2, and S3 soils were determined through laboratory tests. Based on these calculated values, the required amount of soil to be placed into the molds was determined, and sample preparation was carried out accordingly. The S1, S2, and S3 silica sand specimens were placed into PVC molds with a diameter of 38 mm and a height of 76 mm, compacted to achieve an approximate relative density of about 50%.

2.2.1. Bacterium, Liquid Medium and Cementation Solution

The bacteria used in the experiment was Viridibacillus arenosi K64 (GenBank Acceptance No.: KR873397) procured from Ataturk University, Faculty of Science, Department of Biology. It has been confirmed in vitro that bacteria participate in calcite formation [1,2]. The liquid medium was prepared with specific amounts of urea (20 g/L), MnSO4·H2O (12 mg/L), tryptic soy broth ((TSB) (Laboratorios Conda S.A., Madrid, Spain) was used for the cultivation of microorganisms.) (30 g/L), and NiCl2·6H2O (24 mg/L) and dissolved in a measured volume of distilled water [3]. All components of the mixture, except for urea, were sterilized using an autoclave at 121 °C for 45 min. Due to the thermal decomposition of urea during autoclaving, it was sterilized via filtration and subsequently incorporated into the mixture under aseptic conditions. The prepared liquid medium was incubated on a shaker at 120 rpm and a constant temperature of 25 °C for 48 h. The cementation solution (CS) was prepared using CaCl2 (Calcium chloride dihydrate (TEKİM, Istanbul, Türkiye)) and urea (TEKİM, Türkiye) with the aid of a magnetic stirrer. The CS was used without sterilization, as maintaining sterile conditions was not feasible in field applications. In this study, the culture medium was prepared under sterile conditions to ensure the effective growth of bacterial cells. However, the cementation solution was not sterilized, as the sterilization of such solutions may present practical challenges, especially when considering real field applications. Moreover, the literature has shown that effective biocementation can be achieved under low-sterility conditions. For example, one study reported that impure bacterial cultures were able to successfully strengthen the sand without the need to isolate a pure strain. The researchers also demonstrated that biocementation could be effectively achieved under low-sterility conditions. This finding is particularly important for large-scale applications, as it highlights that bacterial cells can be cultivated on-site, eliminating the need for importing pure cultures from external sources [30]. Based on these findings, the use of a non-sterile cementation solution was considered appropriate for the purpose of this research (Figure 3). The CS consists of 0.5 mol/L urea and 0.5 mol/L CaCl2. In the literature, it is reported that using equimolar concentrations of urea and CaCl2 enhances the cementation effect of the MICP process [31]. Several studies have shown that a 1:1 ratio of these two compounds leads to more effective calcite precipitation and improved soil stabilization [32]. Accordingly, the cementation solution used in this study was prepared with 0.50 mol/L of urea and 0.50 mol/L of CaCl2 [33]. In this study, soil improvement was carried out using both injection and mixing methods by applying 120 mL of culture medium (CM) or 120 mL of SA solution (SS), along with 350 mL of CS. In the specimens treated only with sodium alginate, SA was dissolved in distilled water. However, in the specimens treated with both sodium alginate and MICP, SA was intentionally dissolved in the culture medium to ensure direct interaction with the bacterial solution and to achieve a homogeneous distribution within the same nutrient environment. This approach was adopted to support the simultaneous and effective occurrence of biopolymer gelation and microbially induced calcite precipitation.

2.2.2. Sodium Alginate Solution

Sodium alginate (ISOLAB Laborgeräte GmbH, Eschau, Germany) was used in the experimental procedure. SA was obtained from Isolab chemicals (CAS-No.: 9005-38-3), and the properties of SA are given in Table 2. The CM was converted into a gel by adding SA at concentrations (w/v) of 0.5%, 1%, 1.5%, 2%, and 2.5%, as the varying levels of SA can significantly influence the polymorphism and morphology of CaCO3 [4]. Stirring was maintained until the SA was fully dissolved in the CM. This homogeneous mixture was injected into the soil using the injection technique. However, prior to application with the mixing method, the mixture (SA + CM) was mixed with varying volumes of CS (120 and 350 mL) before being applied to the soil. The use of SA in MICP may offer the following benefits: Firstly, in the injection method, the CS contains calcium ions, and the presence of divalent ions (especially calcium) facilitates rapid gel formation by exchanging monovalent sodium ions, quickly converting the low-viscosity solution into a gel structure. Secondly, in the mixing method, the presence of urea in the mixture delayed gel formation, enhancing the workability of the soil mix and simplifying the compaction process [5].

2.2.3. Preparation of Injection Method

In the soil improvement studies conducted using the injection method, extensive preparations were carried out in the laboratory to ensure the proper execution of the experimental program. Initially, the required quantities of soil for each type—S1, S2, and S3—were calculated for use in molds. Subsequently, appropriate molds were prepared, and the soil specimens were produced accordingly. These specimens, with dimensions of 38 mm in diameter and 76 mm in height, were compacted to achieve a relative density of about 50%. To ensure homogeneous distribution within the mold, Scotch-Brite filters were placed at both the bottom and top of the soil samples [34]. For the injection-based improvement, soil specimens were prepared using only MICP, only SA, and a combined SA-MICP approach, applied to the S1, S2, and S3 sand types. The nutrient medium and bacterial solution required for the microbial improvement process were prepared in advance. Additionally, SS was formulated for use as a biopolymer additive. Following this, the CS was prepared, completing the set of chemical components to be integrated into the injection system. In our experimental setup, the injection was performed from the top of the specimen by allowing the solution to percolate downward through the soil column. A peristaltic pump (Bimetron, Buca, Türkiye) was used to control the injection rate and ensure a uniform flow during the percolation process. A peristaltic pump, which plays a critical role in flow regulation, was used to calibrate the injection flow rate. After the setup was assembled, the injection process was carried out in accordance with the predetermined flow rate parameter. Upon completion of the injection procedure, the specimens were placed in an oven for the curing process. Visual representations of the prepared molds and soil specimens are presented in Figure 3a, while the injection test setup is shown in Figure 3b (See Figure 3).

