Green Hydrogels Composed of Sodium Mannuronate/Guluronate, Gelatin and Biointeractive Calcium Silicates/Dicalcium Phosphate Dihydrate Designed for Oral Bone Defects Regeneration

Innovative green, eco-friendly, and biologically derived hydrogels for non-load bearing bone sites were conceived and produced. Natural polysaccharides (copolymers of sodium D-mannuronate and L-guluronate) with natural polypeptides (gelatin) and bioactive mineral fillers (calcium silicates CaSi and dicalcium phosphate dihydrate DCPD) were used to obtain eco-sustainable biomaterials for oral bone defects. Three PP-x:y formulations were prepared (PP-16:16, PP-33:22, and PP-31:31), where PP represents the polysaccharide/polypeptide matrix and x and y represent the weight % of CaSi and DCPD, respectively. Hydrogels were tested for their chemical-physical properties (calcium release and alkalizing activity in deionized water, porosity, solubility, water sorption, radiopacity), surface microchemistry and micromorphology, apatite nucleation in HBSS by ESEM-EDX, FT-Raman, and micro-Raman spectroscopies. The expression of vascular (CD31) and osteogenic (alkaline phosphatase ALP and osteocalcin OCN) markers by mesenchymal stem cells (MSCs) derived from human vascular walls, cultured in direct contact with hydrogels or with 10% of extracts was analysed. All mineral-filled hydrogels, in particular PP-31:31 and PP-33:22, released Calcium ions and alkalized the soaking water for three days. Calcium ion leakage was high at all the endpoints (3 h–28 d), while pH values were high at 3 h–3 d and then significantly decreased after seven days (p < 0.05). Porosity, solubility, and water sorption were higher for PP-31:31 (p < 0.05). The ESEM of fresh samples showed a compact structure with a few pores containing small mineral granules agglomerated in some areas (size 5–20 microns). PP-CTRL degraded after 1–2 weeks in HBSS. EDX spectroscopy revealed constitutional compounds and elements of the hydrogel (C, O, N, and S) and of the mineral powders (Ca, Si and P). After 28 days in HBSS, the mineral-filled hydrogels revealed a more porous structure, partially covered with a thicker mineral layer on PP-31:31. EDX analyses of the mineral coating showed Ca and P, and Raman revealed the presence of B-type carbonated apatite and calcite. MSCs cultured in contact with mineral-filled hydrogels revealed the expression of genes related to vascular (CD31) and osteogenic (mainly OCN) differentiation. Lower gene expression was found when cells were cultured with extracts added to the culture medium. The incorporation of biointeractive mineral powders in a green bio-derived algae-based matrix allowed to produce bioactive porous hydrogels able to release biologically relevant ions and create a suitable micro-environment for stem cells, resulting in interesting materials for bone regeneration and healing in oral bone defects.

Predictive evaluation of multicomponent direct compress model tablets by integrating sphere UV-Vis spectroscopy and chemometrics

BACKGROUND: Control of the pharmaceutical manufacturing process and active pharmaceutical ingredients (API) is essential to product formulation and bioavailability. OBJECTIVE: The aim of this study is to predict tablet surface API concentration by chemometrics using integrating sphere UV-Vis spectroscopy, a non-destructive and contact-free measurement method. METHODS: Riboflavin, pyridoxine hydrochloride, dicalcium phosphate anhydrate, and magnesium stearate were mixed and ground with a mortar and pestle, and 100 mg samples were subjected to direct compression at a compaction pressure of 6 MPa at 7 mm diameter. The flat surface tablets were then analyzed by integrating sphere UV-Vis spectrometry. Standard normal variate (SNV) normalization and principal component analysis were applied to evaluate the measured spectral dataset. The spectral ranges were prepared at 300–800 nm and 500–700 nm with SNV normalization. Partial least squares (PLS) regression models were constructed to predict the API concentrations based on two previous datasets. RESULTS: The regression vector of constructed PLS regression models for each API was evaluated. API concentration prediction depends on riboflavin absorbance at 550 nm and the excipient dicalcium phosphate anhydrate. CONCLUSION: Integrating sphere UV-Vis spectrometry is a useful tool to process analytical technology.

Controlled synthesis of dicalcium phosphate dihydrate (DCPD) from metastable solutions: insights into pathogenic calcification

Calcium phosphate (CaP) compounds may occur in the body as abnormal pathogenic phases in addition to their normal occurrence as bones and teeth. Dicalcium phosphate dihydrate (DCPD; CaPO4·2H2O), along with other significant CaP phases, have been observed in pathogenic calcifications such as dental calculi, kidney stones and urinary stones. While other studies have shown that polar amino acids can inhibit the growth of CaPs, these studies have mainly focused on hydroxyapatite (HAp; Ca10(PO4)6(OH)2) formation from highly supersaturated solutions, while their effects on DCPD nucleation and growth from metastable solutions have been less thoroughly explored. By further elucidating the mechanisms of DCPD formation and the influence of amino acids on those mechanisms, insights may be gained into ways that amino acids could be used in treatment and prevention of unwanted calcifications. The current study involved seeded growth of DCPD from metastable solutions at constant pH in the presence of neutral, acidic and phosphorylated amino acid side chains. As a comparison, solutions were also seeded with calcium pyrophosphate (CPP; Ca2P2O7), a known calcium phosphate inhibitor. The results show that polar amino acids inhibit DCPD growth; this likely occurs due to electrostatic interactions between amino acid side groups and charged DCPD surfaces. Phosphoserine had the greatest inhibitory ability of the amino acids tested, with an effect equal to that of CPP. Clustering of DCPD crystals giving rise to a “chrysanthemum-like” morphology was noted with glutamic acid. This study concludes that molecules containing an increased number of polar side groups will enhance the inhibition of DCPD seeded growth from metastable solutions.

