Oral drug delivery has always been the preferred route for patients and physicians alike. However, the oral route presents several challenges, including low permeability and uptake, particularly for therapeutic proteins, with insulin being a prime example. Oral insulin barely reaches the bloodstream as it is digested and breaks down due to the enzymatic environment and its inability to transverse the gut barrier due to its molecular weight. To the best of our knowledge, this is the first report to investigate the potential of genetically engineered (GE) Arthrospira platensis (i.e., Spirulina) as an oral carrier of human insulin (hINS), as well as the intestinal uptake of recombinant hINS produced by GE Spirulina. Furthermore, we report for the first time the application of acetosyringone (ACE) and salicylic acid (SA) to GE Spirulina to enhance hINS production. This study examines the potential of GE Spirulina as an oral delivery vehicle. Agrobacterium containing a pCAMBIA-based binary vector with the hINS gene in its transfer-DNA (T-DNA) segment was used to transform Spirulina via the co-cultivation method. Acetosyringone (ACE) was investigated for its effect on transformation efficiency, and salicylic acid (SA) was investigated for its ability to increase hINS production. The production of hINS was quantified using the hINS ELISA kit. Furthermore, we investigated the scalability potential of this method using laboratory-scale batch bioreactors. Lastly, the potential of engineered Spirulina to deliver hINS was assessed via intestinal uptake and permeability experiments using rat tissues. Colony-PCR confirmed the presence of pCAMBIA-based plasmid in Agrobacterium tumefaciens GV3101 (used to transform Spirulina). Spirulina was found to be sensitive to hygromycin and cefotaxime antibiotics, enabling the harvesting and separation of transformed Spirulina. The PCR amplification of hINS from genomic DNA indicated stable integration into the Spirulina genome, consistent with previous reports; however, additional junction-level confirmation is recommended. Adding 200 µM ACE during the transformation process increased transfection and hINS production compared to the control (2.3 ± 0.15 ng vs. 0.9 ± 0.31 ng, respectively). A second round of transformation increased hINS production while a third transfection cycle reduced it. The addition of 100 µM SA increased hINS production by 2.9-fold. Interestingly, scaled-up production of genetically engineered (GE) Spirulina using a 2 L glass bioreactor resulted in 277% higher hINS production than expected based on volume increase. The hINS produced by GE Spirulina is bioactive, is released intact, and remains stable under intestinal conditions. Ex vivo experiments with the small intestines of rats exposed to GE Spirulina revealed that hINS was uptaken in the rat's ileum > jejunum > duodenum. In contrast, transverse accumulation of the hINS was the highest for the duodenum, followed by the jejunum and ileum. The results of this study demonstrate the potential of GE Spirulina as an effective oral drug carrier, warranting further exploration.

Although physical, chemical, and biological wastewater treatment technologies have improved in recent years, their overall effectiveness remains limited by various challenges. This study evaluated the potential application of swine manure aerobic digestate effluent (SME) as an alternative medium for cultivating the cyanobacterium Arthrospira (Limnospira) maxima to meet the requirements for sustainable microalgae cultivation. A pilot-scale study was conducted using a 200-L photobioreactor to test the efficacy of SME dilutions of 30%, 50%, and 70% (v/v) over 30 days. The objective was to identify the optimal concentration for microalgae growth and nutrient removal. The 30% SME group showed the highest biomass productivity (1.04 ± 0.02 g L⁻1) and superior nutrient removal, with total nitrogen (T-N) and phosphate (PO₄-P) reduction rates of 42.46 ± 3.96% and 46.19 ± 1.36%, respectively, and chemical oxygen demand removal efficiency of 50.05 ± 6.04%. These findings indicate effective organic matter reduction under high-load conditions. The harvested biomass exhibited high protein content (~ 52%) and stable amino acid composition, supporting its potential as a protein-rich livestock feed. However, trace metal accumulation (Zn and Cu) increased at higher SME concentrations, highlighting the need for optimized dilution strategies. These results demonstrate the feasibility of integrating nutrient removal and biomass valorization using microalgae and show that swine manure-derived SME can be directly used without pretreatment as a scalable, eco-friendly approach in circular agriculture.

