Efficacy of a new delivery system based on solid lipid microparticles for the oral administration of the non-conventional antioxidant IAC on a diabetes mouse model.

PURPOSE: We previously showed the positive effects of the new antioxidant molecule bis(1-hydroxy-2,2,6,6-tetramethyl-4-piperidinyl)-decandioate (IAC) in reducing basal hyperglycaemia and relieving glucose intolerance in a diabetes model. However, the chemical properties of IAC did not allow an efficient oral administration, thus representing the main failing of that study. Here, we tested the effect of a new oral delivery system based on solid lipid microparticles (SLMs) in a diabetes mouse model. METHODS: The diabetes model was induced in C57B1/6J mice using streptozotocin and nicotinamide. Only the animals that overcame the glycaemic threshold of 180 mg/dL were enrolled in the study. Diabetic animals were then randomly assigned to 4 groups (n = 9) and treated once a day for 5 consecutive weeks with IAC (50, 100, and 150 mg/kg b.w.). The control group was composed of (n = 7) healthy mice that received only the vehicle. Glucose level was weekly monitored during the treatment period and up to 3 weeks after the suspension of the treatment. Glucose tolerance and insulin-resistance test were carried out. RESULTS: Our results showed that SLMs maintained the IAC effect in reducing basal hyperglycaemia as well as improving the insulin sensitivity and glucose tolerance. CONCLUSION: The present study confirms that SLMs are promising drug carriers, which allow the oral administration of IAC ensuring its therapeutic efficacy. The concrete possibility to administer IAC per os represents a significant breakthrough in the putative consideration of this multi-radical scavenger in the diabetes therapeutic approach.

Structure--activity relationship in cinnamamides. 3. Synthesis and anticonvulsant activity evaluation of some derivatives of (E)- and (Z)-m-(trifluoromethyl)cinnamamide.

The (E)- and (Z)-m-(trifluoromethyl)-alpha, beta-dimethylcinnamamides and some of their N-alkyl derivatives were prepared and pharmacologically tested as anticonvulsant agents in order to verify if a ring substituent, like the m-CF3 group, different from a halogen but possessing the same electronic effect could lead to equally active compounds. Some (E)-m-(trifluoromethyl)-alpha-methyl- and -non-methyl-substituted-cinnamamides were also prepared and tested. In the alpha, beta-dimethyl series the results show that the m-CF3 group leads to products more active than the ones unsubstituted on the phenyl ring but still less active than the p-halogen-substituted compounds previously studied. In the alpha-methyl and non-methyl-substituted series, the trend shows the m-CF3 group being able to produce less toxic and, in some cases, more active products than the previously studied amides.

Structure-activity relationship in cinnamamides. 2. Synthesis and pharmacological evaluation of some (E)- and (Z)-N-alkyl-alpha, beta-dimethylcinnamamides substituted on the phenyl group.

Several (E)- and (Z)-N-alkyl-alpha,beta-dimethylcinnamamides variously substituted on the phenyl group were synthesized from their corresponding acids and characterized through their NMR spectra. The compounds were tested to determine the relationship existing between their action on the CNS and the activity exhibited by the corresponding amides unsubstituted on the phenyl, previously studied. Substitution with the same group always had the same effects on the biological activity of the (E)-N-alkyl-alpha,beta-dimethylcinnamamides selected; these effects mainly regarded anticonvulsant activity which is the most noteworthy action of these compounds. This activity was reduced by electron-donating substituents and increased by electron-withdrawing ones. In the Z series the p-phenyl substitution with a halogen reduced or suppressed the CNS stimulant activity exhibited by the parent compounds.

Structure-activity relationships in cinnamamides. 1. Synthesis and pharmacological evaluation of some (E)- and (Z)-N-alkyl-alpha,beta-dimethylcinnamamides.