Designing Calcium Phosphate Nanoparticles with the Co-Precipitation Technique to Improve Phosphorous Availability in Broiler Chicks

Dicalcium phosphate (DP) is a mineral involved in the metabolism and development and is used as a dietary source of phosphorus (PT) for poultry. Our study objective is to design nano-dicalcium phosphate (NDP) by co-precipitation. The methodological procedure was divided into two phases: (1) NDP synthesis, and (2) bird performance, digestibility, and Ca-P in chick’s tissues. The sizes of the NDP were 20–80 nm. NDP had the Ca: P ratio of 1:1.12. The birds were divided into control diet (available P (Pa) = 0.13%) and three supplementary P sources [Commercial (Calcium phosphate), analytical grade (DP) and nanoparticles (NDP)] with three Pa levels (0.24, 0.35, 0.46%). Supplementary P sources compared to the control treatment had the highest body weight gain (698.56 vs. 228; p < 0.05) and feed intake (FI) (965.18 vs. 345.82), respectively. The digestibility of PT (67%) improved with 0.35% NDP. The highest contents of PT -breast were with the levels of 0.35 and 0.46% NDP. The PT, ash, and diameters were higher (p < 0.05) with supplementary P compared to the control treatment. As conclusion, the use of 0.35% NDP was the ideal dose in the chicks for the digestibility, absorption values, and the amount of PT in the breast.

Dietary nano-dicalcium phosphate improves immune response and intestinal morphology of broiler chickens

Various levels of substitution of nano-dicalcium phosphate (nCaHPO4) for the calcium and phosphorus supplied by limestone and monocalcium phosphate were evaluated, based on weight change, serum constituents, immune response and intestinal morphology of broiler chickens. The birds were assigned to five dietary treatments containing 0% (control), 40%, 60%, 80%, and 100% substitution levels of nCaHPO4 from days 0 to 42. Data were collected for blood measurement and intestinal morphology and analysed with one-way analysis of variance. Feeds substituted with 40% nCaHPO4 showed a higher mean value of final weight and better feed conversion. The results revealed similar haematological parameters across the treatment groups. At the same time, nCaHPO4 influenced most of the serum biochemical constituents, such as calcium, phosphorus, glucose, albumin, and total protein. High concentrations of total protein were observed for birds fed 60% nCaHPO4 compared with other treatments. The highest comparable means for albumin were observed in birds fed 40% and 60% nCaHPO4. Cholesterol levels were increased when birds were fed 0% and 40% of diets. Elevated serum phosphorus concentration was observed when 0%, 40%, 60% and 100% substitution levels were fed, whereas 80% presented the least mean value. The results also showed that birds fed 40% nano-dicalcium phosphate had the highest villi length, which signified improved absorption of minerals. In conclusion, 40% and 60% levels of nCaHPO4 could be used to boost immune response and functional intestinal morphology of broiler chicken with no negative effect on haematological parameters.

Effects of Hinokitiol and Dicalcium Phosphate on the Osteoconduction and Antibacterial Activity of Gelatin-Hyaluronic Acid Crosslinked Hydrogel Membrane In Vitro

Many hydrogel-based crosslinking membranes have been designed and tailored to meet the needs of different applications. The aim of this research is to design a bifunctional hydrogel membrane with antibacterial and osteoconducting properties to guide different tissues. The membrane uses gelatin and hyaluronic acid as the main structure, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride as the crosslinker, hinokitiol as the antibacterial agent, and dicalcium phosphate anhydrous (DCPA) micron particles for osteoconduction. Results show that the hydrogel membrane with added DCPA and impregnated hinokitiol has a fixation index higher than 88%. When only a small amount of DCPA is added, the tensile strength does not decrease significantly. The tensile strength decreases considerably when a large amount of modified DCPA is added. The stress–strain curve shows that the presence of a large amount of hinokitiol in hydrogel membranes results in considerably improved deformation and toughness properties. Each group impregnated with hinokitiol exhibits obvious antibacterial capabilities. Furthermore, the addition of DCPA and impregnation with hinokitiol does not exert cytotoxicity on cells in vitro, indicating that the designed amount of DCPA and hinokitiol in this study is appropriate. After a 14-day cell culture, the hydrogel membrane still maintains a good shape because the cells adhere and proliferate well, thus delaying degradation. In addition, the hydrogel containing a small amount of DCPA has the best cell mineralization effect. The developed hydrogel has a certain degree of flexibility, degradability, and bifunctionality and is superficial. It can be used in guided tissue regeneration in clinical surgery.