Phycocyanin (PC), a bioactive phycobiliprotein from Arthrospira maxima, exhibits strong antioxidant and therapeutic potential; however, its spectroscopic characterization becomes challenging when PC is incorporated into complex biopolymeric matrices due to matrix-induced signal interference. In this study, photoacoustic spectroscopy (PAS) was evaluated as an alternative, matrix-tolerant technique for detecting PC encapsulated within polysaccharide-based polymeric matrices, including alginate, agavins, κ-carrageenan, and carboxymethyl cellulose. PC-loaded matrices were characterized by using PAS, UV-vis spectrophotometry, FTIR spectroscopy, and optical microscopy to assess optical absorption, structural features, and potential matrix-analyte interactions. PAS successfully reproduced the characteristic PC absorption band near 620 nm across all matrices and PC concentrations, even in optically dense or highly scattering samples, where UV-Vis transmission measurements are unreliable. FTIR spectra enabled differentiation among polymer matrices and revealed formulation-dependent variations attributable to physicochemical interactions with PC. Morphometric analysis showed substantial differences in encapsulate size and surface characteristics among formulations, although these differences did not affect PAS detection. Overall, the results demonstrate that PAS is a reliable, nondestructive method for identifying PC regardless of matrix composition, morphology, or optical heterogeneity, highlighting its potential as an analytical tool for nutraceutical characterization in biopolymeric delivery systems.

Thermal and microstructure insights into the mechanism of coating/agglomeration/microencapsulation to preserve the antioxidant potential of Spirulina powder (Arthrospira maxima) by fluidized bed processing

Arthrospiramaxima (Am) is a filamentouscyanobacterium with multiple nutraceutical components, such as essentialamino acids, proteins, secondary metabolites, and pigments, such asβ-carotene and phycobiliproteins. Am phycobiliproteins are water-solubleproteins and accessory pigments involved in the photosynthetic processwith multiple beneficial health effects, such as antiviral, anticancer,and antioxidant. The aim of this work was to analyze the cardioprotectiveand antihyperlipidemic effects of Am and Am phycobiliproteins ((ExPhy)containing 76% C-phycocyanin (C-PC), 18.4% allophycocyanin (APC),and 3.8% phycoerythrin (PE)) in an acute myocardial infarction model.Methods: Prediabetic Wistar rats were administered with ExPhy (6.75mg/kg, 12.50 mg/kg, 75 mg/kg) and Am (500 mg/kg) for 21 days, thenthe ischemia/reperfusion model was performed (1/4) measuring infarctarea vs healthy area, in addition to biochemical lipid tests. Results:A reduction in cardiac necrotic damage produced by ischemia was found,as well as a reduction in total cholesterol and LDL cholesterol withoutsignificant differences in triglycerides and HDL; however, a reductionin the atherogenic index was observed in the doses of ExPhy 12.50mg/kg and Am 500 mg/kg administered subchronically. Conclusion: Amand ExPhy showed cardioprotective and antihyperlipidemic action inthe prediabetic rat model, because of its ability to modulate keysignaling pathways involved in cell survival, inflammation, and lipidmetabolism, hence acting as potential adjuvants against damage causedby cardiac ischemia and reperfusion.

Effect of dark period and reactor thickness on Arthrospira maxima continuous cultivation under high frequency pulsed light

An high-performance liquid chromatography (HPLC)photodiode array (PDA) method was developed and validated for quantifying phycocyanobilin (PCB) in Arthrospira maxima extracts.While previous methods for PCB analysis lacked comprehensive validation and struggled with coelution issues, this approach demonstrated high specificity, accuracy, sensitivity, and reproducibility.Effective separation was achieved, with a retention time of 18.7 min.The calibration curve exhibited excellent linearity (r 2 = 1.0000) across a concentration range of 3.125-50 g/mL, with a limit of detection and limit of quantification of 0.22 and 0.67 g/ mL, respectively.Recovery rates ranged from 97.75 to 103.36%, with relative standard deviations (RSD) between 0.50 and 2.14%, confirming the accuracy and reliability of the method.The intra-and inter-day precisions yielded mean contents of 0.122 (RSD: 1.61%) and 0.128 mg/g (RSD: 0.71%), respectively.Notably, this study is the first to validate an HPLC-PDA method for direct quantification of PCB in A. maxima with full validation parameters.By offering a standard for PCB analysis, this study contributes to quality control in research and industrial processes, paving the way for advancements in pharmacology and biotechnology.

