Current Status of Alginate in Drug Delivery

Alginate is one of the natural polymers that are often used in drug- and protein-delivery systems. The use of alginate can provide several advantages including ease of preparation, biocompatibility, biodegradability, and nontoxicity. It can be applied to various routes of drug administration including targeted or localized drug-delivery systems. The development of alginates as a selected polymer in various delivery systems can be adjusted depending on the challenges that must be overcome by drug or proteins or the system itself. The increased effectiveness and safety of sodium alginate in the drug- or protein-delivery system are evidenced by changing the physicochemical characteristics of the drug or proteins. In this review, various routes of alginate-based drug or protein delivery, the effectivity of alginate in the stem cells, and cell encapsulation have been discussed. The recent advances in the in vivo alginate-based drug-delivery systems as well as their toxicities have also been reviewed.


Chemistry and Physicochemical Properties of Alginate.
Alginate is a polysaccharide extracted from brown seaweeds, including Laminaria hyperborea, Laminaria digitata, Laminaria japonica, Ascophyllum nodosum, and Macrocystis pyrifera [1,2]. It is composed by a sequence of two (1Ñ4)linked α-L-guluronate (G) and β-D-mannuronate (M) monomers. e proportion of M and G blocks may vary with the type of seaweed from where it is extracted (Figure 1). For example, alginate extracted from Laminaria digitata and Ascophyllum nodosum has been shown to have M/G ratios of 1.16 and 1.82, respectively. Alginate is a biocompatible polymer with very low toxicity [3].
ese are the main advantages that make alginate one of the biopolymers with the widest biomedical applicability [4,5]. One of the most common applications of alginate is their use as an excipient in drug-delivery systems, namely, acting as a stabilizer agent in various pharmaceutical formulations [6,7].
Alginate has carboxyl groups which are charged at pH values higher than 3-4, making alginate soluble at neutral and alkaline conditions to promote the widespread use of alginates. For some drugs which require greater protection with preferential absorption in the intestinal tract or other conditions such as modified drug release, alginate is a preferable polymer. us, solubility and pH sensitivity make alginate a good biomaterial for drug-delivery systems [8]. Sodium alginate is the type of alginate mainly used in the pharmaceutical industry and may be used for the purpose of extending the drug release. Using sodium alginate with different chemical features and degree of viscosities, the slow release of ibuprofen from press-coated tablets was reported [8]. In acidic environments, alginate carboxyl groups are protonated, thereby limiting drug release. Alginate has the ability to crosslink with Ca 2+ ions through an ionotropic gelation process, usually above pH 6. Ba 2+ or Zn 2+ ions are also used as crosslinkers [9][10][11].
Alginate hydrogels are applied in wound healing treatments through the construction of wound dressings [12][13][14][15]. Several studies showed that the bioavailability of drugs encapsulated in alginate hydrogels is greater than that of the free drug applied directly at the lesion site, thus increasing the efficacy of healing. Alginate hydrogels are also used widely in tissue regeneration treatments and cell encapsulation [16][17][18][19][20][21][22]. Alginate may be used in the construction of capsules for cell encapsulation often associated with cytotherapy treatments or simply the creation of cellular microcultures in more complex systems. A new approach to the construction of alginate-based capsules for the incorporation of different types of cells has been demonstrated [23]. Cells were encapsulated in alginate liquefied particles, followed by coating it with chitosan and alginate. Poly(lactic acid) microparticles along with the cells were coencapsulated to protect cell survival with high viability of the encapsulated cells. Hydrogels obtained from alginate nowadays present some advantages of being appropriate materials to be used in tissue engineering and regenerative medicine applications [23][24][25][26][27][28][29][30][31].
Some important uses of alginates in nanomedicines in the forms of dendrimers, nanocrystals, emulsions, liposomes, solid lipid nanoparticles, micelles, and polymeric nanoparticles have provided advantages over conventional medicines including efficacy, safety, physicochemical properties, and pharmacokinetic/pharmacodynamic profiles [32].