2.2.4. Preparation of Mixing Method

For the mixing method, a liquid mixture consisting of MICP, SA and SA-MICP solutions was prepared by blending with the CS. This mixture was filtered and combined with silica sands at different pellet contents (5%, 10%, and 15%) to prepare the test specimens. In the mixing method, a “pellet” refers to the solid product obtained by filtering the sodium alginate–cementation solution mixture or the sodium alginate plus MICP–cementation solution mixture. For each SA/CM (%) ratio, the produced pellets were added to the soil specimens at 5%, 10%, and 15% by soil weight, and the mixing procedure was carried out accordingly. The samples were shaped in molds measuring 76 × 38 mm. In the mixing method, the amount of silica sand (S1, S2, and S3) to be placed in the molds was calculated similarly to the injection method. The filtered liquid mixture, composed of MICP, SA, and their combination (SA-MICP), was mixed with the cementation solution. After filtration, this mixture was combined with silica sands at three pellet content ratios (5%, 10%, and 15%) to form the test samples. In the mixture design process, the dry unit weights corresponding to approximately 50% relative density were determined as 15.01 kN/m3, 14.91 kN/m3, and 14.81 kN/m3 for the S1, S2, and S3 soils, respectively. Cylindrical specimens (38 mm in diameter and 76 mm in height) were prepared by incorporating pellet material at 5%, 10%, and 15% of the dry unit weight, followed by a compaction procedure. The determined amounts of sand and mixture were blended, then divided into five layers. Each layer was placed into the mold and compacted uniformly using a 10 mm diameter rod. Upon completion of the application, the specimens were placed in an oven for the curing process. A visual representation of the mixing test setup is provided in Figure 3c. The soil improvement methods applied to the S1, S2, and S3 silica sand specimens, along with the quantities of solutions used in each method, SA ratios, injection flow rates, and details of the tests conducted on the soils, are presented in Table 3.

2.3. Methods

In this study to determine the mechanical strength properties of the soil specimens, the UCS test was performed in accordance with the ASTM D2166 [35]. To assess the hydraulic conductivity characteristics, the falling head test was carried out based on ASTM D5084 [36]. In addition, calcite content analysis was conducted to determine the amount of calcium carbonate formed as a result of biocementation, and SEM analyses were performed to examine the morphological characteristics of the microbial precipitates. Furthermore, EDS analysis was applied to identify the chemical composition of the formed minerals. In this context, EDS analysis was utilized as a complementary characterization method to support the effectiveness of MICP.

2.3.1. Hydraulic Conductivity Test

The falling head test was conducted to measure the hydraulic conductivity of the soil specimens after improvement (Figure 4). This method is particularly suitable for fine-grained soils with low hydraulic conductivity and is based on evaluating the rate at which water flows through the soil by observing the decreasing water level over time. A peristaltic pump device was used to perform the falling head test. In this setup, the soil specimen within the mold was sealed to ensure it was watertight and then fully saturated. A transparent vertical tube (1 m in length) was filled with water to the point of overflowing to avoid the presence of air bubbles. Once the tube was filled, the peristaltic pump was stopped and the valve was closed. Then, the valve was reopened, and the change in water level over time was recorded. Using this setup, the change in hydraulic head over time across the soil sample was measured, and the coefficient of hydraulic conductivity (k) was calculated based on Darcy’s law. If the water height is h1 at time t1 and h2 at time t2, then the hydraulic conductivity coefficient is calculated as follows:
k = a . L A . t 2 − t 1 ln h 1 h 2
In this equation, k represents the hydraulic conductivity coefficient (cm/s); a is the cross-sectional area of the transparent tube (cm2); L is the height of the soil specimen (cm); A is the cross-sectional area of the specimen (cm2); h1 is the initial water head at time t1 (cm); and h2 is the final water head at time t2 (cm).

2.3.2. Unconfined Compressive Strength (UCS) Test

UCS tests were performed on the soil specimens following the completion of the oven-curing period, in accordance with the ASTM D2166. These tests were conducted to evaluate the effect of the soil improvement applications on the mechanical strength of the soils. In this context, the improved soil specimens were prepared in cylindrical form with a diameter of 38 mm and a height of 76 mm. During the test, load-deformation data were continuously recorded, and the maximum strength value was determined for each specimen. These strength tests were carried out at a loading rate of 1.0 mm/min. The resulting UCS values were used to compare the effects of different additive contents (SA, SA + MICP, and MICP) and different improvement methods on the strength of the soils (Figure 5).

2.3.3. Determination of Calcite Percentage

The weight loss method was used to determine the calcium carbonate content. Weight loss occurred as a result of washing the soil specimens with HCl. Although various methods exist for measuring calcium carbonate percentage, this method was preferred due to its ease of application and compatibility with the available laboratory equipment. In this method, approximately 5–6 g of the soil sample, taken from a specimen previously subjected to UCS testing, was mixed with 20 mL of 1 M HCl acid. The dissolved particles were rinsed using distilled water and filter paper for about 10 min. This washing process ensures the complete removal of all soluble calcium components from the sample. Subsequently, the remaining soil particles were oven-dried again and weighed (Figure 6). The difference between the initial weight of the sample (A) and the weight after washing and drying (B) indicates the amount of calcium carbonate. Finally, the calcium carbonate percentage is calculated using the following formula:
Calcium Carbonate (CaCO3, %) = [(A − B)/A] × 100

2.3.4. Scanning Electron Microscopy (SEM) and Energy Dispersive X-Ray Spectroscopy (EDS) Analysis

The analysis of the existing structures before treatment and the structures formed after treatment was conducted using the Quanta 250 FEG model SEM (Thermo Fisher Scientific, Hillsboro, OR, USA). In addition to SEM, EDS analysis was used to determine the elemental composition of the soil sample. This technique is commonly employed in soil improvement studies such as MICP to verify the chemical composition of the precipitated minerals. Within the scope of the study, SEM and EDS analyses were performed on selected soil samples in line with the specified objective. These analyses were conducted on untreated S1, S2, and S3 soil samples, as well as on samples improved by the injection method (S1-SA, S2-SA, S3-SA; S1-SA + MICP, S2-SA + MICP, S3-SA + MICP) and by the mixing method (S1-SA, S2-SA, S3-SA; S1-SA + MICP, S2-SA + MICP, S3-SA + MICP). The analyzed samples were selected from among those that demonstrated the highest performance based on UCS results.