Assessing the mass-culture system and proximate content of Arthrospira maxima local for biogas and biohydrogen productions

Effects of salt concentration and trophic mode on glucosylglycerol and phycocyanin production by Arthrospira maxima

• Spirulina had a higher protein content followed by carbohydrates and lipids. • The major amino acids were alanine, glycine, glutamate, serine, and aspartate. • Spirulina protein had promising biological value and protein efficiency ratio. • The major fatty acids were α-linolenic, palmitic, linoleic, and γ-linolenic acid. • Spirulina had a promising nutritive value, atherogenicity, and thrombogenicity indices. This research studied Arthrospira ( Spirulina ) maxima for their approximate chemical composition, fatty acid composition, amino acid composition, protein nutritional quality, lipid nutritional quality, and anti-amylase capacity. The study on Arthrospira indicates a significantly high protein content of 45.50 %, along with 21 % carbohydrates, 17 % lipids, and 9.33 % ash content. The amino acid profile shows high levels of alanine (17.38 g), glycine (11.75 g), and glutamic acid (9.69 g) per 100 g protein. The protein quality is noteworthy, with significant amounts of protein efficiency ratio, and essential, non-essential, hydrophobic, ketogenic, branched-chain, flavor, and sulfur amino acids. Additionally, the study highlights a diverse fatty acid profile, including linolenic and palmitic acids, with a high percentage of unsaturated fatty acids (75.76 %) and polyunsaturated fatty acids (71.63 %). Given the lipid nutritional quality indices, Arthrospira had promising unsaturation, peroxidability health-promoting, omega-6/omega-3, hypocholesterolemic, nutritive value, atherogenicity, and thrombogenicity indices. Ultraviolet absorption, fluorescence quenching analysis, and Colorimetric assay revealed that protein and lipid hydrolysate interact with amylase and inhibit amylase activity. Therefore, Arthrospira can be considered a functional food with high nutritional quality and is imperative as an amylase inhibitor for diabetes management.

Emulsifying properties of Maillard reaction products formed using spirulina (Arthrospira maxima) protein extract coupled with various saccharides

This study develops an optimized methodology for extracting food-grade phycocyanin (PC) from Arthrospira maxima using cell disruption techniques, specifically ultrasound and freeze-thaw cycles. The research investigates the impact of various operational parameters, such as pH, temperature, and biomass concentration, on PC stability and extraction yield. Filtration was found to preserve higher PC concentrations compared to centrifugation, while spray drying effectively retained the pigment. Results indicate that pH 6.0 and temperatures below 45°C best preserved PC stability. Ultrasound treatment at 50% amplitude for 7min maximized extraction yield (72.6mg/g) with a purity of 1.18, though extending treatment time did not significantly improve results. Combining freeze-thaw cycles with ultrasound yielded lower extraction efficiency compared to ultrasound alone. The findings offer a scalable approach for producing high-purity PC, with potential applications in the food industry as a natural dye.

Co-encapsulation of hydrophilic and hydrophobic compounds within a single delivery system remains a significant challenge across various scientific and industrial fields. Towards this direction, an encapsulation strategy is proposed, enabling the simultaneous incorporation of both hydrophilic and hydrophobic biomolecules within a hydrogel matrix. Specifically, the cyanobacterial protein phycocyanin (hydrophilic), extracted and purified by dry Arthrospira maxima biomass, and curcumin (hydrophobic) bound to bovine serum albumin (BSA) were utilized. This approach facilitates the indirect entrapment of hydrophobic molecules within the hydrophilic hydrogel network. The structural and physicochemical properties of the resulting hydrogels were characterized using optical analysis, scanning electron microscopy (SEM), and confocal laser scanning microscopy (CLSM). Additionally, the antioxidant potential of the encapsulated biomolecules was evaluated to assess their functionality after the encapsulation. Furthermore, a cell viability assay confirmed the hydrogel's biocompatibility and lack of toxicity, demonstrating its suitability as a multifunctional biomaterial for biomedical and pharmaceutical applications.