Crosslinker for Alginate
Micro/Nanoparticles to Encapsulate Drugs. Typical shapes of alginate are processing through several different techniques, including emulsion, multiple-phase emulsion, and cation crosslinked encapsulation (Ca 2+ , Ba 2+ , or Cu 2+ ) [33][34][35][36][37]. e ability of alginate to create complexes with other biomaterials by electrostatic interactions, chemical modification, or crosslinking can be exploited for building hybrid and more versatile DDSs. Capsules constructed from chitosan/alginate-PEG complexes are reliable models for encapsulating proteins, such as albumin, one of the most common model proteins used in controlled release studies [38][39][40][41][42][43]. is approach can promote higher control release of drugs, proteins, and other biomolecules.

Effect of Different Classes of Crosslinkers on Alginate
Polyelectrolyte Nanoparticle. Mirtic et al. [10] investigated the preparation of alginate nanoparticles using complexation of different classes of crosslinkers (divalent cations, polycations, and positively charged surfactants) and found that alginate nanoparticles were formed across a limited range of molar ratios that were specific for each crosslinker and had different size and stability. Additionally, the ionic strengths of the media influenced the characteristics and stabilities of the polyelectrolyte nanoparticles.

Effect of Divalent Cation on Morphology and Drug-Delivery Efficiency.
A study by Deepika et al. [44] was about the formation of levofloxacin in chitosan-alginate hybrid gel for controlled release and effect of divalent alkaline ions (Mg 2+ , Ca 2+ , Sr 2+ , and Ba 2+ ) on encapsulation efficiency and drug release kinetics from chitosan-alginate nanostructure was investigated. e particle size increases and encapsulation efficiency decreases with the size of the divalent ions. Spherical shaped particles were formed by Mg 2+ and Ca 2+ , whereas Sr 2+ and Ba 2+ produced nonspherical particles. Transformation of microspheres is shown by SEM as truncated tetrahedron by Sr 2+ and clear rod shape by Ba 2+ was identified. is suggested that metal ions have a significant influence on the morphology, drug encapsulation, and release profile of the chitosan-alginate hybrid polymer nanoparticles.

Effect of Zinc-Ion Complex with Alginates.
Kotagale et al. [45] complexed alginates with zinc metal ion to improve beads' physicochemical and biological properties for controlling the drug release. ey found that the atenolol-zinc polymeric beads exhibited pulsed release with increased half-life. Moreover, no significant differences in in vitro and in vivo atenolol release behavior among the N,Odimethyl, N-methyl, or N-benzyl hydroxylamine derivatives of sodium alginate were observed.

Effect of Ferric Ion Crosslinker on Alginates.
Microspheres of acrylamide-(AAm-) grafted poly(vinyl alcohol) (PVA)/sodium alginate (NaAlg) were prepared by crosslinking with FeCl 3 and 5-fluorouracil (5-FU) [46]. Microspheres were characterized by particle diameter, equilibrium swelling values and morphology, elemental analysis, and release profiles. is group studied the effects of PVA-g-PAAm/NaAlg ratio, drug/polymer ratio, crosslinker concentration, and exposure time to FeCl 3 on the release of 5-FU. e highest 5-FU release was found to be as 99.57% after 6 h for PVA-g-PAAm/NaAlg and release kinetics was described by Fickian and non-Fickian approaches.

Purposes of Encapsulation of Drugs Using Alginates.
Alginate can also undergo complexation with natural polymers, like chitosan, to enhance the absorption and cargo protection in oral delivery, for example, for the administration of insulin [47,48]. Alginate was also combined with pectin polymer which has a similar mechanism. is research also showed successfully encapsulated drugs [49][50][51][52]. Alginate-based drugs encapsulated into nanoparticles/microparticles with various purposes are presented in Table 1.

