Targeted intradermal delivery of alpha-arbutin-loaded dissolving polymeric microneedles visualized by three-dimensional Orbitrap secondary ion mass spectrometry (3D OrbiSIMS).

Hyperpigmentation, a prevalent dermatological condition characterized by melanin overproduction, poses treatment challenges due to the hydrophilicity of alpha-arbutin, a widely utilized tyrosinase inhibitor. This study investigates the efficacy of dissolving microneedles (DMNs) in augmenting skin permeation for alpha-arbutin delivery to the targeted epidermal site. Porcine full-thickness skin was employed in a 24-hour Franz cell study, commencing with the assessment of commercial alpha-arbutin-containing products. Solid steel microneedles (CMNs) from Dermapen® were utilized as both pre- and post-treatment modalities to evaluate the influence of different applications on alpha-arbutin delivery. Additionally, alpha-arbutin-loaded polyvinylpyrrolidone-co-vinyl acetate (PVPVA) DMNs, containing 2 % w/w alpha-arbutin, were fabricated and examined for their permeation-enhancing capabilities. HPLC analysis and 3D Orbitrap Secondary Ion Mass Spectrometry (OrbiSIMS) were employed to quantify and visualize alpha-arbutin in various Franz cell components. Results indicate that alpha-arbutin permeation to the skin was restricted (less than 1 %) without microneedle application and significantly increased by 6-fold (4-5 %) with post-treatment CMNs and DMNs, but not with pre-treatment CMNs. Notably, DMNs exhibited a more sustainable and robust capacity than post-treatment CMNs. OrbiSIMS imaging analysis revealed that DMNs visually enhance skin permeation of alpha-arbutin by delivering the compound to the basal layer of the targeted skin location. Overall, this study underscores the potential of DMNs as a promising delivery system for promoting targeted intradermal delivery of alpha-arbutin, providing a comprehensive exploration of various methodologies to identify innovative and improved microneedle approaches for alpha-arbutin permeation.

Distribution of lamivudine into lymph node HIV reservoir.

Efficient delivery of antiretroviral agents to lymph nodes is important to decrease the size of the HIV reservoir within the lymphatic system. Lamivudine (3TC) is used in first-line regimens for the treatment of HIV. As a highly hydrophilic small molecule, 3TC is not predicted to associate with chylomicrons and therefore should have negligible uptake into intestinal lymphatics following oral administration. Similarly, negligible amounts of 3TC are predicted to be transported into peripheral lymphatics following subcutaneous (SC) injection due to the faster flow rate of blood in comparison to lymph. In this work, we performed pharmacokinetic and biodistribution studies of 3TC in rats following oral lipid-based, oral lipid-free, SC, and intravenous (IV) administrations. In the oral administration studies, mesenteric lymph nodes (MLNs) had significantly higher 3TC concentrations compared to other lymph nodes, with mean tissue:serum ratios ranging from 1.4 to 2.9. However, cells and chylomicrons found in mesenteric lymph showed low-to-undetectable concentrations. In SC studies, administration-side (right) draining inguinal and popliteal lymph nodes had significantly higher concentrations (tissue:serum ratios as high as 3.2) than corresponding left-side nodes. In IV studies, lymph nodes had lower mean tissue:serum ratios ranging from 0.9 to 1.4. We hypothesize that following oral or SC administration, slower permeation of this hydrophilic molecule into blood capillaries may result in considerable passive 3TC penetration into lymphatic vessels. Further studies will be needed to clarify the mechanism of delivery of 3TC and similar antiretroviral drugs into the lymph nodes.

Design, Synthesis, and Biological Evaluation of Novel Hybrids Containing Dihydrochalcone as Tyrosinase Inhibitors to Treat Skin Hyperpigmentation.

Excessive melanin deposition may lead to a series of skin disorders. The production of melanin is carried out by melanocytes, in which the enzyme tyrosinase performs a key role. In this work, we identified a series of novel tyrosinase inhibitor hybrids with a dihydrochalcone skeleton and resorcinol structure, which can inhibit tyrosinase activity and reduce the melanin content in the skin. Compound 11c possessed the most potent activity against tyrosinase, showing IC50 values at nanomolar concentration ranges, along with significant antioxidant activity and low cytotoxicity. Furthermore, in vitro permeation tests, supported by HPLC analysis and 3D OrbiSIMS imaging visualization, revealed the excellent permeation of 11c. More importantly, compound 11c reduced the melanin content on UV-induced skin pigmentation in a guinea pig model in vivo. These results suggest that compound 11c may serve as a promising potent tyrosinase inhibitor for the development of a potential therapy to treat skin hyperpigmentation.