The antifouling performance of a zwitterionic sulfobetaine-hydroxyethyl-containing polymethylmethacrylate ter-co-polymer (PSBM) is evaluated against three photosynthetic strains, namely Chlorella sp., Nannochloropsis sp., and Arthrospira maxima. PSBM-coated polymethylmethacrylate (PMMA) surfaces displayed a significantly reduced propensity for biofilm formation compared to rough and untreated controls, leaving clean surfaces after 7 days of exposure. A tribological approach was adopted to estimate the long-term durability of the PSBM coating. Repeated cycles of exposure to Chlorella sp., Nannochloropsis sp., and A. maxima biomass subject the coating to stress and continuous biofilm challenges. After several cycles, the PSBM coating maintains a higher antifouling efficacy than the untreated PMMA surface, suggesting stability and high potential in photobioreactor applications.

Phycocyanin (C-PC) is a hydrophilic blue pigment found in cyanobacteria, rhodophytes, and cryptophytes. Its intense blue color and antioxidant capacity make phycocyanin an attractive substance for food and cosmetic industries. In this study, phycocyanin extracts from Arthrospira maxima, obtained by different extraction methods, were used to investigate the influence of preserving agents on thermostability of phycocyanin. C-PC remained stable at 60°C, but rapidly degraded at 70°C, and was completely lost at 80°C within 5 minutes. Addition of sorbitol, sucrose, and glucose at 32% (w/v) to phycocyanin extracted through ultrasonication and flocculation with chitosan in organic acid provided the highest stabilizing effects, with CR values of 96.67 ± 0.01 %, 83.41 ± 0.67 %, and 76.21 ± 1.34 %, respectively. The corresponding half-live values increased from 13.77 ± 0.21 min (control) to between 76.74 ± 5.12 and 614.83 ± 1.18 min, followed by xylitol, fructose, and maltose. Xylose, lactose, citric acid, and ascorbic acid showed no effects. The stabilizing effect of NaCl was dependent on pH of extracts, remaining stable at pH 6 – 7, but showing no significant effect at pH 3 – 4, and leading to precipitation. Incorporating 14 – 20% (w/v) NaCl in C-PC at pH 6.49 increased CR values from 88.61 ± 0.31 to 99.10 ± 0.01 %, with half-life values ranging from 172.03 ± 4.92 to 2291.19 ± 14.51 minutes. Conversely, the stabilizing effect of sugars was most effective in C-PC at pH 3 – 4, and least effective in C-PC obtained by freeze/thawing at pH 6 – 7.

The antifouling performance of a zwitterionic Sulfobetaine-Hydroxyethyl containing-Polymethylmethacrylate ter-co-polymer (PSBM) is evaluated against three photosynthetic strains, namely Chlorella sp., Nannochloropsis sp., and Arthrospira maxima. PSBM coated polymethylmethacrylate (PMMA) surfaces displayed significantly reduced propensity for biofilm formation, compared to rough and untreated controls, leaving clean surfaces after 7 days of exposure. A tribological approach was adopted to estimate the long-term durability of the PSBM coating. Repeated cycles of exposure to Chlorella sp., Nannochloropsis sp., and A. maxima biomass subject the coating to stress and continuous biofilm challenge. After several cycles, the PSBM coating maintains a higher antifouling efficacy than untreated PMMA surfaces, suggesting stability and high potential in Photobioreactors application.