3. Results and Discussion

This study, which was conducted by taking various parameters into account, resulted in the creation of 186 different designs: injection (36 specimens) and mixing (150 specimens). The experimental program was carried out on specimens prepared with three different types of additives: only SA, a combination of SA and MICP, and only MICP. However, during the injection-based improvement process, the use of standard peristaltic pumps and associated equipment limited the application of SA to a maximum of 1.0% for S1 soil and 2.0% for S2 soil; higher concentrations could not be successfully injected. To evaluate the effects of these soil improvement techniques on the geotechnical properties of the soils, UCS tests, falling head hydraulic conductivity tests, and calcite content determinations were conducted. SEM was performed to examine the morphological characteristics of the precipitates, and EDS analysis was conducted to evaluate their chemical composition.

3.1. Hydraulic Conductivity Results

In the experimental studies conducted using the injection method, three different soil types (S1, S2, and S3) were improved by applying five different additive ratios—0%, 0.5%, 1.0%, 1.5%, 2.0%, and 2.5%—of SA, in combination with MICP. The effects of different SA ratios and SA + MICP combinations on the hydraulic conductivity (cm/s) of the S1, S2, and S3 soil specimens improved by the injection method are presented in Figure 7. The results obtained after treatment generally indicate that increasing the SA content significantly reduces hydraulic conductivity. Except for the S3 soil, the hydraulic conductivity values further decreased in the combinations where SA was applied together with MICP. The lowest hydraulic conductivity values were recorded at the highest SA ratios. This suggests that the viscous structure of SA and the calcite precipitation induced by MICP play a complementary role in reducing soil hydraulic conductivity. The obtained data showed that the addition of SA was effective in improving low hydraulic conductivity across all soil groups. As the SA content increased, the gel-like structure narrowed the water flow paths, resulting in a decrease in the hydraulic conductivity coefficient, with this effect becoming particularly evident up to a 1% SA ratio. However, further increases in SA content, in some cases, adversely affected hydraulic conductivity reduction due to the non-uniform distribution of the additive. The addition of MICP to the SA treatment resulted in a further decrease in hydraulic conductivity. The biologically induced calcite precipitates formed by MICP more effectively filled the pores within the soil, significantly restricting water flow. However, in some mixtures, the effect of MICP appeared to be limited, which was likely due to factors such as insufficient microbial activity, pH imbalance, or inadequate precipitation time.
In the experimental studies conducted using the mixing method, three different soil types (S1, S2, and S3) were improved using a combination of SA at 0%, 0.5%, 1.0%, 1.5%, 2.0%, and 2.5% and pellet contents of 5%, 10%, and 15%, along with MICP. Figure 8 illustrates the effects of different SA ratios and 5%, 10%, and 15% pellet contents on the hydraulic conductivity of S1, S2, and S3 soils treated using the mixing method, under both SA and SA + MICP treatments. In general, increasing SA and pellet contents tended to reduce hydraulic conductivity. However, in some cases, the SA + MICP combinations resulted in higher hydraulic conductivity values compared to only SA. This can be interpreted as SA blocking the soil pores more extensively, while MICP, despite inducing mineral precipitation, leaves voids that still allow fluid flow. In the mixtures where only SA was applied to the S1, S2, and S3 soil groups, a general increasing trend in low hydraulic conductivity was observed with increasing SA content. This effect can be attributed to the gel-like structure that SA forms upon contact with water, filling the voids in the soil and narrowing the water flow paths. In the S1 soil group, the lowest hydraulic conductivity coefficient of 1.9 × 10−5 cm/s was obtained in the mixture containing 1% SA and 10% pellet content. This value indicates that the additive provided an optimal level of low hydraulic conductivity. Although SA also improved low hydraulic conductivity in the S2 and S3 soils, the hydraulic conductivity values obtained were higher compared to those in the S1 soil. In the mixtures where MICP was applied in addition to SA, low hydraulic conductivity became more pronounced. MICP contributed positively to low hydraulic conductivity by producing calcium carbonate through microbial activity, which filled the soil porosity with mineral precipitates. When these additives were used together, the dense structure formed by the gel effect of SA became more stable due to the cementing effect of MICP, leading to further enhancement in low hydraulic conductivity. In the S2 soil, the lowest hydraulic conductivity coefficient, 1.2 × 10−5 cm/s, was measured in the mixture containing 1% SA, 15% pellet, and MICP. However, in some specimens, the addition of MICP did not result in the expected reduction in hydraulic conductivity. This was likely due to factors such as insufficient microbial activity, a short precipitation period, or the non-uniform distribution of the additives. Overall, SA + MICP combinations proved to be highly effective in reducing hydraulic conductivity. When comparing the untreated and treated soils, it is clearly observed that the treatment process causes a significant reduction in the hydraulic conductivity of the sand. While the untreated soil has a hydraulic conductivity of approximately 10−1 m/s, this value decreases to around 10−5 m/s in the treated soils.
Through different mechanisms, both injection and mixing procedures provide effective results in research aimed at reducing soil hydraulic conductivity. The injection method, particularly when combined with SA and MICP, enables the controlled delivery of additives into the soil, thereby providing more consistent and effective low hydraulic conductivity outcomes. The synergistic effect of the gel-like structure of SA and the calcite precipitation induced by MICP effectively fills soil voids, narrowing water flow paths. For instance, it has been reported that in situ-formed Ca-alginate gel reduced the hydraulic conductivity coefficient of sandy soil from 5.0 × 10−4 m/s to 2.2 × 10−9 m/s, demonstrating the potential of this method to significantly enhance low hydraulic conductivity [28]. On the other hand, the mixing method has the potential to improve low hydraulic conductivity by ensuring a more homogeneous distribution of additives within the soil. However, the success of this method strongly depends on the uniform dispersion of the additives and their application at optimal ratios. While high pellet contents can enhance soil stability, the non-uniform distribution of the additives may result in irregularities in hydraulic conductivity reduction. Moreover, the biological processes involved in MICP may not always proceed with the desired efficiency, leading in some cases to lower hydraulic conductivity values in mixtures containing only SA. It has been emphasized that high SA concentrations can increase viscosity, hinder the homogeneous distribution of additives, and adversely affect low hydraulic conductivity [37]. Although the combination of SA and MICP generally reduces hydraulic conductivity, the effectiveness of this combination is directly influenced by the distribution of the additives within the soil and the duration of precipitation [38]. In conclusion, both methods possess significant potential for enhancing soil low hydraulic conductivity. However, the sustainability of their effectiveness depends on the appropriate dosage of additives, application under method-specific conditions, soil properties, and environmental factors. In particular, for SA + MICP combinations, the careful control of microbial activity levels, precipitation time, and ion balance is critical to ensuring long-term low hydraulic conductivity performance.