Evaluation of cannabidiol nanoparticles and nanoemulsion biodistribution in the central nervous system after intrathecal administration for the treatment of pain.

We investigated how the biodistribution of cannabidiol (CBD) within the central nervous system (CNS) is influenced by two different formulations, an oil-in-water (O/W) nanoemulsion and polymer-coated nanoparticles (PCNPs). We observed that both CBD formulations administered were preferentially retained in the spinal cord, with high concentrations reaching the brain within 10 min of administration. The CBD nanoemulsion reached Cmax in the brain at 210 ng/g within 120 min (Tmax), whereas the CBD PCNPs had a Cmax of 94 ng/g at 30 min (Tmax), indicating that rapid brain delivery can be achieved through the use of PCNPs. Moreover, the AUC0-4h of CBD in the brain was increased 3.7-fold through the delivery of the nanoemulsion as opposed to the PCNPs, indicating higher retention of CBD at this site. Both formulations exhibited immediate anti-nociceptive effects in comparison to the respective blank formulations.

Development of nanoparticle loaded microneedles for drug delivery to a brain tumour resection site.

Systemic drug delivery to the central nervous system (CNS) has been historically impeded by the presence of the blood brain barrier rendering many therapies inefficacious to any cancer cells residing within the brain. Therefore, local drug delivery systems are being developed to overcome this shortfall. Here we have manufactured polymeric microneedle (MN) patches, which can be anchored within a resection cavity site following surgical removal of a tumour such as isocitrate dehydrogenase wild type glioblastoma (GBM). These MN patches have been loaded with polymer coated nanoparticles (NPs) containing cannabidiol (CBD) or olaparib (OLA) and applied to an in vitro brain simulant and ex vivo rat brain tissue to assess drug release and distance of penetration. MN patches loaded with methylene blue dye were placed into a cavity of 0.6 % agarose to simulate brain tissue. The results showed that clear channels were generated by the MNs and the dye spread laterally throughout the agarose. When loaded with CBD-NPs, the agarose showed a CBD concentration of 12.5 µg/g at 0.5 cm from the MN insertion site. Furthermore, high performance liquid chromatography of ex vivo brain tissue following CBD-NP/MN patch insertion showed successful delivery of 59.6 µg/g into the brain tissue. Similarly, OLA-NP loaded MN patches showed delivery of 5.2 µg/g OLA into agarose gel at 0.5 cm distance from the insertion site. Orbitrap secondary ion mass spectrometry (OrbiSIMS) analysis confirmed the presence of OLA and the MN patch at up to 6 mm away from the insertion site following its application to a rat brain hemisphere. This data has provided insight into the capabilities and versatility of MN patches for use in local brain drug delivery, giving promise for future research.

The clinical and translational prospects of microneedle devices, with a focus on insulin therapy for diabetes mellitus as a case study.

Microneedles have the clinical advantage of being able to deliver complex drugs across the skin in a convenient and comfortable manner yet haven't successfully transitioned to medical practice. Diabetes mellitus is a complicated disease, which is commonly treated with multiple daily insulin injections, contributing to poor treatment adherence. Firstly, this review determines the clinical prospect of microneedles, alongside considerations that ought to be addressed before microneedle technology can be translated from bench to bedside. Thereafter, we use diabetes as a case study to consider how microneedle-based-technology may be successfully harnessed. Here, publications referring to insulin microneedles were evaluated to understand whether insertion efficiency, angle of insertion, successful dose delivery, dose adjustability, material biocompatibility and therapeutic stability are being addressed in early stage research. Moreover, over 3,000 patents from 1970 to 2019 were reviewed with the search term '"microneedle" AND "insulin"' to understand the current status of the field. In conclusion, the reporting of early stage microneedle research demonstrated a lack of consistency relating to the translational factors addressed. Additionally, a more rational design, based on a patient-centred approach is required before microneedle-based delivery systems can be used to revolutionise the lives of people living with diabetes following regulatory approval.

Development of a nanocapsule-loaded hydrogel for drug delivery for intraperitoneal administration.