Two series of (E)- and (Z)-N-alkyl-alpha,beta-dimethylcinnamamide derivatives were prepared and the biological activity of these compounds was investigated in a series of pharmacological tests. All compounds tested had clear activity on the CNS; generally, this was depressant with E isomers, while Z isomers always caused marked stimulation (tremors and convulsions). Some of the E isomers also had a clear-cut anticonvulsant activity as shown by the antagonistic effect on pentylenetetrazole-induced seizures in the mouse. The NMR spectra of these compounds, which confirm their configurations, are discussed.

An investigation into the effects of residual water on the glass transition temperature of polylactide microspheres using modulated temperature DSC.

The objective of the study was to ascertain residual water levels in polylactide and polylactide-co-glycolide microspheres prepared using the solvent evaporation technique and to investigate the effects of that water on the glass transitional behaviour of the microspheres. Microspheres were prepared from polylactic acid (PLA) and polylactide-co-glycolide (PLGA) 50:50 and 75:25 using a standard solvent evaporation technique. The glass transition was measured as a function of drying conditions using modulated temperature DSC. The microspheres were found to contain very low levels of dichloromethane, while residual water levels of up to circa 3% w/w were noted after freeze or oven drying, these levels being higher for microspheres containing higher glycolic acid levels. The residual water was found to lower the T(g) following the Gordon-Taylor relationship. The data indicate that the microparticles may retain significant water levels following standard preparation and drying protocols and that this drying may markedly lower the T(g) of the spheres.

Fractal analysis of beta-cyclodextrin-indomethacin particles compacted by ultrasound.

An association between indomethacin and beta-cyclodextrin (beta-CD) was obtained by compacting a 1:2 molar physical mixture by ultrasound. The product prepared by this technique was compared with the initial physical mixture and with materials having the same composition but prepared by a simple compaction and kneading process. The samples examined by scanning electron microscopy revealed morphological differences related to the methods of preparation. In particular the material obtained by ultrasound had a smooth surface and on milling produced particles of uniform size; moreover EDAX analysis (energy dispersion analysis by X-rays) also revealed a homogeneous distribution of the two components in each particle examined. Surface fractal dimension of the surface of these particles is very low, suggesting the presence of a regular and smooth surface whereas the sample obtained by kneading had a higher value. The reactive dimension related to dissolution was much higher than the corresponding surface dimension in all cases and had comparable values for all samples regardless of the techniques used. This behavior was attributed to the hydrophilicity of beta-CD, which levels the differences during dissolution and which could be originated by surface imperfections. Experimental results suggested that the material obtained by ultrasound had a dissolution rate comparable to that measured with the kneaded material. The use of ultrasound, however, can both reduce the production time and improve the homogeneity of the association between indomethacin and beta-CD.

Effects of ultrasound-assisted compaction on Ketoprofen/Eudragit S100 mixtures.

Ketoprofen alone and in binary mixtures with Eudragit S100 was compacted by an ultrasound-assisted (US) tableting machine at an energy ranging from 50 to 400 J. The final material was analysed by TLC and HPLC: no decomposition product of the active agent was found. IR spectra and HSM revealed the absence of any interaction between the two components. Thermal analysis (DSC) evidenced that ketoprofen inside the mixtures was transformed into an amorphous state, documented by the decreasing of the DeltaHfus as the Eudragit/ketoprofen ratio increases and as US energy increases. While pure ketoprofen recovers its crystalline state quickly after the US treatment, the presence of Eudragit was found to slow down or possibly to prevent the regeneration of the crystallinity.

Ultrasound-compacted and spray-congealed indomethacin/polyethyleneglycol systems.