Use of Alginates in the Pharmaceutical Industry.
Many application areas of sodium alginate-based drug-delivery systems, and these systems can be formulated as gels, matrices, membranes, nanospheres, microspheres, and others [2,81]. Researchers are exploring possible applications of alginates as a coating material and preparation of controlled release drug-delivery systems.

Alginate for Protein Delivery and Cell Encapsulation.
Alginate microparticles as a carrier for protein delivery prepared by spray-drying processes have been studied for their application in nasal and pulmonary drug delivery [85][86][87] prepared inhalable alginate particles (of an average diameter 3.23 ± 0.25 μm) with a high encapsulation efficiency of 97% with the preserved structure and bioactivity of BSA. e alginate particles released approximately 20% of the loaded BSA over 24 h and then a slow release occurred, reaching a cumulative release of only 35% after 180 h. Möbus et al. [88] prepared Zn 2+ -crosslinked alginate microparticles containing the model protein BSA via a simple one-step spray-drying process to produce microparticles of 2-4 µm size. ey found BSA release into the simulated lung fluid increased with an increasing content of protein in the alginate microparticles. Alginate hydrogels have also been studied for oral delivery of proteins [89,90]. Hariyadi et al. [91] prepared alginate microspheres containing lysozyme and insulin resulting in 30 to 60 μm in size with high protein loadings. Moreover, it was found to retain 75% activity using the ARCHITECT ® assay and exhibit at least 80% bioactivity using the Micrococcus lysodeikticus assay. Another study using BSA demonstrated that the BSA release from the hydrated microparticles reached less than 7% in the simulated gastric fluid over 2 h, whereas 90% of the protein load was gradually released in the simulated intestinal fluid over 10 h. Another cell viability study was also conducted by Morachis et al. [92]; Severino et al. [93]; Joddar et al. [94]; Ciriza et al. [95]; Yoncheva et al. [96]; and Gurruchaga [18]. Applications of alginates for protein delivery and cell encapsulation are presented in Tables 2 and 3.

Alginate Particles with Ovalbumin (OVA)
(1) Peptide as a Carrier and Adjuvant. Ovalbumin (OVA) peptide 323-339 encapsulated in alginate has been reported to be involved in immune response as carrier and adjuvant for the immune therapy of cancer [53]. A tumor model was established in C57BL/6J mice via subcutaneous injection of 3 × 105 B16-OVA tumor cells. Alginate/OVA peptide inhibited tumor progression more effectively than using the peptide alone. e viability and uptake study illustrated that this particle is safe and nontoxic. Furthermore, alginate particles can promote the activation of surface markers on macrophages. ELISA assay showed that the particles with peptide can promote the secretion of inflammatory and effector cytokines from macrophages.

Liposomal Alginate for Bupivacaine Delivery and MSC
Function. Mesenchymal stromal cell (MSC) therapies have become potential treatment options for multiple ailments and traumatic injuries. Davis et al. [103] developed and characterized a sustained release delivery formulation comprised of alginate-encapsulated liposomal bupivacaine to evaluate the effect of this formulation on the secretion of three key MSC regulatory molecules, interleukin 6 (IL-6), prostaglandin E2 (PGE2), and transforming growth factorbeta 1 (TGF-β1). Bupivacaine release profile analyses indicated that the mode of drug delivery controlled the liposomal-alginate (LA) concentration over time and pathway analysis identified several shared and cytokine-specific molecular mediators for IL-6, PGE2, and TGF-β1. ese studies support the potential utility of LA for anti-inflammatory cell therapy coadministration.

Curcumin-Alginate-Based Composite Sponges.
Alginate-based composite sponges were developed as carriers to prolong the gastric retention time and controlled release of curcumin-loaded self-microemulsifying drug-delivery systems (Cur-SMEDDS) [104]. Researchers used adsorbent (colloidal silicon dioxide) and additional polymers such as sodium carboxymethyl cellulose (SCMC) and hydroxypropyl methylcellulose (HPMC) to form composite e formulation exhibited a droplet size of approximately 30 nm and provided a sustained release.