Intraperitoneal (IP) drug delivery of chemotherapeutic agents, administered through hyperthermal intraperitoneal chemotherapy (HIPEC) and pressurized intraperitoneal aerosolized chemotherapy (PIPAC), is effective for the treatment of peritoneal malignancies. However, these therapeutic interventions are cumbersome in terms of surgical practice and are often associated with the formation of peritoneal adhesions, due to the catheters inserted into the peritoneal cavity during these procedures. Hence, there is a need for the development of drug delivery systems that can be administered into the peritoneal cavity. In this study, we have developed a nanocapsule (NCs)-loaded hydrogel for drug delivery in the peritoneal cavity. The hydrogel has been developed using poly(ethylene glycol) (PEG) and thiol-maleimide chemistry. NCs-loaded hydrogels were characterized by rheology and their resistance to dilution and drug release were determined in vitro. Using IVIS® to measure individual organ and recovered gel fluorescence intensity, an in vivo imaging study was performed and demonstrated that NCs incorporated in the PEG gel were retained in the IP cavity for 24 h after IP administration. NCs-loaded PEG gels could find potential applications as biodegradable, drug delivery systems that could be implanted in the IP cavity, for example at a the tumour resection site to prevent recurrence of microscopic tumours.

Detection of Label-Free Drugs within Brain Tissue Using Orbitrap Secondary Ion Mass Spectrometry as a Complement to Neuro-Oncological Drug Delivery.

Historically, pre-clinical neuro-oncological drug delivery studies have exhaustively relied upon overall animal survival as an exclusive measure of efficacy. However, with no adopted methodology to both image and quantitate brain parenchyma penetration of label-free drugs, an absence of efficacy typically hampers clinical translational potential, rather than encourage re-formulation of drug compounds using nanocarriers to achieve greater tissue penetration. OrbiSIMS, a next-generation analytical instrument for label-free imaging, combines the high resolving power of an OrbiTrapTM mass spectrometer with the relatively high spatial resolution of secondary ion mass spectrometry. Here, we develop an ex vivo pipeline using OrbiSIMS to accurately detect brain penetration of drug compounds. Secondary ion spectra were acquired for a panel of drugs (etoposide, olaparib, gemcitabine, vorinostat and dasatinib) under preclinical consideration for the treatment of isocitrate dehydrogenase-1 wild-type glioblastoma. Each drug demonstrated diagnostic secondary ions (all present molecular ions [M-H]− which could be discriminated from brain analytes when spiked at >20 µg/mg tissue. Olaparib/dasatinib and olaparib/etoposide dual combinations are shown as exemplars for the capability of OrbiSIMS to discriminate distinct drug ions simultaneously. Furthermore, we demonstrate the imaging capability of OrbiSIMS to simultaneously illustrate label-free drug location and brain chemistry. Our work encourages the neuro-oncology community to consider mass spectrometry imaging modalities to complement in vivo efficacy studies, as an analytical tool to assess brain distribution of systemically administered drugs, or localised brain penetration of drugs released from micro- or nano-scale biomaterials.

Surface-controlled spatially heterogeneous physical properties of a supramolecular gel with homogeneous chemical composition.

Controlling supramolecular self-assembly across multiple length scales to prepare gels with localised properties is challenging. Most strategies concentrate on fabricating gels with heterogeneous components, where localised properties are generated by the stimuli-responsive component. Here, as an alternative approach, we use a spiropyran-modified surface that can be patterned with light. We show that light-induced differences in surface chemistry can direct the bulk assembly of a low molecular weight gelator, 2-NapAV, meaning that mechanical gel properties can be controlled by the surface on which the gel is grown. Using grazing incidence X-ray diffraction and grazing incidence small angle X-ray scattering, we demonstrate that the origin of the different gel properties relates to differences in the architectures of the gels. This provides a new method to prepare a single domain (i.e., chemically homogeneous) hydrogel with locally controlled (i.e., mechanically heterogeneous) properties.

Biomedical engineering approaches to enhance therapeutic delivery for malignant glioma.