The product obtained by ultrasound (US)-assisted compaction was compared with a solid dispersion for systems containing polyethyleneglycols (PEGs) of different molecular weights and indomethacin (IMC), at the weight ratio 9:1, obtained by traditional melting and followed by a new US-assisted spray-congealing technique. US-discharge during compaction affects crystallinity of both IMC and PEG: pure IMC changes to an amorphous form and, when in mixture with PEG, partially dissolves in the excipient: this causes an increase of the dissolution rate of the drug. Differential scanning calorimetry (DSC) thermograms do not reveal any endothermic peak associated with the melting of the drug, while X-ray diffractograms show a loss of crystallinity of both IMC and PEG in the US-compacted granules. The extent of a back-crystallisation, which reduces the dissolution rate, as a function of the ageing of the material, depends on the type of the selected PEG. When a molten IMC/PEG mixture was transformed into microspheres by an US-assisted spray-congealing technique, the behaviour at dissolution almost recalls that of US-compacted granulates and some differences are briefly discussed.

Formation of ion-pairs in aqueous solutions of diclofenac salts.

In this work we studied the ability of the diclofenac anion to form ion-pairs in aqueous solution in the presence of organic and inorganic cations: ion-pairs have a polarity and hydrophobicity more suitable to the partition than each ion considered separately and can be extracted by a lipid phase. The cations considered were those of the organic bases diethylamine, diethanolamine, pyrrolidine, N-(2-hydroxyethyl) pyrrolidine and N-(2-hydroxyethyl) piperidine; the inorganic cations studied were Li(+), Na(+), K(+), Rb(+), Cs(+). Related to each cation we determined the equilibrium constant (K(XD)) for the ion-pair formation with the diclofenac anion in aqueous solution and the water/n-octanol partition coefficient (P(XD)) for each type of ion-pair formed. Among the alkali metal cations, only Li(+) shows some interaction with the diclofenac anion, in agreement with its physiological behaviour of increasing clearance during the administration of diclofenac. The influence of the ionic radius and desolvation enthalpy of the alkali metal cations on the ion-pair formation and partition was briefly discussed. Organic cations promote the formation of ion-pairs with the diclofenac anion better than the inorganic ones, and improve the partition of the ion-pair according to their hydrophobicity. The values of the equilibrium parameters for the formation and partition of ion-pairs are not high enough to allow the direct detection of their presence in the aqueous solution. Their formation can be appreciated in the presence of a lipid phase that continuously extracts the ion-pair. Extraction constants (E(XD)=P(XD) times K(XD)) increase passing from inorga to organic cations. This study could help to clarify the mechanism of the percutaneous absorption of diclofenac in the form of a salt, a route where the formation of ion-pairs appears to play an important role.

Description and preliminary evaluation of a new ultrasonic atomizer for spray-congealing processes.

A new atomizer that operates with ultrasonic energy is described. This apparatus is intended to obtain microparticulate drug delivery systems through spray-congealing or spray-drying technologies. In this work, some experimental results are reported on model systems submitted to spray-congealing. The formulations under examination contained theophylline and fenbufen as model drugs and stearic acid, carnauba wax, Cutina HR(R) and Compritol 888 ATO(R) as low melting excipients. Non-aggregate and spherical-shaped microparticles were obtained with all the materials tested; moreover, they had smooth surface and good flowability. The particle sizes depend on the amount of drug present and in each case the maximum size value of the distribution frequency was found to be 375 mu. In vitro release of the drug depends on its solubility and on the excipient lipophilicity. The results suggest that the ultrasound-assisted atomizer could be proposed as a possible alternative to traditional atomizers used for spray-congealing in the pharmaceutical field.

Preparation and characterization by morphological analysis of diclofenac/PEG 4000 granules obtained using three different techniques.