3.2. Unconfined Compressive Strength (UCS) Results

Figure 9 shows the effects of different SA ratios and SA + MICP combinations on the UCS of the S1, S2, and S3 soil specimens treated using the injection method. In general, an increasing trend in UCS values was observed with a higher SA content, and in some cases, the addition of MICP further enhanced the strength. However, this effect varied depending on the soil type. Moreover, for the S1 soil, specimens treated with 0% and 0.5% SA could not retain their shape after treatment and therefore were not subjected to UCS testing. In the specimens prepared with only SA, an increase in UCS was observed with an increasing SA content, regardless of the soil particle size. The UCS values obtained for the S1, S2, and S3 soils reached maximum values of 43 kPa, 121 kPa, and 52 kPa, respectively. In the injection applications where SA and MICP were used together, a general increase in strength was observed in the soil specimens. However, in the fine-grained S1 soil, the desired structure could not be formed; the specimens disintegrated after injection and could not be brought to the testing stage. A general increasing trend in strength was observed with higher SA content, but this effect was limited in the coarse-grained S3 soil. This may be attributed to the insufficient accumulation of binding material between the larger particles.
Figure 10 presents the effects of different SA ratios and 5%, 10%, and 15% pellet contents on UCS of S1, S2, and S3 soil specimens improved using the mixing method, under both SA and SA + MICP applications. In general, SA + MICP combinations produced significantly higher strength values compared to SA alone, especially at higher pellet contents. This indicates that biological cementation is effective in enhancing mechanical strength. While only SA resulted in limited strength improvement, the addition of MICP contributed to the formation of stronger and more durable structures. As a result of SA-based stabilization processes applied using the mixing method, a clear increasing trend in UCS values was observed with higher SA and pellet contents. The maximum UCS values obtained for the S1, S2, and S3 soil types were 398 kPa, 247 kPa, and 91 kPa, respectively. For all soil types, the highest strength values were obtained in specimens treated with 2.5% SA and 15% pellet contents, indicating that this additive combination provided the most effective stabilization conditions in terms of mechanical performance. An increasing trend in soil strength was observed with higher SA and MICP ratios in the S1, S2, and S3 soils. These results indicate that the additives had the most pronounced effect in the S1 soil, while also providing a significant improvement in the S2 soil. In all soil types, the highest strength values were obtained with the combination of 2.5% SA and 15% pellet content, demonstrating that this was the most effective additive configuration in terms of mechanical performance.
In terms of UCS, both methods exhibit potential for enhancing the mechanical properties of soils; however, the mixing method has demonstrated higher strength values, particularly at elevated additive ratios. In SA + MICP combinations applied via the injection method, although low hydraulic conductivity performance improved due to the controlled delivery of additives into the soil, UCS values remained limited. This outcome is attributed to the non-uniform distribution of additives and the insufficient duration of microbial activity [39]. Conversely, the mixing method allows for the better integration of additives with the soil matrix, leading to significant increases in UCS values, especially in SA + MICP combinations containing 2.5% SA and 15% pellet. For instance, UCS values as high as 398 kPa were achieved in the S1 soil series. This indicates that biological cementation is more efficiently realized through the mixing approach [38]. Moreover, existing studies have shown that the optimum SA content typically lies within the range of 1.5–2.5%, and maximum strength enhancement is achieved within this interval [40].

3.3. Calcite Percentage Results

The effects of SA and SA + MICP combinations on the amount of calcite precipitation in S1, S2, and S3 soil specimens treated by the injection method, depending on various SA ratios, are shown in comparison in Figure 11. In general, variations in calcite precipitation were observed with changing SA content. In the S1 soil, the highest calcite content was obtained at 1% SA, while in the S2 and S3 soils, more balanced and higher precipitation values were recorded between 1.5% and 2.5% SA ratios. These results indicate that, in addition to the amount of SA, the type of soil also plays a significant role in calcite precipitation. In the literature, soil improvement studies using the injection method have primarily reported data for specimens containing only the SA + MICP additive combination. While this combination successfully promoted calcite formation across all soil types, the amount of calcite produced varied depending on the type of soil. In the S1 soil, the calcite content ranged between 1.30% and 2.72%, representing the highest values among all soil types. This suggests that the S1 soil has a structure more conducive to biological precipitation and that microbial calcite accumulation via the injection method was more effective in this soil. In the S2 soil, the calcite content ranged from 1.36% to 1.58%, indicating that the method was still effective, although the level of mineralization was more limited compared to S1. For the S3 soil, the calcite content varied between 0.31% and 1.73%, showing that the effectiveness of the injection method in promoting calcite formation was lower in this case. Differences in the pore structure, grain size distribution, and hydraulic conductivity characteristics of the soils are considered the main factors explaining the variation in calcite content.
The results of SA + MICP combinations, applied with varying SA ratios and pellet amounts, on calcite precipitation in S1, S2, and S3 soil specimens enhanced by the mixing method are compared in Figure 12. Higher calcite contents were obtained at 0.5% and 1% SA ratios, particularly with 10% and 15% pellet contents. However, increasing SA and pellet ratios did not always result in higher precipitation; in some combinations, a decrease was observed. This indicates that achieving maximum calcite precipitation depends critically on selecting appropriate ratios of SA and pellet content. In the S1 soil, the calcite content ranged from 1.39% to 5.12%. This relatively wide range indicates that both low and high levels of mineralization are possible, suggesting that both the biopolymer and microbial activity functioned efficiently in this soil type. In the S2 soil, the calcite content was measured between 0.91% and 4.51%, indicating that although the maximum values were lower compared to S1, a significant amount of calcite accumulation still occurred. The S3 soil exhibited the lowest range of calcite content, varying from 0.90% to 3.84%. This finding suggests that while the mixing method was also effective in S3, the soil may present more limiting conditions, particularly in terms of porosity or the available surface area for bonding.
SA + MICP combinations applied via the injection method have been observed to result in higher calcite accumulation, particularly in fine-grained soils. This suggests that the S1 soil offers favorable porosity and a binding surface area conducive to microbial precipitation. In a related study, it was reported that Ca-alginate polymeric gel effectively filled the voids in sandy soils containing limestone, thereby narrowing water flow paths and significantly enhancing low hydraulic conductivity [28]. This condition also increases the efficiency of biological precipitation. Nevertheless, the mixing method allows for a more homogeneous distribution of additives within the soil, and when supported by a high pellet content, it can lead to a wide range of calcite precipitation. For example, in the S1 soil, calcite contents ranging from 1.39% to 5.12% were obtained through this method. This wide range indicates that both low and high levels of mineralization are achievable, demonstrating the high potential of SA + MICP combinations applied via mixing for calcite precipitation. However, increasing the pellet or SA content did not always result in higher precipitation; in some mixtures, a decline in calcite content was observed. The relevant literature highlights that in MICP applications using the mixing method, the distribution of additives and the duration of precipitation directly influence the amount of mineralization [38].