Application of Alginates in Context of the Routes of Drug Administration
Alginates have been extensively investigated for delivering drugs via oral, parenteral, pulmonary, and transdermal routes ( Table 4). Using alginate as a single polymer or the combined polymer, controlled or sustained release delivery of quercetin, isoniazid, rifampicin, ciprofloxacin, bovine insulin, and lentivectors has been investigated. All formulations showed increased entrapment efficiency of drugs, increased dissolution and bioavailability, and reduced degradation of drugs [105-107, 109-112, 130-132]. Some chemotherapeutic agents encapsulated in alginate polymer showed enhanced penetration in the target cells. Antigen-encapsulated alginate showed enhanced immune response [8,115,116,133,134]. Alginates have been also widely investigated for pulmonary drug delivery [99,117,[119][120][121][122][123][124][125][126][127][128]. Alipour et al. developed paclitaxel-alginate microparticles which increased the site-specific efficacy of drugs with reduced toxicity [117]. Using alginate and PLGA polymers, Abdelaziz et al. studied inhalable particulate delivery of cisplatin and doxorubicin for lung cancer therapy [120]. e alginate-based BSA and BCG vaccines have been used to study the efficacy of smaller inhalable vaccines, which provided better protection and more immunogenic effect [99,124,125]. Applications of alginate in transdermal delivery for wound dressing or wound healing were shown to be effective to produce a high porosity and sustained release and able to inhibit preinfection [126][127][128]135].

Alginate-Based Hybrid Aerogel Microparticles for Mucosal
Drug Delivery. Some polysaccharides (e.g., alginate, chitosan, and pectin) have been applied as biopolymer aerogels to have mucoadhesive properties for mucosal drug delivery [136] Alginate-based hybrid aerogels of microparticles (<50 μm) were produced. Low methoxyl pectin and κ-carrageenan were also cogelled with alginate and further dried with supercritical CO 2 (sc-CO 2 ). Spherical mesoporous aerogel microparticles were obtained for alginate, hybrid alginate/pectin, and alginate/κ-carrageenan aerogels, presenting high specific surface area and mucoadhesive properties. e microparticles were loaded with ketoprofen and quercetin. Release of both drugs from alginate/κ-carrageenan aerogel was slightly faster compared to alginate/ pectin indicating that alginate-based aerogel microparticles are potential for mucosal drug-delivery applications.

Alginates for Ocular Drug Delivery.
To develop potential ocular drug delivery, mucoadhesive microspheres is one of the best approaches to prolong the drug residence inside the  Pancreatic rat islets Alginates Cell encapsulation by zwitterionic group Alginates improved outcome of islet encapsulation in a chemically induced diabetic mouse model [22] Riboflavin Sodium alginate and furfurylamine Coupling and photo-crosslinked method Photo-crosslinked F-alginate resulted in slow release and potential for cell growth enhancement for medical application, biomaterials, soft and hard tissue applications, and tissue interfaces [102] Advances in Pharmacological and Pharmaceutical Sciences 5 cul-de-sac, consequently increasing the bioavailability. us, some researchers worked to overcome the limitations of ocular drug delivery [137][138][139]. e chitosan-sodium alginate microspheres or other polymers encapsulating of ocular drugs have been investigated widely. Sodium alginate microspheres prepared were in particle size range suitable for ocular purpose and were able to improve the therapeutic efficacy.