The steam granulation is a new wet granulation technique, which involves the use of steam water instead of traditional liquid water as granulation liquid. The aim of this work was to evaluate the possibility of using this new technique to prepare diclofenac-polyethylene glycol 4000 accelerated-release granules. Steam granules were prepared in a laboratory scale high-shear mixer, and their properties were then compared to those of granules, having the same composition, obtained by traditional granulation techniques (wet and melt granulation). The results showed that, selecting the proper process parameters, it was possible to obtain granules using all the three methods; however, the total process time was significantly shorter for steam granulation (30 min) in comparison to traditional wet granulation (70 min), due to the lower amount of used water. The morphological characterization of steam, water and melt granules, performed by scanning electron microscopy (SEM) and image analysis, revealed that steam granules had a more spherical shape and a larger surface area with respect to water and melt ones, suggesting a possible difference in dissolution behavior. Moreover, differential scanning calorimetry (DSC) and X-ray powder diffraction analysis evidenced the transformation of the drug from its originally crystalline form into the amorphous one. Finally, the in vitro dissolution tests showed an increased dissolution rate of the drug from the granules (in particular steam granules) in comparison to pure drug and physical mixture. In conclusion, the results of this study suggested that the steam granulation technique could be considered an interesting alternative to traditional wet granulation to improve the dissolution rate of diclofenac.

Characterization of ciclosporin A loaded poly (D,L lactide-co-glycolide) microspheres using modulated temperature differential scanning calorimetry.

The aim of this study was to investigate the physical structure of poly (D,L lactide-co-glycolide) (PLGA) microspheres loaded with ciclosporin A in terms of the amorphous properties of the individual components and the phase separation characteristics of the binary systems. Microspheres were prepared using a standard oil-in-water emulsion technique. The thermal properties of the PLGA, ciclosporin A and loaded spheres were investigated using modulated temperature differential scanning calorimetry (MTDSC) using a TA Instruments MTDSC 2920, with scanning electron microscopy (SEM), X-ray powder diffraction (XRD) and high-performance liquid chromatography used as supportive techniques. MTDSC indicated a glass transition for ciclosporin A in the reversing heat flow signal at 107 degrees C, supported by temperature cycling studies, while XRD showed clear evidence for diffraction peaks, thereby indicating that the material as received is semi-crystalline. The unloaded PLGA spheres showed a glass transition (Tg) at 43 degrees C, with no reduction in Tg being observed on loading the peptide up to 50%, w/w. Similarly, no evidence for diffraction peaks were seen for the drug-loaded systems, although the glass transition corresponding to the peptide was observed for the loaded microspheres, suggesting that the drug is present as a separate amorphous phase. Similarly, SEM studies showed the appearance of distinct "islands" on the surface of the spheres that are suggested to correspond to the drug phase, with the size of the islands increasing with drug loading. Evidence is therefore presented that ciclosporin A may exist in a range of solid states, with the degree of crystallinity being altered by processing. In addition, there appears to be little or no miscibility between the drug and PLGA using the manufacturing protocol employed here. These findings may have implications for the choice of manufacturing protocol, the release of peptide drugs from PLGA microspheres and the chemical and physical stability of such drugs.

The effect of polymer coatings on physicochemical properties of spray-dried liposomes for nasal delivery of BSA.

This work describes the development of spray dried polymer coated liposomes composed of soy phosphatidylcholine (SPC) and phospholipid dimyristoyl phosphatidylglycerol (DMPG) coated with alginate, chitosan or trimethyl chitosan (TMC), that are able to penetrate through the nasal mucosa and offer enhanced penetration over uncoated liposomes when delivered as a dry powder. All the liposome formulations, loaded with BSA as model antigen, were spray-dried to obtain powder size and liposome size in a suitable range for nasal delivery. Although coating resulted in some reduction in encapsulation efficiency, levels were still maintained between 60% and 69% and the structural integrity of the entrapped protein and its release characteristics were maintained. Coating with TMC gave the best product characteristics in terms of entrapment efficiency, glass transition (T(g)) and mucoadhesive strength, while penetration of nasal mucosal tissue was very encouraging when these liposomes were administered as dispersions although improved results were observed for the dry powders.

A new approach to enhance oral bioavailability of Silybum Marianum dry extract: association of mechanochemical activation and spray congealing.

