Method for manufacturing a laboratory object suitable for conducting experiments with pharmaceutical or cosmetic active ingredients

Abstract

The invention relates to a method for manufacturing a laboratory object suitable for conducting experiments with pharmaceutical or cosmetic active ingredients without same being absorbed or adsorbed into the surface of the object. The method comprises 3D printing, coating the objects and drying. More specifically, the invention relates to a diffusion cell for in vitro and ex vitro testing.

Description

[0001] DESCRIPTION

[0002] Method of manufacturing a laboratory object suitable for carrying out experiments with pharmaceutical or cosmetic active ingredients

[0003] TECHNICAL SECTOR

[0004] The invention relates to a method of manufacturing a laboratory object suitable for carrying out experiments with pharmaceutical or cosmetic active ingredients, preventing them from being absorbed or adsorbed to its surface.

[0005] BACKGROUND OF THE INVENTION

[0006] In the research of new active molecules and in the development of medicines and cosmetics, in vitro studies are necessary to predict in vivo behavior and to establish in vitro-in vivo correlations. During preformulation development, in vitro studies make it possible to discard formulations that do not meet optimal requirements and, more importantly, correctly conducted in vitro studies make it possible to reduce the number of animals needed to test new formulations in future in vivo studies, helping to comply with the 3Rs principle in animal testing: REPLACEMENT, REDUCTION, REFINEMENT. Furthermore, cosmetic product legislation prohibits animal testing, making it necessary to replace it with in vitro or ex vivo tests.

[0007] The most commonly used objects for in vitro assays of drug and cosmetic release, diffusion and penetration through biological and synthetic membranes are the so-called Franz glass cells and side-by-side cells consisting mainly of a donor part (donor chamber) and a recipient part (receiver chamber) separated by a membrane or tissue.

[0008] Furthermore, it is important to have different types of diffusion cells available depending on the formulation and intended route of administration, with different shapes to adapt to the desired membrane or tissue type, sample volume, receptor medium, and agitation type. For example, some studies have focused on developing models that mimic the physiology of the eye or skin for in vitro modeling. Therefore, the manufacture of more robust, affordable, and customized diffusion cells and other objects that meet the same pharmaceutical requirements as currently marketed glass cells is an advantage. Cells with different shapes and adapted to the needs of the experiment can be prepared using 3D printing. However, the main problem with this technology lies in the use of photocurable resins, composed primarily of acrylates.Acrylic materials are permeable to various substances, including solvents and solutes (e.g., pharmaceuticals). Furthermore, these materials can absorb water and other solvents and, depending on their degree of crosslinking, can even swell. As a result, drug adsorption and absorption problems can arise when they come into contact with a pharmaceutical formulation.

[0009] Thus, it has been shown that these types of acrylic cells adsorb the drugs with which the studies are carried out, so the result of the experiment is distorted, unreliable, robust, and useful for the intended purpose. In Sil BC, Alvarez MP, Zhang Y, Kung CP, Hossain M, Lliopoulos F, et al. 3D-printed Franz type diffusion cells. Int J Cosmet Sci. 2018 Dec;40(6):604-9, an acrylic resin is used in the process of forming the diffusion cell pieces, using the 3D printing technique by stereolithography. The results obtained using different active ingredients show that in all cases there is a significant retention of the drugs and even the use of hydrophobic coatings were ineffective in reducing drug retention.

[0010] In a subsequent study (Sil BC, Belgrave RG, Alvarez MP, Luo L, Cristofoli M, Penny MR, et al. 3D-Printed Franz cells - update on optimization of manufacture and evaluation. International Journal of Cosmetic Science. 2020;42(4):415-9), caffeine was used as a hydrophilic model drug in uncoated 3D printed objects. Although the use of this hydrophilic drug improved the compatibility with acrylic resin, hydrophobic compounds still do not show compatibility with the proposed model.

[0011] For these reasons, a solution is still needed that allows diffusion objects of different shapes, ad hoc, designed according to the needs of the experiment and that do not absorb or adsorb drugs and active substances on their surface.

[0012] BRIEF DESCRIPTION OF THE INVENTION

[0013] The authors of the present invention have found that by manufacturing a laboratory object using 3D printing and then coating it using the method described in the invention, they ensure that said object does not absorb active pharmaceutical ingredients or adsorb them on its surface. Furthermore, said materials, coated using the method described in the invention, do not absorb or adsorb hydrophilic or hydrophobic active pharmaceutical ingredients. The laboratory objects of the present invention confer superior advantages even compared to glass, the material traditionally used in laboratory objects. This is demonstrated in Example 2, where bevacizumab, known for its interaction with glass, does not interact with the objects of the present invention.

[0014] Thus, in a first aspect, the invention relates to a laboratory object that is characterized by being formed by an acrylic resin coated with a Bisphenol.