We review the challenges of next-generation therapeutics for both systemic and localised delivery to brain tumours and discuss how recent engineering advances may be used to enhance brain penetration of systemic delivery therapies. The unmet clinical need which drug delivery seeks to address is discussed with reference to the therapy obstacles that the intra-tumour heterogeneity of glioma present. The unmet chemistry and biomedical engineering challenge to develop controlled release therapeutics is appraised, with commentary on current success/failures in systemic carrier-mediated delivery, including receptor-targeted, cell-based, blood-brain-barrier disrupting and MRI-guided focused ultrasound. Localised therapeutic delivery is a relatively under-studied research avenue and is discussed with reference to existing technologies in preclinical development. These include convection-enhanced delivery, alternative catheter delivery, and neuro-surgically applied delivery systems such as polymeric hydrogels and interstitial spray. A myriad of nano-scale therapeutic delivery systems is emerging as potential future medicines for malignant brain tumours. Such biomedically-engineered systems will increasingly feature in next-generation neuro-oncological clinical trials to deliver repurposed and experimental therapeutics, aimed at achieving therapeutic drug concentrations in the brain, with associated mortality and morbidity benefits for patients.

Etoposide and olaparib polymer-coated nanoparticles within a bioadhesive sprayable hydrogel for post-surgical localised delivery to brain tumours.

Glioblastoma is a malignant brain tumour with a median survival of 14.6 months from diagnosis. Despite maximal surgical resection and concurrent chemoradiotherapy, reoccurrence is inevitable. To try combating the disease at a stage of low residual tumour burden immediately post-surgery, we propose a localised drug delivery system comprising of a spray device, bioadhesive hydrogel (pectin) and drug nanocrystals coated with polylactic acid-polyethylene glycol (NCPPs), to be administered directly into brain parenchyma adjacent to the surgical cavity. We have repurposed pectin for use within the brain, showing in vitro and in vivo biocompatibility, bio-adhesion to mammalian brain and gelling at physiological brain calcium concentrations. Etoposide and olaparib NCPPs with high drug loading have shown in vitro stability and drug release over 120 h. Pluronic F127 stabilised NCPPs to ensure successful spraying, as determined by dynamic light scattering and transmission electron microscopy. Successful delivery of Cy5-labelled NCPPs was demonstrated in a large ex vivo mammalian brain, with NCPP present in the tissue surrounding the resection cavity. Our data collectively demonstrates the pre-clinical development of a novel localised delivery device based on a sprayable hydrogel containing therapeutic NCPPs, amenable for translation to intracranial surgical resection models for the treatment of malignant brain tumours.

Intradermal delivery of imiquimod using polymeric microneedles for basal cell carcinoma.

Despite being one of the most efficacious drugs used in the treatment of basal cell carcinoma (BCC), imiquimod has limited cutaneous permeation. The current work presents the development of polyvinylpyrrolidone-co-vinyl acetate (PVPVA) microneedles loaded with imiquimod for improving intradermal delivery of imiquimod for the treatment of nodular BCC. In vitro permeation studies, using full thickness ex vivo porcine skin were used to evaluate the effectiveness of these imiquimod loaded polymeric microneedles in comparison to the topical application of commercial Aldara™ cream. HPLC analysis demonstrated similar intradermal permeation of imiquimod from Aldara™ cream and imiquimod-loaded microneedles despite the microneedle having a six-fold lower drug loading than the clinical dose of Aldara™ used for BCC management. In addition, ToF-SIMS analysis of skin cross sections demonstrated intradermal localisation of imiquimod following microneedle-based delivery while the Aldara™ treated skin showed the drug localised predominantly within the stratum corneum. ToF-SIMS analysis also demonstrated intradermal co-localisation of the PVPVA polymer, used in fabricating the microneedle, with imiquimod within the microneedle channels in a label-free manner. This study demonstrates that a polymeric microneedle system may be a viable approach to improving the intradermal delivery of imiquimod for the treatment of nodular BCC with lower drug loading.

Intradermal Delivery of an Immunomodulator for Basal Cell Carcinoma; Expanding the Mechanistic Insight into Solid Microneedle-Enhanced Delivery of Hydrophobic Molecules.