Various Techniques to Produce Alginate Micro/Nanoparticles for Drug Delivery
Over the years, various methods have been developed to fabricate drug-delivery particles of bioactive substances. Using superhydrophobic surfaces, it is possible to produce polymer particles suitable as DDSs. is method allowed loading drugs into spherical structures with an encapsulation efficiency close to 100% [145,146]. Goncalves et al. [136] developed alginate microparticles which were shown to have Resveratrol Chitosan, alginate, and poly(d,llactide-co-glycolide) Transdermal Nanoprecipitation [127] Metronidazole Alginate Transdermal Ionotropic gelation combination with freezethawing cycle [128,129] 6 Advances in Pharmacological and Pharmaceutical Sciences perfluorocarbon breakthrough capacity when subjected to vibration by ultrasound waves. Results showed a disruption of these microparticles after 15 min of exposure, suggesting that such structures are promising DDSs controlled externally by acoustic stimuli. Another strategy to synthesize particles relies on complexation, based on the electrostatic interactions between alginate at neutral and alkaline pH values, bioactive agents, and other kinds of naturally occurring polymers, such as the polycation chitosan [147][148][149].

Preparation Techniques for Production of
Alginate Nanoparticles

Oligopeptide-Side Chained Alginate via the Amidation Method.
A melittin-targeting drug carrier was successfully synthesized by the grafting of sodium alginate to an oligopeptide via an amidation method at different oligopeptide: alginate unit molar ratios [150]. e average sizes of the oligopeptide-alginate nanoparticles formed decreased with increasing oligopeptide contents, indicating intramolecular interactions between oligopeptide-side chains. e results confirm that the derivation of an oligopeptide-side chain in alginate offers a specific binding site for melittin and effectively works in cancer chemotherapy.

Chitosan/Alginate Nanoparticles by Emulsification and
Ionotropic Gelification. Curcumin-diglutaric acid (CG) is a prodrug of curcumin encapsulated into chitosan/alginate polysaccharide-based nanoparticles [151]. CG-loaded chitosan/alginate nanoparticles were prepared by o/w emulsification and ionotropic gelification, with the conditions optimized using response surface methodology. e CGloaded chitosan/alginate nanoparticles showed better stability compared to a CG dispersion in water. e nanoparticles showed slow cumulative release and the release pattern was mainly controlled by Fickian diffusion and erosion of polymer materials. CG-loaded chitosan/alginate nanoparticles showed higher in vitro cellular uptake in human epithelial colorectal adenocarcinoma (Caco-2 cells) and better anticancer activity against Caco-2, human hepatocellular carcinoma (HepG2), and human breast cancer (MDA-MB-231) cells.

Nutraceutical Nanodelivery
System. Alginate nano/ microspheres were produced by emulsification/internal gelation of sodium alginate within vegetable oils containing surfactant, followed by CaCl 2 addition resulting in hardened particles [153]. Size of nanoparticles decreased at higher oil and surfactant contents, higher molarity of CaCl 2 , and lower alginate concentrations. Moreover, encapsulation efficiency was inversely proportional to the size of nanoparticles.

Alginate/Chitosan Formulations for Ciprofloxacin-Controlled Delivery.
Kyziol et al. loaded ciprofloxacin in alginate beads with an emulsification technique in combination with an internal gelation method [154]. Hydrodynamic diameter and zeta potential showed of 160 nm and −32 mV in the case of AL_CP and ca. 240 nm and ca. +14 mV in the case of AL_CP_CS, respectively. ey found that alginate beads with encapsulated ciprofloxacin covered with chitosan were effective oral delivery system since limited ciprofloxacin was release in gastric.
Various techniques which have been used to produce alginate nanoparticles are presented in Table 5.