3.3.1. Calcite Percentage to Unconfined Compressive Strength Ratio

The highest calcite content (2.72%) was obtained in the S1 soil treated via the injection method; however, the samples subjected to both SA and SA + MICP treatments failed to maintain structural integrity after curing, making UCS testing unfeasible. This finding clearly demonstrates that the amount of calcite alone is not sufficient to enhance mechanical performance. Instead, it highlights the critical importance of parameters such as the micro-scale distribution of calcite, pore structure, and the formation of binding bridges [41]. In contrast, when the same soil type was treated using the mixing method, calcite contents ranging from 1.39% to 5.12% were obtained, indicating a positive correlation with UCS values of up to 398 kPa. Similarly, in the S2 soil, the calcite contents obtained via the injection method ranged between 1.36% to 1.58%, indicating UCS values reaching as high as 121 kPa. On the other hand, with the mixing method, calcite contents varied from 0.91% and 4.51%, indicating UCS values up to 247 kPa. For the S3 soil, both methods resulted in relatively lower calcite contents and UCS values. However, a more pronounced trend of strength improvement with increasing calcite content was observed in the mixing method. These findings indicate that the mixing method enables a more homogeneous distribution of additives within the soil matrix, thereby allowing the precipitated calcite to form more effective binding structures.

3.3.2. Calcite Percentage to Hydraulic Conductivity Ratio

Experimental findings have revealed that the amount of precipitated calcite plays a decisive role in reducing soil hydraulic conductivity. In both injection and mixing methods, an increase in calcite content corresponded with a decrease in hydraulic conductivity. The mixing method facilitated greater calcite accumulation at high SA and pellet ratios due to the homogeneous distribution of additives within the soil. In contrast, the injection method provided effective low hydraulic conductivity in fine-grained soils by promoting more targeted precipitation. However, the success of hydraulic conductivity reduction depends not only on the quantity of precipitated calcite but also on the extent to which this precipitation fills soil pores and its structural uniformity. This relationship has been supported by various studies in the literature. For instance, MICP has been reported to significantly reduce hydraulic conductivity by decreasing soil porosity [42]. Similarly, it has been demonstrated that the effect of calcite precipitation on hydraulic conductivity in sands with different grain sizes is directly related to its pore-filling capacity [43]. Another study reported up to a 90% reduction in hydraulic conductivity in MICP-treated sands, with this effect being dependent on both the volume of precipitation and the quality of its micro-distribution [44]. These findings emphasize that not only the amount of calcite, but also its homogeneous distribution and structural integration within the soil matrix, are critical determinants of hydraulic conductivity reduction [39].