[0015] In a particular embodiment, the invention relates to a laboratory object consisting of an acrylic resin coated with a Bisphenol.

[0016] In a preferred embodiment, the laboratory object is a diffusion cell.

[0017] In a second aspect, the invention relates to a method of manufacturing a laboratory object, said laboratory object being suitable for carrying out experiments with pharmaceutical or cosmetic active ingredients, preventing them from being absorbed or adsorbed to its surface, comprising: i. 3D printing of the object,

[0018] i. coating of the printed objects using a solution comprising Bisphenol, iii. drying of the applied coating.

[0019] In a third aspect, the invention relates to the use of a laboratory object according to the present invention for performing in vitro or ex vivo tests in the presence of active pharmaceutical or cosmetic ingredients. In particular, to prevent the absorption and adsorption of active pharmaceutical or cosmetic ingredients on the surface.

[0020] DESCRIPTION OF THE FIGURES

[0021] Figure 1. 3D models of diffusion cells and vials designed using CAD software from measurements of actual glass diffusion cells. Figure A is a model of a vial similar to those used in chemistry and pharmacy laboratories. Figure B is a model of a vertical Franz diffusion cell. Figure C is a model of a side-by-side diffusion cell.

[0022] Figure 2. Photographs of diffusion cells and vials fabricated by stereolithography 3D printing and with coating. Figure A is a vertical Franz diffusion cell. Figure B is a side-by-side diffusion cell. Figure C is a vial similar to those used in chemistry and pharmacy laboratories. Figure 3. SEM (scanning electron microscopy) images of (A) the uncoated resin surface, (B) the coated resin surface, and (C) the side view of the opaque Bisphenol A coating applied to the resin material.

[0023] Figure 4. XDR diffractograms of the surface of (A) uncoated objects and (B) coated with opaque coating comprising 10-25% (w / v) Bisphenol with fillers.

[0024] Figure 5. Raman spectrum of the surface of (A) the objects coated with opaque coating comprising 10-25% (w / v) of Bisphenol with fillers, and (B) the uncoated ones.

[0025] Figure 6. XDR diffractograms of the washed coated objects and the opaque coating comprising 10-25% (w / v) Bisphenol with bulking agents. (A) Dexamethasone base - washed with neutral detergent, (B) Dexamethasone base - washed with neutral detergent and ethanol, (C) Hydroxocobalamin - washed with neutral detergent, (D) Hydroxocobalamin - washed with neutral detergent and ethanol, (E) Voriconazole - washed with neutral detergent, (F) Voriconazole - washed with neutral detergent and ethanol, (G) Insulin - washed with neutral detergent, (H) Insulin - washed with neutral detergent and ethanol (I) Bevacizumab - washed with neutral detergent, (J) Bevacizumab - washed with neutral detergent and ethanol, (K) coating.

[0026] Figure 7. Raman spectrum of the surface of washed coated objects and of the opaque coating comprising 10-25% (w / v) Bisphenol with fillers, and of the uncoated ones. (A) Dexamethasone base - washed with neutral detergent, (B) Dexamethasone base - washed with neutral detergent and ethanol, (C) Hydroxocobalamin - washed with neutral detergent, (D) Hydroxocobalamin - washed with neutral detergent and ethanol, (E) Voriconazole - washed with neutral detergent, (F) Voriconazole - washed with neutral detergent and ethanol, (G) Insulin - washed with neutral detergent, (H) Insulin - washed with neutral detergent and ethanol (I) Bevacizumab - washed with neutral detergent, (J) Bevacizumab - washed with neutral detergent and ethanol, (K) coating.

[0027] Figure 8. SEM (scanning electron microscopy) images of the surface of the opaque coating comprising 10-25% (w / v) Bisphenol with fillers after exposure to (A) liquid paraffin, (B) acetone, (C) ethanol, (D) isopropanol, (E) neutral detergent, (F) methanol.

[0028] Figure 9. SEM (scanning electron microscopy) images of (A) the resin surface coated with the opaque coating comprising 10-25% (w / v) Bisphenol combined with fillers to induce opacity, (B) the resin surface coated with the transparent coating comprising 10-25% (w / v) Bisphenol, and (C) the side view of the transparent coating applied to the resin material.

[0029] Figure 10. XDR diffractogram of (A) the opaque coating surface and (B) the transparent coating.

[0030] Figure 11. Raman spectrum of the surface of (A) the opaque coating and (B) the transparent coating.

[0031] DETAILED DESCRIPTION OF THE INVENTION

[0032] 3D printing of the object

[0033] In this invention, the term "3D printing" refers to additive manufacturing technologies, specifically three-dimensional (3D) printing. Three-dimensional printing has proven to be an excellent tool for the production of replacement parts and small pieces of laboratory equipment. The affordability and design flexibility offered by this technology, along with the wide variety of low-cost materials that can be used, make 3D printers essential in any research laboratory.