Basal cell carcinoma (BCC) is the most common cutaneous malignancy in humans. One of the most efficacious drugs used in the management of BCC is the immunomodulator, imiquimod. However, imiquimod has physiochemical properties that limit its permeation to reach deeper, nodular tumor lesions. The use of microneedles may overcome such limitations and promote intradermal drug delivery. The current work evaluates the effectiveness of using an oscillating microneedle device Dermapen either as a pre- or post-treatment with 5% w/w imiquimod cream application to deliver the drug into the dermis. The effectiveness of microneedles to enhance the permeation of imiquimod was evaluated ex vivo using a Franz cell setup. After a 24-h permeation experiment, sequential tape strips and vertical cross-sections of the porcine skin were collected and analyzed using time-of-flight secondary ion mass spectrometry (ToF-SIMS). In addition, respective Franz cell components were analyzed using high-performance liquid chromatography (HPLC). Analysis of porcine skin cross-sections demonstrated limited dermal permeation of 5% w/w imiquimod cream. Similarly, limited dermal permeation was also seen when 5% w/w imiquimod cream was applied to the skin that was pretreated with the Dermapen, this is known as poke-and-patch. In contrast, when the formulation was applied first to the skin prior to Dermapen application, this is known as patch-and-poke, we observed a significant increase in intradermal permeation of imiquimod. Such enhancement occurs immediately upon microneedle application, generating an intradermal depot that persists for up to 24 h. Intradermal colocalization of isostearic acid, an excipient in the cream, with imiquimod within microneedle channels was also demonstrated. However, such enhancement in intradermal delivery of imiquimod was not observed when the patch-and-poke strategy was used with a non-oscillating microneedle applicator, the Dermastamp. The current work highlights that using the patch-and-poke approach with an oscillating microneedle pen may be a viable approach to improve the current treatment in BCC patients who would prefer a less invasive intervention relative to surgery.

Intradermal and transdermal drug delivery using microneedles - Fabrication, performance evaluation and application to lymphatic delivery.

The progress in microneedle research is evidenced by the transition from simple 'poke and patch' solid microneedles fabricated from silicon and stainless steel to the development of bioresponsive systems such as hydrogel-forming and dissolving microneedles. In this review, we provide an outline on various microneedle fabrication techniques which are currently employed. As a range of factors, including materials, geometry and design of the microneedles, affect the performance, it is important to understand the relationships between them and the resulting delivery of therapeutics. Accordingly, there is a need for appropriate methodologies and techniques for characterization and evaluation of microneedle performance, which will also be discussed. As the research expands, it has been observed that therapeutics delivered via microneedles has gained expedited access to the lymphatics, which makes them a favorable delivery method for targeting the lymphatic system. Such opportunity is valuable in the area of vaccination and treatment of lymphatic disorders, which is the final focus of the review.

Mechanistic investigations into the encapsulation and release of small molecules and proteins from a supramolecular nucleoside gel in vitro and in vivo.

Supramolecular gels have recently emerged as promising biomaterials for the delivery of a wide range of bioactive molecules, from small hydrophobic drugs to large biomolecules such as proteins. Although it has been demonstrated that each encapsulated molecule has a different release profile from the hydrogel, so far diffusion and steric impediment have been identified as the only mechanisms for the release of molecules from supramolecular gels. Erosion of a supramolecular gel has not yet been reported to contribute to the release profiles of encapsulated molecules. Here, we use a novel nucleoside-based supramolecular gel as a drug delivery system for proteins with different properties and a hydrophobic dye and describe for the first time how these materials interact, encapsulate and eventually release bioactive molecules through an erosion-based process. Through fluorescence microscopy and spectroscopy as well as small angle X-ray scattering, we show that the encapsulated molecules directly interact with the hydrogel fibres - rather than being physically entrapped in the gel network. The ability of these materials to protect proteins against enzymatic degradation is also demonstrated here for the first time. In addition, the released proteins were proven to be functional in vitro. Real-time fluorescence microscopy together with macroscopic release studies confirm that erosion is the key release mechanism. In vivo, the gel completely degrades after two weeks and no signs of inflammation are detected, demonstrating its in vivo safety. By establishing the contribution of erosion as a key driving force behind the release of bioactive molecules from supramolecular gels, this work provides mechanistic insight into the way molecules with different properties are encapsulated and released from a nucleoside-based supramolecular gel and sets the basis for the design of more tailored supramolecular gels for drug delivery applications.

Expanding the applications of microneedles in dermatology.