3.4. Scanning Electron Microscopy (SEM) and Energy Dispersive X-Ray Spectroscopy (EDS) Analysis Results

SEM and EDS analyses were conducted on soil samples improved by injection and mixing methods (S1, S2, S3; S1-SA, S2-SA, S3-SA; S1-SA + MICP, S2-SA + MICP, S3-SA + MICP). The results are presented in Figure 13 and Figure 14. In the S1 soil specimen, since the particle size is smaller compared to the S2 and S3 soil specimens, the SEM images presented in Figure 13 and Figure 14 were taken at a magnification of ×1600 for S1, while ×100 magnification was used for S2 and S3. These analyses were performed to visually and chemically verify the microstructural changes and calcite precipitations induced by the addition of SA and MICP within the soil particles. In this context, the effects of the applied improvement methods on soil performance were explained at the micro scale, and the observed increases in strength and low hydraulic conductivity were scientifically substantiated.
In the S1 series, SA applied via the injection method was homogeneously distributed within the microstructure by forming regular film structures on the particle surfaces. A similar observation was reported in a study involving biopolymer and MICP applications, where, in moderately bio-cemented sand specimens, clusters of rhombohedral CaCO3 crystals were observed on the smooth surfaces and surface grooves of sand particles. These crystals were found to precipitate in significant amounts on the particle surfaces, promoting interparticle bridging and resulting in a notable increase in erosion resistance [45]. In the S2 series, SA applied via the injection method adhered to the particle surfaces and formed fibrous film structures within the pores, effectively reducing hydraulic conductivity. Following the application of SA in combination with MICP, irregular yet effective CaCO3 precipitations were observed. These structures are believed to have enhanced interparticle bonding, thereby improving microstructural strength. In the S3 series, however, due to the coarse and smooth surfaces of the particles, the SA and MICP additives could not be homogeneously distributed throughout the soil. Even though the injection method was employed, the effectiveness of the treatment remained limited. The resulting calcite precipitates were localized on the surface, failed to form binding bridges between particles, and thus led to low cementation efficiency. Overall, the effectiveness of injection-based soil improvement is considered to be directly dependent on the surface characteristics and grain size of the soil particles. In coarse-grained soils (S3), the influence of the additives remained largely restricted to the surface, and sufficient cementation could not be achieved. In contrast, in fine- and medium-grained soils (S1 and S2), both chemical cementation and physical gel bonding were observed more prominently.
EDS analysis indicates that SA adheres physically to the particle surfaces; however, crystallization is chemically confirmed through MICP and cementation additives. In the S1 soil, the SA additive was observed to cover the grain surfaces with fibrous film structures in SEM images, and this adhesion is considered to be chemically supported by the presence of Cl elements detected in EDS analysis. The crystalline calcite precipitations observed in SEM images following the application of the SA + MICP combination were further validated by a notable increase in carbon (C) and calcium (Ca) levels in the EDS results. Additionally, the decrease in the silicon (Si) content suggests that the grain surfaces were partially covered with additive products. The consistent presence of aluminum (Al) across all EDS spectra is thought to originate from environmental contamination, clay phases naturally present in the soil, or as a secondary signal from the sample mounting plate.
The mixing method is considered to enable the more homogeneous distribution of SA and more effective calcite precipitation in fine-grained soils such as S1, due to the direct contact it provides between the additives and soil particles. However, despite this relatively positive effect, the desired performance level was not fully achieved. This limitation is thought to result from the irregular adhesion of additives to particle surfaces during the mixing process, which in turn prevents SA from forming a continuous film layer within the microstructure. In the S2 series, SA and MICP additives were able to reach the particle surfaces through the mixing method and induced partial microstructural changes. Nevertheless, SEM images indicate that the binding film structures and calcite precipitations were mostly localized. This may be attributed to the viscoelastic nature of SA, which can lead to its uneven distribution within the pores, particularly in low-controlled mixing applications. In coarse-grained soils such as S3, the additives applied through the mixing method did not produce sufficient bonding effects at the microstructural level. The large and smooth particle surfaces hindered the adhesion of SA and the localized formation of microbial precipitates. As a result, calcite precipitations remained mostly confined to the surface, and the interparticle cementation effect was weak.
EDS analysis indicates that the notable presence of carbon (10.28%) and calcium (7.41%) reflects partial CaCO3 precipitation resulting from the application of SA and MICP additives via the mixing method. The high levels of oxygen and silicon on the surface, along with limited chloride (Cl) content, suggest that the effect of the additives remained localized and that the formation of binding structures occurred in an irregular manner. In a study supporting this finding, the effects of MICP applied through the mixing method on soil stabilization were investigated, and it was observed that CaCO3 precipitates were unevenly distributed in both horizontal and vertical directions [38]. This heterogeneity was attributed to spatial variations in oxygen content, which influenced the mineralization reaction. Furthermore, the study emphasized that the mineralization process alters the pore structure of the soil, which is directly related to the stabilization efficiency of MICP-treated soils.
The high concentrations of oxygen (49.16%) and silicon (28.86%) observed in EDS analysis indicate that the soil surface remains largely covered with natural silica. This suggests that the surface coating effect of SA and MICP products is limited. This finding is consistent with the irregular crystallization and weak binding effects observed in SEM images. The low calcium content indicates that sufficient calcite precipitation did not occur during the MICP process, which is likely due to the non-uniform distribution of SA within the soil matrix. Previous studies have emphasized that for the SA + MICP combination to produce effective results, the components must be applied to the soil in a controlled and layered manner [46,47,48]. According to EDS analysis, the presence of carbon (10.13%) and calcium (3.08%) indicates the formation of CaCO3 as a result of the SA additive and the MICP process. However, this calcium content is lower than that observed in the S3-SA-MICP specimens treated via the injection method. This suggests that the additives applied through the mixing method were not homogeneously distributed within the coarse-grained S3 soil, and that calcite precipitation remained mostly confined to the surface. Overall, it can be concluded that the SA + MICP system applied via the mixing method is less effective in coarse-grained soils such as S3 due to the limited surface area, resulting in lower performance in terms of binding efficiency and low hydraulic conductivity.