[0034] The advantage of 3D printing is that it allows for the production of objects in different shapes, adapted and customized to the needs of laboratory experimentation.

[0035] Thus, the method of the invention also comprises a step prior to step i, which is the design of the laboratory object using CAD software. In this design phase, the person skilled in the art can design the object with the appropriate shape and size for the experiment to be performed, for example, an in vitro or ex vivo experiment in the presence of active pharmaceutical ingredients.

[0036] In a preferred embodiment, the laboratory object is a diffusion cell.

[0037] A "diffusion cell" refers to a laboratory object used in the preformulation stage that allows for in vitro or ex vivo diffusion studies of pharmaceutical or cosmetic formulations. The most common diffusion cells are Franz cells and side-by-side cells, but other types exist. These cells consist of two compartments: a donor compartment, in which the sample is placed, and a recipient compartment, separated by a membrane or living tissue. The compartments can be arranged vertically (vertical diffusion cells), with the donor compartment placed above the recipient compartment, where the medium is agitated, or horizontally (side-by-side cells), with both compartments arranged horizontally and both agitated.The different parameters of diffusion cells, for example, cell type and shape, size of the donor and recipient compartments, type of agitation, recipient medium, sampling time, membrane or tissue used, or orifice diameter, are selected according to the type of study and the route of administration to be tested.

[0038] Among the available 3D printing technologies, stereolithography (SLA) 3D printing creates objects layer by layer through the selective solidification of a liquid resin composed of monomers and a photoinitiator. This technology is well-suited for 3D printing small parts with complex geometries at very high resolutions (< 10 µm). This allows for the manufacture of objects with very tight layer heights and no gaps between them, creating completely solid objects. This prevents the risk of leaks when liquids are introduced into the object, which is very relevant if the object to be printed is a diffusion cell.Furthermore, unlike other 3D printing techniques such as Fused Deposition Modeling (FDM), where a molten thermoplastic polymer is deposited layer by layer, stereolithography 3D printing is a scalable technology in both time and production, since it takes the same amount of time to print one object as it does to print several.

[0039] Thus, in a preferred embodiment, the method of the invention employs stereolithographic 3D printing. And in a more preferred embodiment, in step i of the method of the invention, stereolithographic 3D printing is used to prepare a diffusion cell.

[0040] In a more particular embodiment of the invention, 3D printing by stereolithography uses liquid resins composed of acrylic monomers and mixtures thereof with functional monomers, crosslinking agents, and polymerization photoinitiators. More preferably, resins made of acrylic materials that solidify selectively upon application of ultraviolet or visible light at a wavelength determined by the activation wavelength of the photoinitiator.

[0041] Using the 3D printing method described in this invention, translucent laboratory objects are obtained.

[0042] Coating of the object

[0043] In the present invention, the term “coating” or “coating” refers to the application of a material to the surface of 3D-printed objects during step i, which will be exposed in experiments with active pharmaceutical or cosmetic ingredients.

[0044] The coating, according to the present invention, comprises a bisphenol dissolved in a highly volatile organic solvent to accelerate the coating process. The coating has properties that include abrasion resistance, organic solvent resistance, pH resistance, temperature resistance, reusability, compatibility with various materials used in 3D printing, inertness, translucent, or opaque.

[0045] In a particular embodiment of the method of the invention, a solution comprising Bisphenol is used in the coating of step 1i; more particularly, the solution has a Bisphenol percentage of between 1% and 75% w / v. More preferably, between 2% and 70%, even more preferably between 2.5% and 65%.

[0046] In the present invention, the term "Bisphenol" refers to an organic chemical compound with two phenol functional groups linked by a C1-C4 alkyl group, which may be branched with alkyl groups. In a more preferred embodiment, the two phenyl groups are linked by an isopropylidene group. The term Bisphenol also includes derivatives in which the hydroxyl groups of the phenyls are substituted, forming ether groups with alkyl chains or glycidyl chains, which in their terminal position may be substituted by different groups such as a methacrylic group. The term Bisphenol also includes block copolymers in which there is a structure of two phenyl groups linked by an isopropylidene group, which may be substituted on the hydroxyl groups of the phenols.

[0047] Preferably, the Bisphenol is selected from the group consisting of glycidyl-terminal Poly(Bisphenol A-co-epichlorohydrin) copolymer, Bisphenol A-diglycidylether methacrylate (bis-GMA), 2,2-bis(4-(2-methacryloxyprop-1-oxy)phenyl) propane (CHsBis-GMA).