The aim of the work was to produce a delivery system for Silybum Marianum dry extract with enhanced oral bioavailability by combining two technologies (mechanochemical activation and spray congealing). Initially, the active was coground with sodium croscarmellose in a planetary mill in order to reach an activated state more prone to dissolution. DSC, XRD, FT-IR and LD analyses showed the formation of nanosized particles of dry extract, with reduced degree of crystallinity of the main crystalline flavolignans (silybine A and B). Then, microparticles containing the activated coground and, as comparison, the corresponding physical mixture of extract and polymer and the dry extract alone were produced by spray congealing technology using Gelucire(®) 50/13 as a hydrophilic low m.p. carrier. Microparticles containing the activated coground were produced spherical in shape, achieved satisfactory yield and high encapsulation efficiency. These microparticles, in addition to a favourable in vitro solubilisation kinetic, in a preliminary in vivo study in five rats demonstrated their ability to improve very significantly the oral bioavailability of the main flavolignans of Silybum Marianum dry extract (silybin A and B). These results suggested that the association of mechanochemical activation and spray congealing could be considered an innovative and very useful approach to the oral delivery of Silybum Marianum. Furthermore, for the first time the possibility of successfully applying the spray congealing technology for the preparation of a herbal drug delivery system was shown.

Poloxamer 407 microspheres for orotransmucosal drug delivery. Part II: In vitro/in vivo evaluation.

Aim of this research was to evaluate novel microspheres based on poloxamer 407, alone or in mixture with Gelucire(®) 50/13, as possible buccal delivery system for atenolol (AT). The microspheres have been prepared by spray congealing and investigated to assess AT in vitro delivery through cellulose membranes and ex vivo permeation using porcine buccal mucosa. The microparticles were tested as such or directly compacted to obtain tablets. For comparison the physical mixtures, tablets of the physical mixtures and an AT solution were examined. Finally, the microparticles were sublingually administered in rabbits to evaluate AT pharmacokinetics compared to a market oral tablet (reference). The AT release from microspheres through the synthetic membrane was delayed with respect to the drug solution, more markedly when microparticles contained poloxamer as unique adjuvant; this formulation enhanced AT transmucosal permeation. The enhancement effect of poloxamer was confirmed by the permeation experiments on the corresponding physical mixture. Tabletting hindered both release through cellulose membranes and transmucosal permeation of drug. In vivo studies revealed that the absolute bioavailability of microsphere formulations was higher than that of reference in spite of a lower dosage of drug, suggesting a possible dose reduction by AT microparticles orotransmucosal administration.

Poloxamer 407 microspheres for orotransmucosal drug delivery. Part I: formulation, manufacturing and characterization.

The two-part article aimed to investigate poloxamer 407-based microspheres as a novel platform for enhancing and controlling the delivery of atenolol across the oromucosal tissue. In the Part I of the work, atenolol-loaded poloxamers 407 microparticles were prepared by the solvent free spray congealing technology. This approach was feasible upon the high viscosity of the systems allowing for high loaded (20% w/w) non-aggregated microspheres. Several formulations were studied and the results demonstrated that the drug release patterns, solubility data, mucoadhesion to buccal tissue and gelling properties in saliva could be modified by adding different amount of an amphiphilic polymer-lipid excipient (Gelucire(®) 50/13) to poloxamer 407. Particularly, microspheres based only on poloxamer 407 exhibited very high solubility, mucoadhesive strength and gelling behaviour. To assess their potential as matrix for buccal application, the gelling property and the drug release from tablets obtained from direct compression of the microparticles were further evaluated. The microspheres were then characterized by differential scanning calorimetry, X-ray powder diffraction and Fourier transform-infrared spectra analysis. No solid state modifications and chemical interactions were detectable in the microspheres after manufacturing and during storage, suggesting their stability and use as orotransmucosal delivery systems.

Solid state mechanochemical activation of Silybum marianum dry extract with betacyclodextrins: Characterization and bioavailability of the coground systems.