4. Conclusions

In this study, two different soil improvement methods—injection and mixing—were applied to three different soil types (S1: fine sand, S2: medium sand, S3: coarse sand) with varying particle sizes, and the effects of these methods on the geotechnical properties of the soils were investigated. To evaluate the impact of the improvement methods on geotechnical behavior, various tests were conducted on the treated soil specimens, including hydraulic conductivity, UCS, and calcite content measurements. Additionally, SEM analyses were performed to examine the morphological characteristics of the microbial precipitates. Furthermore, EDS was used to determine the chemical composition of the formed minerals. The key findings obtained from the experimental studies are summarized below:
  • Both methods—injection and mixing—proved effective in enhancing soil strength through the use of SA and MICP additives. While the injection method improved UCS values across all soil types, the combination of SA and MICP yielded successful results only in the S2 soil. In contrast, the mixing method ensured more homogeneous distribution of the additives within the soil matrix, achieving high strength values up to 398 kPa in the S1 soil. The best performance across all soil types was obtained with the combination of 2.5% SA and 15% pellet content. Moreover, the mixing method provided more consistent and effective strength improvements compared to the injection method. These findings indicate that the influence of SA on UCS is more pronounced when applied via the mixing method.
  • Experimental results demonstrated that both injection and mixing methods have significant potential to enhance soil low hydraulic conductivity. In the injection method, increased SA content and MICP contributed to the clogging of soil pores, thereby reducing hydraulic conductivity. However, factors such as uneven additive distribution, restricted flow in certain zones, and insufficient structural integrity occasionally hindered the achievement of the desired low hydraulic conductivity levels.
  • The combination of 1–1.5% SA + 10–15% pellet + MICP was found to yield the most effective results in terms of reducing hydraulic conductivity. Nevertheless, due to the variability in soil properties, it is essential to optimize application parameters specifically for each soil type. The success of low hydraulic conductivity improvement depends not only on the quantity of precipitated calcite but also on its spatial distribution within the soil matrix, soil type, and microbe–soil interactions.
  • The sustained effectiveness of SA and MICP additives requires careful management of parameters such as proper dosage, controlled application conditions, microbial activity level, precipitation time, and ion balance. In this context, optimizing SA + MICP systems according to site-specific conditions is considered a key factor in achieving long-term and efficient low hydraulic conductivity performance.
  • Overall, the injection method offers advantages in terms of low hydraulic conductivity improvements, while the mixing method delivers higher performance in mechanical strength enhancement. However, the success of both methods depends on various parameters such as additive ratios, microbial activity duration, application conditions, and soil type. Therefore, to ensure sustainable and effective UCS improvement, the selection of the appropriate method should be carefully made based on the soil properties and the intended application purpose.
  • When both the UCS (unconfined compressive strength) and hydraulic conductivity (k) results are considered together, it is evident that both methods are effective in reducing soil hydraulic conductivity; however, the mixing method provides higher strength values. As the optimal treatment configuration, we recommend applying the SA + MICP combination with 2.5% SA and 15% pellet contents using the mixing method. This combination ensures low hydraulic conductivity while significantly enhancing soil stability and strength. Although the injection method is effective in reducing hydraulic conductivity through controlled additive delivery, it yields lower UCS values compared to the mixing method. Therefore, for achieving high strength and low hydraulic conductivity simultaneously, the mixing method with an appropriate SA concentration and pellet ratio is considered the optimum solution.
  • The calcite contents obtained through the injection method indicate that this approach provides relatively controlled and homogeneous calcite formation across all soil types. In contrast, the SA + MICP combination applied via the mixing method resulted in higher calcite accumulation. This broader variation suggests that the mixing method offers a higher yield potential but is also more sensitive to soil properties and application conditions. Therefore, the choice of method should be made based on the desired strength level, soil characteristics, and requirements for uniformity.
  • The amount of calcite obtained through the mixing method was associated with a greater increase in mechanical strength compared to the injection method.
  • It strongly demonstrates that, in the improvement of unconfined compressive strength, not only the amount of precipitated calcite but also its spatial distribution within the soil and the soil–microorganism interaction is among the critical determining factors.
  • Calcite precipitation induced by MICP acts as a complementary mechanism to the gelation effect of SA by filling the remaining voids and enhancing interparticle bonding. The distinct calcium peaks observed in the EDS spectra support the occurrence of this precipitation. Therefore, it is concluded that the gel structure formed by sodium alginate is the primary factor responsible for permeability reduction, while MICP-induced calcite formation provides a secondary sealing contribution.
  • Injection and mixing methods yielded distinct microstructural outcomes: In fine-grained soils (S1), injection led to uniform film formation and dense calcite bridges, while in coarse soils (S3), both methods showed localized and limited precipitation. SEM and EDS analyses confirmed that homogeneous additive distribution and sufficient CaCO3 crystallization are critical for effective cementation—highlighting that soil type and method compatibility are key to microstructural improvement.
Based on the findings of this study, future research is recommended to focus on optimizing additive ratios according to soil type, developing application techniques that ensure homogeneous distribution, and thoroughly investigating the conditions that enhance microbial activity in the MICP process. Furthermore, the long-term performance of bio-based soil improvement should be evaluated under environmental conditions such as freeze–thaw cycles, wetting–drying, and repeated loading. Exploring the combined use of different biopolymer types (e.g., xanthan, chitosan) with MICP can also expand the method’s applicability in field-scale implementations. In this context, the present study provides a valuable foundation in the literature for biopolymer-assisted MICP applications. In addition to the findings of this study, it is evident that not only the amount of calcite but also its spatial distribution plays a critical role in the overall strength. Uneven precipitation may create weak zones within the soil, potentially leading to sudden failure under load. Therefore, developing a Discrete Element Method (DEM)-based modeling approach that links bond strength and distribution to macroscopic behavior could significantly enhance the understanding of this mechanism. It is recommended that future studies incorporate DEM-based analyses. Although the mixing method demonstrated higher unconfined compressive strength under laboratory conditions, its application is generally limited to shallow depths and is difficult to implement over large areas. In contrast, the injection method can be applied in-situ to deeper soil layers using pressurized injection equipment. Therefore, while the mixing method offers certain advantages in terms of strength, the injection method is a more feasible and scalable alternative for field applications, particularly in deep ground improvement. In this context, it is recommended that future studies conduct comprehensive evaluations of the performance of both mixing and injection methods at various depths. Additionally, creating analytical or numerical models that can replicate the slow decline in hydraulic conductivity brought on by microbially induced pore-clogging is a crucial avenue for future research. In biocemented soils, microbial activity and the accumulation of metabolic byproducts have the potential to gradually alter the internal pore network, leading to long-term reductions in permeability. Modeling these dynamic changes would provide valuable insights into the long-term behavior and durability of treated soils. In this regard, the permeability–porosity model proposed by Cao et al. [49], which takes into account oxidative precipitation phenomena, offers a promising framework that could be adapted to biogenic clogging processes in bio-mediated soil systems. What is more, in future studies, we recommend conducting comparative environmental and cost assessments, particularly in conjunction with large-scale field applications, to more comprehensively evaluate the practical feasibility and sustainability of such biocementation techniques.

Author Contributions

M.Ş.: She conducted the experiments, analyzed the results and performed the writing process. S.Ç.: He conducted review, editing and funding acquisition. H.A.: He conducted review and editing, and draft writing. All authors have read and agreed to the published version of the manuscript.

Funding

This study was supported by Atatürk University Research Grant (Grant Number: FDK-2023-12755).

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

The original contributions presented in this study are included in the article. Further inquiries can be directed to the corresponding author.

Acknowledgments

This study is derived from the Ph.D. thesis conducted by Mutlu Şimşek under the supervision of Semet Çelik and co-supervision of Harun Akoğuz at Atatürk University, Graduate School of Natural and Applied Sciences, Department of Civil Engineering.

Conflicts of Interest

The authors declare no conflicts of interest.