[0048] In another particular embodiment, the solution is prepared with a solvent selected from ethyl acetate, dichloromethane, trichloromethane, toluene, xylene, ethylbenzene, butanone, 4-hydroxy-4-methyl-2-pentanone and diacetone alcohol.

[0049] In another particular embodiment, the coating method is selected from among dip coating, air atomization coating, high-volume, low-pressure gun coating, and spray gun coating. Using any of these techniques, a homogeneous application of the solution to all points on the surface of the object is achieved.

[0050] Drying

[0051] After the coating step, the object is dried. Thus, in a particular embodiment of the invention, the drying step can be carried out by convection drying, room temperature drying, or vacuum drying. After drying, the object obtained by the method of the present invention is suitable for preventing the adsorption and absorption of drugs or bioactive substances or cosmetics when used for in vitro or ex vivo testing.

[0052] In a particular embodiment, the invention relates to the use of a laboratory object obtainable according to the invention, or of the diffusion cell according to the invention, for carrying out in vitro or ex vivo tests in the presence of pharmaceutical or cosmetic active ingredients.

[0053] In a particular embodiment, the invention relates to the use of a laboratory object obtainable according to the invention, or of the diffusion cell according to the invention, to prevent the absorption and adsorption of pharmaceutical or cosmetic active ingredients on the surface.

[0054] EXAMPLES

[0055] MATERIALS AND METHODS

[0056] 405 nm Clear UV resin supplied by Anycubic (Shenzhen, China). Ciclopirox olamine and dexamethasone supplied by Acofarma® (Barcelona, ​​Spain). Voriconazole purchased from Normon® (Madrid, Spain). Bevacizumab (Avastin®) purchased from Roche (Basel, Switzerland). Insulin supplied by Merck (Darmstadt, Germany). Hydroxocobalamin acetate supplied by HealthTech BioActives (Barcelona, ​​Spain). 2-Hydroxypropyl-p-cyclodextrin (HPpCD), Kleptose®, degree of substitution 0.65 M Molecular Mass 1399 Da) supplied by Roquette® Laisa SA (Valencia, Spain). Hyaluronic acid (HA) obtained from Acofarma® (Barcelona, ​​Spain).

[0057] For the bisphenol-based coating, several varieties of TKROM glass 2C epoxy primer by TKROM® (Murcia, Spain) were used in some cases. The white primer, which is opaque, contains fillers, and the transparent primer, which does not. Bisphenol is present in a proportion between 10 and 25% in both varieties, depending on the manufacturer. In other cases, a solution was prepared with glycidyl-terminated poly(Bisphenol A-co-epichlorohydne), supplied by Sigma® (Misuh, USA).

[0058] Example 1. Retention test of dexamethasone base and ciclopirox olamine using coatings of different nature

[0059] A 25 pg / mL dexamethasone base solution (pH 7.25) and a 50 pg / mL ciclopirox olamine solution (pH 8.5) were prepared. Vials were printed by stereolithography from the model in Figure 1A, using Clear UV Resin (Anycubic). The vials were then coated with the following materials: polytetrafluoroethylene (PTFE)-based coating, Plioway® resin-based coating, self-crosslinking acrylic elastic emulsion coating, epoxy-polyamidoamine resin-based coating, and opaque Bisphenol epoxy resin-based coating. All coatings were applied according to the manufacturer's specifications. The PTFE coating was spray-applied onto the surface of the printed vials.In the case of bisphenol-A epoxy resin coatings, prior to application, the coating component was mixed with the catalyst in the proportions recommended by the manufacturer. The remaining coatings were applied as supplied by the manufacturers. All coatings, except the PTFE coatings, were applied using the immersion technique. To do this, the vials were left in contact with the coatings for 30 minutes, and then the excess material was removed. All vials were left to dry for 48 hours at room temperature.

[0060] After the coatings were dry, the coated, uncoated and glass vials were filled with the dexamethasone or ciclopirox olamine solution and kept in the absence of light at 4°C to prevent evaporation. Drug retention was evaluated compared to uncoated vials and type I borosilicate glass vials by measuring the drug concentration at different time points (1, 2, 5 and 8 days) by ultra-high performance liquid chromatography (UHPLC) (ACQUITY UPLC H-Class Plus, Waters, Milford, Massachusetts, USA). Drug recovery was calculated according to the equation:

[0061] _. , [Drug] in coated vials. > > / r - . .

[0062] Drug recovery (%) = — - - - — — : — — - 100 (Equation 1)

[0063] [Drug] in glass vials

[0064] Table 1 shows the recovery values ​​for dexamethasone base and ciclopirox olamine. It can be seen that only the opaque coating based on bisphenol epoxy resin maintains the initial concentration of both drugs. A significant decrease is observed in the other drugs, which is more pronounced in the case of ciclopirox olamine.