Silybum marianum dry extract, whose therapeutic use is partially restricted by the insolubility in water of its main flavonolignans, was subjected to a mechanochemical activation process in planetary mill using betacyclodextrins as carriers. After optimization of the operating conditions according to an established theoretical model, the best active-to-carrier proportion was selected from the preliminary trials. When using the optimized conditions, the mechanochemical process permits an improvement of the physico-chemical properties of the active, which reaches an "activated" solid state, that is stable for at least 1 year. In fact, XRD, DRIFT and Raman spectroscopy analyses showed that the main extract component, Silybin, completely lost its crystalline structure after co-grinding with betacyclodextrins and formed weak interactions with the carrier. The powder characteristics remarkably changed after co-grinding, leading to a sample with a very small mean diameter and with a twofold increase of the specific surface area in comparison to the dry extract. The activated solid state of the coground systems remarkably enhanced the in vitro drug dissolution kinetics with consequent improved oral bioavailability. Furthermore, the in vivo studies on rats revealed a 6.6-fold bioavailability increase respect to the S. marianum Italian commercial product used as reference (Silirex 200 capsules).

[Chemical and pharmacological research on pyran derivatives. XV. Phenyl-substituted 2-(dialkylamino)chromones]. / Ricerche chimiche e farmacologiche su derivati piranici. Nota XV-2-(Dialchilammino)cromoni fenilsostituiti.

Some 2-(dialkylamino)chromones phenyl substituted in position 6 or 7 or 8 were synthesized by reaction of N,N-dialkylethoxycarbonylacetamides with 4- or 3- or 2-biphenylol, respectively, in the presence of phosphorus oxychloride. The pharmacological activity of these compounds was then evaluated and compared with that shown by naphthopyran derivatives of structures (I), (II), and (III). With this same purpose also 6- and 8-benzyl derivatives of 2-(dialkylamino)chromones were prepared by using in the reaction 4- or 2-benzylphenol. Pharmacological screening showed that 6-phenyl substituted 2-(dialkylamino)chromones maintained the antireserpine and antimetrazole acti-activities which were possessed by the 1H-naphtho[2,1-b]pyran derivatives (I) and the 4H-naphtho[2,3-b]pyran derivatives (II), whereas 7-phenyl substituted compounds were devoid of any activity. 8-Phenyl substituted 2-(dialkylamino)chromones maintained to some extent the antiamphetamine activity which was clearly shown by the 4H-naphtho[1,2-]pyran derivatives (III). Furthermore, the compounds bearing the benzyl substituent both in the 6 or 8 position showed only antimetrazole activity.

[Chemical and pharmaceutical research on pyran derivatives. XI. Synthesis of 2-dialkylamino-4-oxo-10-methyl-4H-naphtho[2,3b]pyrans]. / Ricerche chimiche e farmacologiche su derivati piranici. Nota XI-Sintesi di 2-dialchilammino-4-oxo-10-metil-4H-nafto[2,3-b]pirani.

Reaction of N,N-dialkylethoxycarbonylacetamides with 1-methyl-2-naphthol, in the presence of phosphorus oxychloride, gave rise to the formation of 2-dialkylamino-4-oxo-10-methyl-4H-naphtho[2,3-b] pyrans through the preliminary attack of the amide-phosphorus oxychloride reactant at the phenolic hydroxyl and cyclization at position 3 of the naphthalene moiety. However when (N-alkyl,N-phenyl)ethoxycarbonylacetamides were used in the reaction an ortho position of the N-phenyl group was involved in the cyclization and 1-alkyl-2(1'-methyl-2'-naphthoxy)-4-quinolones were achieved. Pharmacological investigation showed that some naphtho[2,3-b]pyran derivatives have neurotropic activity of the sedative, anticonvulsant and antidepressant type very similar to that shown by previously studied 1-oxo-3-dialkylamino-1H-naphtho[2,1-b]pyrans (5).