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Figure 1. Grading curves of sandy soils.
Figure 1. Grading curves of sandy soils.
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Figure 2. Visual appearances and SEM images of the soil types S1, S2, and S3 used in the study.
Figure 2. Visual appearances and SEM images of the soil types S1, S2, and S3 used in the study.
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Figure 3. Experimental processes (a) Preparation (b) Injection (c) Mixing phase.
Figure 3. Experimental processes (a) Preparation (b) Injection (c) Mixing phase.
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Figure 4. Falling head test conducted on soil specimens to determine hydraulic conductivity.
Figure 4. Falling head test conducted on soil specimens to determine hydraulic conductivity.
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Figure 5. UCS test performed on soil specimens.
Figure 5. UCS test performed on soil specimens.
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Figure 6. Laboratory experiment used to determine the calcite content in improved soil samples.
Figure 6. Laboratory experiment used to determine the calcite content in improved soil samples.
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Figure 7. Hydraulic conductivity results of soil specimens improved by the injection method (S1: fine sand, S2: medium sand, S3: coarse sand).
Figure 7. Hydraulic conductivity results of soil specimens improved by the injection method (S1: fine sand, S2: medium sand, S3: coarse sand).
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Figure 8. Hydraulic conductivity results of soil specimens improved by the mixing method.
Figure 8. Hydraulic conductivity results of soil specimens improved by the mixing method.
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Figure 9. UCS results of soil specimens improved by the injection method.
Figure 9. UCS results of soil specimens improved by the injection method.
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Figure 10. UCS results of soil specimens improved by the mixing method.
Figure 10. UCS results of soil specimens improved by the mixing method.
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Figure 11. The effect of SA and SA + MICP applications on calcite precipitation under different SA ratios in the injection method.
Figure 11. The effect of SA and SA + MICP applications on calcite precipitation under different SA ratios in the injection method.
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Figure 12. Effect of SA and SA + MICP applications on calcite precipitation under different SA ratios in the mixing method.
Figure 12. Effect of SA and SA + MICP applications on calcite precipitation under different SA ratios in the mixing method.
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Figure 13. (a) SEM, and (b) EDS results of the soil samples that improved by injection method.
Figure 13. (a) SEM, and (b) EDS results of the soil samples that improved by injection method.
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Figure 14. (a) SEM, and (b) EDS results of the soil samples that improved by mixing method.
Figure 14. (a) SEM, and (b) EDS results of the soil samples that improved by mixing method.
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Table 1. Basic physical characteristics of the sandy soils.
Table 1. Basic physical characteristics of the sandy soils.
Physical PropertiesS1S2S3
D10 (mm)0.080.390.65
D30 (mm)0.130.460.77
D60 (mm)0.190.530.99
Coefficient of uniformity Cu2.381.351.52
Coefficient of curvature Cc1.031.030.92
Unified Soil Classification System (USCS)SPSPSP
Dry unit weights (kN/m3)15.0114.9114.81
Minimum void ratio emin0.610.630.65
Maximum void ratio emax0.860.870.86
Void ratio e0.740.750.76
~Dr (%)505050
Explanation: The void ratio (e) represents the condition at which the specimen was prepared, corresponding to an approximate relative density (Dr) of 50%. This reflects an intermediate state between the loosest and densest configurations commonly encountered under field conditions.
Table 2. Properties of SA.
Table 2. Properties of SA.
Test NameSpecificationUnit
Appearance (Form)Solid
Loss on drying≤15.5%
Viscosity (c = 1%, Water at 25 °C)5.0–40.0cps
pH (c = 1%, Water at 25 °C)5–8
Table 3. Experimental details.
Table 3. Experimental details.
Test GroupInjection OptionMixing Option Soil TypeUCS TestHydraulic Conductivity TestCaCO3 Test
Culture Medium or Deionized Water (CM or DW)Cementation Solution (CS)Mixing Solution
Solution (mL)Flow Rate (mL/min)SA/CM
(%) *		Solution (mL)Flow Rate (mL/min)SA/CM
(%) *		CM or DW (mL)CS
(mL)
S1S2S3
MICP120203500.20120350✓✓✓✓✓✓
MICP + SA12020.53500.20.5120350✓✓✓✓✓✓
120213500.21120350✓✓✓✓✓✓
12021.53500.21.5120350✓✓✓✓✓✓
120223500.22120350✓✓✓✓✓✓
12022.53500.22.5120350✓✓✓✓✓✓
SA12020.53500.20.5120350✓✓✓✓✓✓
120213500.21120350✓✓✓✓✓✓
12021.53500.21.5120350✓✓✓✓✓✓
120223500.22120350✓✓✓✓✓✓
12022.53500.22.5120350✓✓✓✓✓✓
* Explanation: In the mixing method, for each SA/CM (%) ratio “SA/CM Solution (%)” refers to the weight/volume percentage (w/v%)), pellets were produced by filtering mixtures of SA or MICP solution combined with the cementation solution (i.e., filtering the CM + SA + CS, SA + CS, and CM + CS mixtures). These pellets were then added to the soil specimens at 5%, 10%, and 15% by weight of the soil for each respective ratio, and the mixing application was carried out accordingly.
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Şimşek, M.; Çelik, S.; Akoğuz, H. Comparative Study of Mechanical and Microstructural Properties of Biocemented Sandy Soils Enhanced with Biopolymer: Evaluation of Mixing and Injection Treatment Methods. Appl. Sci. 2025, 15, 8090. https://doi.org/10.3390/app15148090

AMA Style

Şimşek M, Çelik S, Akoğuz H. Comparative Study of Mechanical and Microstructural Properties of Biocemented Sandy Soils Enhanced with Biopolymer: Evaluation of Mixing and Injection Treatment Methods. Applied Sciences. 2025; 15(14):8090. https://doi.org/10.3390/app15148090

Chicago/Turabian Style

Şimşek, Mutlu, Semet Çelik, and Harun Akoğuz. 2025. "Comparative Study of Mechanical and Microstructural Properties of Biocemented Sandy Soils Enhanced with Biopolymer: Evaluation of Mixing and Injection Treatment Methods" Applied Sciences 15, no. 14: 8090. https://doi.org/10.3390/app15148090

APA Style

Şimşek, M., Çelik, S., & Akoğuz, H. (2025). Comparative Study of Mechanical and Microstructural Properties of Biocemented Sandy Soils Enhanced with Biopolymer: Evaluation of Mixing and Injection Treatment Methods. Applied Sciences, 15(14), 8090. https://doi.org/10.3390/app15148090

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