In this study, a Size Exclusion High-Performance Liquid Chromatography (HPLC-SEC) method was evaluated for its application in determining the apparent molecular weight of biopolymers, specifically exopolysaccharide (EPS) released by the cyanobacterium Arthrospira maxima. The primary objective of this research was to develop a simple, reliable, and reproducible analytical method for measuring the molecular weight of polymers, based on a calibration curve that would characterize polymers using different polyethylene glycols (PEGs) as standards. This approach is important for two reasons, firstly, it provides an alternative to the use of dextran, and secondly, traditional methods of molecular weight measurement can be complex especially when applied to high molecular weight polymers or biopolymers with diverse and variable structures. The HPLC-SEC method was optimized to ensure the separation of various PEGs based on their retention times and to minimize any interferences that could affect the accuracy of the results. Quantification was carried out after the PEGs had been separated, each characterized by a number that typically indicates the average molecular weight of the molecules. Although PEGs are relatively high in molecular mass, the chromatographic conditions were adjusted to enable their clear resolution, which is crucial for ensuring reliable and precise quantification of these standards. Separation was achieved using an Ultra-hydrogel DP column (Waters), which specializes in size exclusion and is designed to allow effective separation of polymers of varying sizes. The mobile phase consisted of a water-acetonitrile mixture in a 65:35 ratio. This composition ensured the optimal separation and enhanced resolution of the samples. The separated PEGs were detected using a refractive index detector, which enabled highly accurate quantitative analysis of the various standards. The six PEG standards employed in this work were PEG300, PEG400, PEG1500, PEG4000, PEG6000, and PEG8000. Remarkably, all these standards were successfully separated in less than 10 minutes, demonstrating the efficiency and rapidity of the developed method. To ensure the robustness and reliability of the method, a set of validation parameters was rigorously assessed. Specificity was confirmed by verifying the absence of interferences from other components in the sample matrix. Linearity was evaluated by analyzing the detector response to different PEG standards across a concentration range, and a considerable degree of correlation was obtained. Precision and intermediate precision were confirmed through repeated analyses, with minimal variation observed between repetitions, thus ensuring the reproducibility of the method. Furthermore, accuracy of the method was validated by comparing the experimental results with theoretical molecular weight values, which showed an excellent agreement between the two, reinforcing the method’s validity. This present study makes a significant contribution to the field of analytical methods for characterizing biopolymers. The optimization of the HPLC-SEC method, coupled with a full validation, enables the precise and rapid determination of the molecular weight of the studied biopolymer, which is essential for its potential use in various industrial applications. Moreover, this methodology could be useful for the analysis of other biopolymers, paving the way for further advancements in the analysis of polymers in different contexts, including biomaterials, biotechnological products, and environmental studies.

Background/Objectives: In recent years the global incidence of cancer during pregnancy is rising, occurring in 1 out of every 1000 pregnancies. In this regard, the most used chemotherapy drugs to treat cancer are alkylating agents such as cyclophosphamide (Cp). Despite its great efficacy, has been associated with the production of oxidative stress and DNA damage, leading to embryotoxicity, genotoxicity, and teratogenicity in the developing conceptus. Therefore, this study aimed to investigate the protective role of phycobiliproteins (PBP) derived from Arthrospira maxima (spirulina) in reducing Cp-induced embryotoxicity and genotoxicity in pregnant CD1 mice. Methods: Pregnant CD1 mice were divided into five groups: control, Cp 20 mg/kg, and three doses of PBP (50, 100, and 200 mg/kg) + Cp co-treatment. PBP were administered orally from day 6 to 10.5 dpc, followed by a single intraperitoneal dose of Cp on 10.5 dpc. Embryos were collected at 12.5 dpc to assess morphological development and vascular alterations, while maternal DNA damage was evaluated using micronucleus assays and antioxidant enzyme activity in maternal plasma. Results: PBP exhibited a dose-dependent protective effect against Cp-induced damage. The 200 mg/kg PBP dose significantly reduced developmental abnormalities, micronucleated polychromatic erythrocytes, and oxidative stress, (as evidenced by increased SOD and GPx activity). Conclusions: Phycobiliproteins from Arthrospira maxima (spirulina) effectively reduced Cp-induced morphological and vascular alterations in embryos and genotoxicity in pregnant mice. These findings highlight their potential as a complementary therapy to mitigate teratogenic risks during chemotherapy. Further research is needed to optimize dosing and explore clinical applications.

The biostimulant effect of Spirulina (Arthrospira maxima) on various agricultural crops is well-documented, demonstrating improvements in yield, productivity, and product quality. However, limited research has explored its impact on bioactive compound stimulation, and no studies to date have examined its effect on pigmented corn cultivars high in anthocyanins. Mexico harbors a diverse array of pigmented maize varieties rich in anthocyanin content, with antioxidant activity, which decreases oxidative stress preventing degenerative diseases. In this study, the biostimulant effect of commercial hydrolyzed Spirulina (Ficocyan®) was evaluated on polyphenols, flavonoid, anthocyanins contents, and antioxidant capacity (DPPH and ABTS assays) in three maize cultivars (black, red and purple). The application of Spirulina significantly influenced polyphenol and flavonoid of red variety (P < 0.05). Conversely, anthocyanins were affected in the black variety. Regarding antioxidant capacity, the DPPH assay revealed no significant differences due to Spirulina treatment; however, the ABTS assay showed a difference in antioxidant capacity for red and purple varieties treated with Spirulina.

Emulsifying and Physicochemical Properties of Maillard Reaction Products Formed Using Spirulina (Arthrospira Maxima) Protein Extract Coupled with Various Saccharides