[0065] Table 1a. Mean recovery values ​​(%) and standard deviation (SD) for dexamethasone and ciclopirox olamine with each coating (n=3).

[0066] Table 1b. Mean recovery values ​​(%) and standard deviation (SD) for dexamethasone and ciclopirox olamine with each coating (n=3). Example 2. Retention test of different drugs with 3D printed vials coated with opaque coating based on Bisphenol epoxy resin

[0067] Vials were designed using CAD software and manufactured by 3D printing using a stereolithography 3D printer using Clear UV Resin (Anycubic). The solution for the application of the opaque Bisphenol coating was prepared according to the manufacturer's specifications. For this purpose, the two components A and B supplied were mixed. Component A is composed of 4,4'-lsopropylidenediphenol, a polymer with 2,2-Bis(p-(2,3-Epoxypropoxy¡)Phen¡l)Propane, reaction mass of ethylbenzene and ethylbenzene, xylene, hydrocarbons, C9, aromatics, methyl propanol, methoxymethylethyl acetate, butyl acetate, fatty acids, formaldehyde and maleic anhydride. The catalyst or component B is composed of mixtures of fatty acids, C18-unsaturated, dimers, oligomeric reaction products with fatty acid oil, triethylenetetramine dissolved in a mixture of xylene, 2-methoxy-1-methylethyl acetate and butanol.The bisphenol-A epoxy resin base was prepared by mixing components A and B in a 4:1 ratio, with a final bisphenol content of 10–25% (w / v). Subsequently, a portion of the 3D-printed vials were coated by immersion in the prepared coating solution, while the remaining portion was left uncoated. The retention of different drugs with distinct physicochemical properties over 8 days was compared between the coated cylindrical vial group and the borosilicate type I glass vial group, expressed in terms of drug recovery according to equation 1.

[0068] Solutions of 25 pg / mL dexamethasone base (pH 7.25), 50 pg / mL ciclopirox olamine (pH 8.5), 100 pg / mL human insulin (pH 2.3), 25 pg / mL bevacizumab (pH 6.15), 50 pg / mL voriconazole with 0.1% (w / v) hydroxypropyl-p-cyclodextrin (pH 7.12) and 50 pg / mL hydroxycobalamin acetate (pH 5.71) were prepared.

[0069] The containers corresponding to the different groups were filled with the corresponding solutions, covered with a Parafilm® M sealing film, and kept at 4°C to prevent evaporation. Drug retention was evaluated compared to uncoated vials and borosilicate type I glass vials by measuring the drug concentration at different time points (1, 2, 5, and 8 days) using ultrahigh-performance liquid chromatography (UHPLC). The results are shown in Table 2.

[0070] Table 2. Results of average recovery values ​​(%) of different drugs in the cylindrical containers made by 3D printing using Clear UV Resin (Anycubic) after coating them with an opaque coating based on Bisphenol epoxy resin (n=3).

[0071] No significant differences were observed in the concentrations recovered at different times for hydroxocobalamin acetate, voriconazole, ciclopirox olamine, dexamethasone base, and insulin (α ns). However, for bevacizumab, retention in glass vials was higher compared with coated objects (α <0.05). This is because adsorption and absorption did not occur on coated printed objects, but did occur on the Type I glass vials used as a reference. Different works have been published describing the retention of proteins and monoclonal antibodies in glass containers, as in our example (Wei Q, Becherer T, Angioletti-Uberti S, Dzubiella J, Wischke C, Neffe AT, et al. Protein Interactions with Polymer Coatings and Biomaterials. Angewandte Chemie International Edition. 2014;53(31):8004-31), (Hoehne M, Samuel F, Dong A, Wurth C, Mahler HC, Carpenter JF, et al.Adsorption of monoclonal antibodies to glass microparticles. Journal of Pharmaceutical Sciences. 2011;100(1):123— 32). Consequently, the Bisphenol coating demonstrates the potential to prevent the adsorption or absorption of the different drugs tested by the acrylic material that makes up the 3D printed objects. Specifically, this experiment shows that it is capable of preventing the surface adsorption of the protein-based active ingredients insulin and bevacizumab, which are known for their high adsorption to glass.

[0072] Example 3. Coatability testing of surfaces of Bisphenol-based coatings Cylindrical vials were designed using CAD software according to the model in Figure 1A and fabricated by 3D printing using a stereolithography 3D printer and Clear UV Resin (Anycubic). The 3D printed vials were subsequently coated by dipping method with an opaque coating comprising 10-25% (w / v) Bisphenol combined with fillers to induce opacity and another group of vials were left uncoated.

[0073] The coating capacity was determined by SEM (scanning electron microscopy) (Figure 3) combined with energy dispersive X-ray analysis (Table 3), X-ray diffraction (XRD) (Figure 4) and Raman spectroscopy (Figure 5). The results show good and complete coating capacity.