Preparation Techniques for Production of Alginate
Microparticles. Some techniques were used to produce alginate microparticles. Production is by conventional emulsification using sodium alginate single or combination polymer with chitosan to encapsulate a variety of drugs including glucose oxidase [167], paclitaxel [168], cocoa extract [169], and diclofenac sodium [170] or double emulsification techniques [171].
Another method is internal gelation technique, which by using sodium alginate polymer to entrap drug of doxorubicin was done by Giovagnoli et al. [35], diclofenac by Ahmed et al. [172], L-α-phosphatidylcholine by Semmling et al. [173], and sulfasalazine by Tavakol et al. [174]. Extrusion dripping method was also used to optimize sphericity of particles and shape deformation [175]. e more recent technique to produce microparticles was an impinging aerosol technique to successfully encapsulate propranolol HCl by Hariyadi et al. [89] and high-voltage electrostatic bead generator for BSA-alginate microparticles by Ørning et al. [176]. Mishra et al. [177] used gas blowing technique to contain verapamil HCl resulting in faster/burst drug release; however, importantly a strong mechanical strength and drug integrity were maintained in hydrogel polymeric network.
from chitosan and glycerophosphate [179]. ere are different mechanisms of release of a bioactive agent from the carrier, such as through variations of temperature and pH and the use of biodegradable materials or enzymatic degradation, among other chemical and physical stimuli-responsive methods [42,[185][186][187][188][189]. Hadijev et al. [180] studied hydrogels which mostly applied drug diffusion as a release mechanism; however, this can be changed with the properties to broadly change the solute diffusion coefficient as the gel system swells. According to Gao et al. [183], mechanism of release of hydrogels can be modified to have more steady release behavior by adding some copolymer which is able to interact and may change the chemical structure, morphology, and rheology characteristics, thus affecting release behavior and mechanism.

Toxicity.
Alginate nanoparticles and microparticles were considered safe, although some studies about safety and toxicity were widely conducted. For example, Spadari et al. [120] investigated alginate nanoparticles as a nontoxic delivery system for miltefosine (MFS) in the treatment of candidiasis and cryptococcosis. Alginate nanoparticles were produced using the external emulsification/gelation method and toxicity on red blood cells and Galleria mellonella larvae were assessed. MFS in alginate nanoparticles presented no hemolytic effect and no toxicity in G. mellonella larvae. ese results showed the potential and nontoxic use of alginatebased drug-delivery systems as carriers to control the fungal infection in the in vivo model of G. mellonella. In vivo study verified that the embolic efficacy of microspheres was similar to that of commercially available alginate microsphere embolic agents [14]. For colon study, Patole and Pandit entrapped mesalamine in variety of polymers including alginate, HPMC, and Eudragit FS-30D and found histopathologically no signs of ulceration or bleeding of the released microspheres [190]. Other in vivo studies including anti-inflammatory, mucoadhesion test, and histopathological were conducted by researchers [191][192][193][194][195].
For vaccine delivery, research using chitosan, trimethyl chitosan (TMC), and alginate was conducted by Mosafer et al. using inactivated PR8 influenza virus for mucosal vaccine delivery. PR8-chitosan formulation elicited higher IgG2a and IgG1 antibody titers compared with PR8-TMC. Alginate coating significantly decreased the antibody titers and less immune response was induced [121].
In vivo study for the transdermal application was done by Hariyadi et al. [196]. ey showed the effectiveness of glutathione-alginate microspheres in decreasing matrix metalloproteinase-1 (MMP-1) expression in the dermis tissue of mice.
Natural products have been investigated by researchers in vivo. Alginate polymer-encapsulated black seed oil for intestine-targeted drug delivery has been studied by Azad et al. (2020) in the forms of gastrointestinal distribution study [197]. ey found uniform distribution of beads after oral administration in rats.
Beside in vivo investigation, ai et al. indicated low toxicity of lovastatin-alginate and chitosan nanoparticles in mice in the acute toxicity test [198].

Conclusions
is paper provides a comprehensive review of the current status of alginate and its progress in drug and protein delivery. Alginate as a potential carrier has been investigated for the delivery of a variety of low and high molecular weight drugs. Applications of alginate polymer in pharmaceutical and biomedical research have a promising future. e most important properties of alginate include safety, biocompatibility, and simple methods of preparations. is review highlights the recent advances in the alginate polymers in pharmaceutical and biomedical fields. Because of its biocompatibility, biodegradability, and nontoxicity, it is applied to various drug-delivery technologies.
us, researchers need to update the advances in the alginate-based drugdelivery systems and this review is a source of guidance for future research. Disclosure e authors alone are responsible for the content and writing of this article.

Conflicts of Interest
e authors declare no conflicts of interest.