Since the first patent for microneedles was filed in the 1970s, research on utilising microneedles as a drug delivery system has progressed significantly. In addition to the extensive research on microneedles for improving transdermal drug delivery, there is a growing interest in using these devices to manage dermatological conditions. This review aims to provide the background on microneedles, the clinical benefits, and challenges of the device along with the potential dermatological conditions that may benefit from the application of such a drug delivery system. The first part of the review provides an outline on benefits and challenges of translating microneedle-based drug delivery systems into clinical practice. The second part of the review covers the application of microneedles in treating dermatological conditions. The efficacy of microneedles along with the limitations of such a strategy to treat diseased skin shall be addressed.

A novel low molecular weight nanocomposite hydrogel formulation for intra-tumoural delivery of anti-cancer drugs.

Herein, an injectable formulation composed of a low molecular weight gelator (LMWG) based hydrogel and drug-loaded polymeric nanocapsules (NCs) is described. The NCs, made of hyaluronic acid and polyglutamic acid and loaded with C14-Gemcitabine (GEM C14), showed a size of 40 and 80 nm and a encapsulation efficiency >90%. These NCs exhibited a capacity to control the release of the encapsulated drug for >1 month. GEM C14-loaded NCs showed activity against various cancer cell lines in vitro; cell growth inhibition by 50% (GI50) values of 15 ±â€¯6, 10 ±â€¯9, 13 ±â€¯3 and 410 ±â€¯463 nM were obtained in HCT 116, MIA PaCa-2, Panc-1 and Panc-1 GEM resistant cell lines respectively. Nanocomposite hydrogels were prepared using the LMWG - N4-octanoyl-2'-deoxycytidine and loaded for the first time with polymeric NCs. 2% and 4% w/v nanocapsule concentrations as compared to 8% w/v NC concentrations with 2% and 3% w/v gelator concentrations gave mechanically stronger gels as determined by oscillatory rheology. Most importantly, the nanocomposite formulation reformed instantly into a gel after injection through a needle. Based on these properties, the nanocomposite gel formulation has potential for the intratumoural delivery of anticancer drugs.

Insight into imiquimod skin permeation and increased delivery using microneedle pre-treatment.

Basal cell carcinoma (BCC) is the most common skin cancer in humans. Topical treatment with imiquimod provides a non-invasive, self-administered treatment with relatively low treatment cost. Despite displaying excellent efficacy, imiquimod is only licensed by the FDA for superficial BCC. The current work employed HPLC and ToF-SIMS analysis to provide a novel assessment of imiquimod permeation from Aldara™ cream in skin depth and lateral distribution. Using Aldara™ cream and in vitro Franz cell studies with subsequent HPLC analysis, it is apparent that most of the topically applied imiquimod cream is left on the skin surface with more than 80% of the drug being recovered from skin wash. In addition, ToF-SIMS chemical imaging of recovered tape stripped skin samples illustrated significant detection of imiquimod signal over the entire skin area for the upper tape strips, whereas the deeper strips show large portions of the skin area without detected imiquimod. Given the limited permeation depth and non-uniform permeation observed at tape strips 6-18 when applied as a topical imiquimod cream, a permeation enhancement strategy utilising a skin pre-treatment with a microneedle device was investigated as a method to improve intradermal delivery. The recovered amount of imiquimod in tape strips and remaining skin determined by HPLC was approximately three times higher when Aldara™ was applied on microneedle pre-treated skin relative to intact skin. The ToF-SIMS ion images of the tape strips and cross-sections illustrated the existence of imiquimod in the microchannels which then laterally diffuses to peripheral epidermal strata. The current work demonstrates the first known attempt to enhance intradermal delivery of imiquimod using a microneedle device as well as underscoring the complementary role of ToF-SIMS analysis in chemically mapping imiquimod permeation into the skin with high sensitivity.

Low Molecular Weight Nucleoside Gelators: A Platform for Protein Aggregation Inhibition.

Low molecular weight nucleoside gelators hold great promise in drug delivery and particularly for the delivery of biologics because of their excellent biocompatibility. However, the influence of these gelators on protein aggregation inhibition has not yet been studied. Protein aggregation is the most significant cause of protein instability and can severely impact the biological activity of the protein, impairing the quality and safety of the formulation. Herein, we report the ability of a nucleoside-based gelator, N4-octanoyl-2'-deoxycytidine, to inhibit protein aggregation. Using turbidimetric, spectroscopic, and microscopic methods, we demonstrate that protein aggregation inhibition is dependent on gelator concentration. Moreover, we have found that the protein is still functionally active in the hydrogel.