[0074] Table 3. Surface elemental analysis of the resin used in 3D printing and the opaque coating.

[0075] Example 4. Permeation assay of a voñconazole eye drop comparing 3D printed Franz cells with the coating and glass Franz cells

[0076] A Franz cell was designed using CAD software with the same dimensions as a conventional glass Franz cell (Figure 1 B). It was 3D printed using a stereolithography 3D printer, and an opaque coating comprising 10–25% (w / v) Bisphenol combined with fillers to induce opacity was dip-applied. The printed objects were convectively dried for 24 h and used in the experiments.

[0077] To compare the suitability of 3D printed coated Franz cells and conventional glass Franz cells for in vitro drug release testing, an in vitro release study was conducted using a voriconazole eye drop. The eye drop was prepared by adding 0.4% (w / v) hyaluronic acid to a 1% (w / v) voriconazole solution containing 20% ​​(w / v) hydroxypropyl-p-cyclodextrin. 0.5 mL of formulation was added to the donor compartment, while the recipient compartment was filled with 6 mL of phosphate-buffered saline (PBS) with a pH of 7.4. Visking® dialysis membranes with a cut-off limit of 12–14 KDa (with an available surface area of ​​0.784 cm 2) between the donor and recipient compartments. Franz cells were maintained at a constant temperature of 37°C and the medium was homogenized by placing them on an orbital shaker at 100 rpm during the assay. 1 mL samples were removed at predefined time points and replaced with fresh PBS medium. Voriconazole concentration was determined using a validated ultra-performance liquid chromatography (UHPLC) method. The results of the study are shown in Table 4. No significant differences in voriconazole release from the eye drops were observed between the two Franz cell types (a ns).

[0078] Table 4. In vitro released amount (pg) of a voriconazole eye drop in 3D printed coated Franz cells and in glass Franz cells (n=3).

[0079] Example 5. Retention testing of different drugs with 3D printed laboratory objects coated with glycidyl-terminated Poly(Bisphenol A-co-epichlorohydrine). Vials Figure 1 A were designed using CAD software and fabricated by 3D printing using a stereolithography 3D printer. The 3D printed vials were subsequently coated by a dipping method with a transparent coating with various concentrations of glycidyl-terminated Poly(Bisphenol A-co-epichlorohydrine) in ethyl acetate: 0.1% (w / v), 1% (w / v), 2.5% (w / v), 5% (w / v), 7.5% (w / v), and 10% (w / v). Dexamethasone base retention was compared according to Example 2 over 8 days between the coated and glass vial groups, expressed as % drug recovery according to Equation 1.Table 5 shows that the drug concentration is adequate when using a 1% (w / v) concentration and remains constant and invariable from a 10% (w / v) concentration of glycidyl-terminated Poly(Bisphenol A-co-epichlorohydrine) in the coated vessels. Concentrations of up to 70% (w / v) are possible. Above this concentration, the coating properties are not adequate for coating 3D printed objects; the mixture is too viscous and not suitable for the purposes described. Table 5. Results of average recovery values ​​(%) of dexamethasone base in vessels made by 3D printing with Clear resin after coating with solutions of different concentrations of glycidyl-terminated Poly(Bisphenol A-co-epichlorohydrine) (n=3).

[0080] Subsequently, the 3D printed vials were coated by dipping method with a clear coating comprising 10% (w / v) glycidyl-terminated Poly(Bisphenol A-co-epichlorohydrin) in ethyl acetate, and the other group was left uncoated. The retention of different drugs with distinct physicochemical properties prepared according to Example 2 over 8 days was compared between the coated and glass vial groups, expressed as drug recovery according to Equation 1.

[0081] Table 6 shows how the concentration of drugs remains constant and unchanged in the coated containers and decreases in the uncoated ones. Table 6. Results of average recovery values ​​(%) of different drugs in the containers made by 3D printing with Clear resin without and after being coated with Poly(Bisphenol A-co-epichlorohydrin) with glycidyl terminal (n=3).

[0082] Example 6. Evaluation of the reusability of coated containers.

[0083] The 3D-printed vials coated with the opaque coating comprising 10–25% (w / v) Bisphenol-A with bulking agents were cleaned using a neutral detergent or a combination of neutral detergent and alcohol. EDX analysis was performed to detect possible changes in the elemental composition of the coating that could have resulted from drug retention during contact. The analysis focused on identifying specific atoms of the drugs that had come into contact with the coating, including nitrogen (ciclopirox olamine, bevacizumab, human insulin, and voriconazole), cobalt (hydroxocobalamin), and fluorine (dexamethasone base and voriconazole). EDX analysis did not detect the presence of any target atoms, indicating that the drugs did not adhere to the coating surface.Furthermore, there were no significant differences in composition between unwashed items, items washed with neutral detergent, and items washed with neutral detergent and alcohol.

[0084] XRD analysis was performed to examine possible changes in coating crystallinity after washing and to detect any signals corresponding to drug crystallization. Figure 7 shows the diffractograms of all tested objects. All observed signals corresponded to the coating matrix, confirming the absence of drug retention within the coating. Although there were variations in phase crystallinity after cleaning, all phases remained present. A slight broadening of the signals was observed after washing, suggesting a minor increase in crystallinity. The Scherrer formula was used to calculate the change in coating crystallinity before and after washing. The crystallite sizes of barite, chlorite, and rutile were determined (Table 7) and compared between the different objects.The washed coated objects showed larger crystal domain sizes for all components, regardless of the washing method (a<0.05). It is possible that some components of the Bisphenol coating with fillers dissolved during washing with water or ethanol. During the drying process, these compounds could crystallize, leading to the formation of larger crystals with different conformations.

[0085] Table 7. Results of Scherrer's formula on washed and unwashed coated vials. Raman spectroscopy analysis was performed to evaluate possible changes in the functional groups present on the surface of the washed coated objects. Figure 7 shows the Raman spectra of the washed objects compared to the bisphenol coating. Signals associated with Ti-O bonds showed reduced intensity in the washed objects, suggesting that the cleaning process might remove some TiC>2 particles from the coating surface. Furthermore, XRD results suggested an increase in rutile crystallite size. Consequently, the analyzed area may have larger crystals, although in lower quantity than before washing, which could explain the decrease in intensity of the Ti-O bond signals. However, no new signals were observed for any of the drugs in the Raman analysis, indicating that the bisphenol coating did not cause drug retention.

[0086] The 3D-printed and coated recycled vials were subjected to drug retention testing by introducing a 50 pg / mL solution of dexamethasone base after cleaning and drying. Table 8 illustrates that the recycled objects showed complete recovery of dexamethasone base, with no drug retention observed at any time point. No significant differences were found between time points for dexamethasone base (a ns). These results demonstrate that the coated objects are reusable, as the cleaning process did not affect drug retention.

[0087] Table 8. Dexamethasone base recovery (%) from 3D-printed vials coated with opaque coating comprising 10–25% (w / v) Bisphenol with bulking agents and reused. (n=3).

[0088] Example 7. Resistance to pH, temperature and solvents.

[0089] Vials coated with the opaque coating comprising 10–25% (w / v) Bisphenol with bulking agents were exposed to a variety of commonly used solvents for 24 hours, including liquid paraffin, acetone, ethanol, isopropanol, neutral detergent, and methanol. SEM images of the vials after exposure to the solvents are shown in Figure 8. Before exposure, the solvents were visually examined for any signs of turbidity, and no visual disturbances were observed after contact with the coating. The SEM images were inspected for possible changes in the surface morphology of the coating, such as pores or imperfections, but no significant alterations were observed for any of the solvents. These results indicate that the coating is compatible with the tested solvents.

[0090] The coating's temperature resistance was evaluated by exposing the 3D-printed vials to different temperature conditions. Tested conditions included 4°C, 25°C, and 37°C. A 50 pg / mL dexamethasone base solution was added to the 3D-printed coated vials, and drug retention was assessed following the method described in Example 2.

[0091] On day 8, dexamethasone base recovery was approximately 100% at all temperatures tested except 37 °C, where recovery was 104.72 ± 2.31% (Table 8). This slight increase in recovery at 37 °C was likely due to solvent evaporation in the 3D-printed vials, which were covered with Parafilm® M sealing film instead of a cap like the glass vials.

[0092] Table 9. Recovery of dexamethasone base (%) from 3D printed vials coated with opaque coating comprising 10-25% (w / v) Bisphenol with bulking agents and at different temperatures (n=3).

[0093] The coating resistance to pH changes was investigated by exposing the 3D-printed vials to neutral, acidic, and basic pH buffers. The buffers consisted of a phosphate solution at pH 7.5 and pH 9, and a simulated gastric fluid at pH 1.2 without enzymes. The buffers were subsequently removed, and a 50 pg / mL dexamethasone base solution was added to the 3D-printed coated vials. Drug retention was assessed following the method described in Example 2. The coated objects showed complete recovery of dexamethasone base with no drug retention at all time points (Table 10). On day 8, dexamethasone base recovery was approximately 100% for all pH values ​​tested.

[0094] Table 10. Recovery of dexamethasone base (%) from 3D printed vials coated with opaque coating comprising 10-25% (w / v) Bisphenol with fillers and at different pH (n=3)

[0095] Example 8. Comparison of retention capacity of opaque coating comprising 10-25% (w / v) of Bisphenol combined with fillers to induce opacity and transparent coating comprising 10-25% (w / v) of Bisphenol.

[0096] The microscopic appearance of the clear coating surface is shown in Figure 9. As with the opaque coating, no large pores or defects were observed. This indicates that the clear coating covers the entire resin surface. Small crystals can be observed on the surface of the clear coating, the presence of which was confirmed by XRD analysis.

[0097] The elemental composition of the clear coating surface was analyzed by EDX and compared with that of the opaque coating. Table 11 presents the elemental composition comparison between the coatings. The opaque coating contains titanium, which is part of the rutile molecules used as a white pigment. In addition, the opaque coating contains chlorite, which in turn contains the elements Mg, Al, and Si. In contrast, the clear coating has a much lower proportion of these elements than the opaque coating, suggesting that chlorite is not present in its composition. The presence of Ba suggests that barite is present in both coatings, although it appears to be present in a higher proportion in the clear coating.

[0098] Table 11. Elemental analysis of the surface of opaque and transparent coatings containing 10-25% Bisphenol.

[0099] The clear coating also contains calcium (Ca). To determine which compound the calcium belongs to, XRD analysis was performed. Figure 10 shows the diffractograms of the opaque and clear coatings. The previously analyzed calcium corresponds to calcium carbonate or calcite molecules (CaCO2), probably included to act as filler material and provide hardness and flexibility to the coating polymer. As can be seen, the signals corresponding to rutile are not visible, demonstrating that rutile is not part of the clear coating composition. Raman spectra (Figure 11) confirmed the absence of TiO2 in the clear coating, since the signals corresponding to the Ti-O bonds are missing. Despite this difference, the spectra of both coatings are similar, suggesting a similar composition.

[0100] A retention study was performed with a 50 pg / mL solution of dexamethasone base in 3D-printed vials coated with the clear coating. Table 12 shows that the clear coating showed complete recovery of dexamethasone base with no drug retention at all time points, with no significant differences found between time points for dexamethasone base (ns). Furthermore, no significant differences in dexamethasone base retention were found between the opaque and clear coatings (ns). These results suggest that the clear coating would not cause drug retention.

[0101] Table 12. Recovery of dexamethasone base (%) from 3D printed vials coated with clear coating comprising 10-25% (w / v) Bisphenol.

Claims

CLAIMS 1. Laboratory object that is characterized by being made of an acrylic resin coated with a Bisphenol.

2. Laboratory object according to claim 1, which is a diffusion cell.

3. Method for manufacturing a laboratory object according to claim 1, said laboratory object being suitable for carrying out experiments with pharmaceutical or cosmetic active ingredients, preventing them from being absorbed or adsorbed to its surface, comprising: i. 3D printing of the object, i. coating of the printed objects using a solution comprising Bisphenol, iii. drying of the applied coating.

4. Manufacturing method according to claim 3, wherein the 3D printing is a stereolithographic printing.

5. Manufacturing method according to any of claims 3-4, wherein the printing by stereolithography uses a liquid resin of acrylic monomers.

6. Manufacturing method according to any of claims 3-5, wherein the solution has a Bisphenol percentage of between 1% and 75% w / v.

7. Manufacturing method according to any of claims 3-6, wherein Bisphenol is selected from the group consisting of Poly(Bisphenol A-co-epichlorohydrin) copolymer with glycidyl terminal, Bisphenol A-diglycidi leter methacrylate (bis-GMA), 2,2-bis(4-(2-methacryloxyprop-1-oxy)phenyl) propane (CHsBis-GMA).

8. Manufacturing method according to any of claims 3-7, wherein the solution is prepared with a solvent selected from ethyl acetate, dichloromethane, trichloromethane, toluene, xylene, ethylbenzene, butanone, 4-hydroxy-4-methyl-2-pentanone and diacetone alcohol.

9. Manufacturing method according to any of claims 3-8, wherein the coating method is selected from dip coating, air atomization system coating, high volume low pressure gun coating, and spray gun coating.

10. Manufacturing method according to claim 3, wherein the drying method is convection drying, room temperature drying or vacuum drying.

11. Manufacturing method according to claim 3, further comprising a step prior to step i, which is the design of the laboratory object by CAD software.

12. Diffusion cell obtainable by the method according to any of claims 3- 13. Use of a laboratory object according to claim 1, obtainable according to claims 1-9, or of the diffusion cell according to claim 10, for carrying out in vitro or ex vivo tests in the presence of pharmaceutical or cosmetic active ingredients.

14. Use of a laboratory object according to claim 1, obtainable according to claims 1-10, or of the diffusion cell according to claim 11, for preventing the absorption and adsorption of pharmaceutical active ingredients on the surface.

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