The in vitro remineralizing effect of CPP-ACP and CPP-ACPF after 6 and 12 weeks on initial caries lesion

Abstract Objective: The aim of this in vitro study was to determine the effects of remineralization promoting agents containing casein phosphopeptide-stabilized amorphous calcium phosphate (CPP-ACP), or CPP-ACP in combination with fluoride (CPP-ACPF) on artificial white spot lesions (WSLs) after 6 and 12 weeks. Methodology: White spot lesions were created on 123 sectioned premolars (246 specimens) with a demineralization solution during a 96 hours pH-cycling regime. Two experimental groups were created: a CPP-ACP group (Tooth Mousse™), and a CPP-ACPF group (Mi Paste Plus™). Additionally, two control groups were created, one using only a conventional toothpaste (1450 ppm fluoride) and another one without any working agents. All teeth were also daily brushed with the conventional toothpaste except the second control group. Tooth Mousse™ and Mi Paste Plus™ were applied for 180 seconds every day. The volume of demineralization was measured with transverse microradiography. Six lesion characteristics regarding the lesion depth and mineral content of WSLs were also determined. Results: The application of CPP-ACP and CPP-ACPF had a significant regenerative effect on the WSLs. Compared to Control group 1 and 2 the volume of demineralization after 6 weeks decreased significantly for CPP-ACP (respectively p<0.001 and p<0.001) and CPP-ACPF (respectively p=0.001 and p=0.003). The same trend was observed after 12 weeks. For the CPP-ACPF group, WSL dimensions decreased significantly between 6 and 12 weeks follow-up (p=0.012). The lesion depth reduced significantly after application of CPP-ACP and CPP-ACPF but increased significantly in the Control groups. Mineral content increased for CPP-ACP and CPP-ACPF after an application period of 12 weeks, but this was only significant for CPP-ACP. Conclusions: Long-term use of CPP-ACP and CPP-ACPF in combination with a conventional tooth paste shows beneficial effects in the recovery of in vitro subsurface caries lesions.

Effects of in-office bleaching agent combined with different desensitizing agents on enamel

Abstract Objective: To analyze color change, microhardness and chemical composition of enamel bleached with in-office bleaching agent with different desensitizing application protocols. Materials and Methods: One hundred and seventeen polished anterior human enamel surfaces were obtained and randomly divided into nine groups (n = 13). After recording initial color, microhardness and chemical composition, the bleaching treatments were performed as G1: Signal Professional White Now POWDER&LIQUID FAST 38% Hydrogen peroxide(S); G2: S+Flor Opal/0.5% fluoride ion(F); G3: S+GC Tooth Mousse/Casein Phosphopeptide-Amorphous Calcium Phosphate (CPP-ACP) paste(TM); G4: S+UltraEZ/3% potassium nitrate&0.11% fluoride(U); G5: S+Signal Professional SENSITIVE PHASE 1/30% Nano-Hydroxyapatite (n-HAP) suspension(SP); G6: S-F mixture; G7: S-TM mixture; G8: S-U mixture; G9: S-SP mixture. Color, microhardness and chemical composition measurements were repeated after 1 and 14 days. The percentage of microhardness loss (PML) was calculated 1 and 14 days after bleaching. Data were analyzed with ANOVA, Welch ANOVA, Tukey and Dunnett T3 tests (p<0.05). Results: Color change was observed in all groups. The highest ΔE was observed at G7 after 1 day, and ΔE at G8 was the highest after 14 days (p<0.05). A decrease in microhardness was observed in all groups except G6 and G7 after 1 day. The microhardness of all groups increased after 14 days in comparison with 1 day after bleaching (p>0.05). PML was observed in all groups except G6 and G7 after bleaching and none of the groups showed PML after 14 days. No significant changes were observed after bleaching at Ca and P levels and Ca/P ratios at 1 or 14 days after bleaching (p>0.05). F mass increased only in G2 and G6, 1 day after bleaching (p<0.05). Conclusions: The use of desensitizing agents containing fluoride, CPP-ACP, potassium nitrate or n-HAP after in-office bleaching or mixed in bleaching agent did not inhibit the bleaching effect. However, they all recovered microhardness of enamel 14 days after in-office bleaching.

Influence of biofilm formation on the mechanical properties of enamel after treatment with CPP-ACP crème

Abstract: The study aimed to investigate the effects of bacterial biofilms on changes in the surface microhardness of enamel treated with casein phosphopeptide—amorphous calcium phosphate (CPP-ACP) with and without fluoride. Human enamel blocks with incipient caries-like lesions were divided into four groups of 13: G1: Saliva (Control); G2: fluoride dentifrice (Crest™, 1100 ppm as NaF); G3: CPP-ACP (MI Paste; Recaldent™); and G4: CPP-ACPF (MI Paste Plus; Recaldent™ 900 ppm as NaF). The specimens were soaked in demineralizing solution for 6 h and remineralized in artificial saliva for 18 h alternately for 10 days. The dentifrice was prepared with deionized water in a 1 : 3 ratio (w/w) or applied undiluted in the case of the CPP-ACP group. The surface microhardness (SMH) was evaluated at baseline, after artificial caries, after pH cycling and treatment with dentifrices, and after incubation in media with Streptococcus mutans for biofilm formation. The biofilms were exposed once a day to 2% sucrose and the biofilm viability was measured by MTT reduction. The percentage of change in surface microhardness (%SMHC) was calculated for each block. The data were analyzed by nonparametric test comparisons (α = 0.05). The %SMHC values observed in G2 were different from those of G1, G3, and G4 (p < 0.05). After biofilm formation, %SMHC was positive in G2 and G4 when compared to G1 and G3, but resistance to demineralization after biofilm formation was similar in all groups. In conclusion, the presence of biofilms did not influence the treatment outcomes of anticaries products.

Evaluation of the remineralization capacity of CPP-ACP containing fluoride varnish by different quantitative methods

ABSTRACT Objective The aim of this study was to evaluate the efficacy of CPP-ACP containing fluoride varnish for remineralizing white spot lesions (WSLs) with four different quantitative methods. Material and Methods Four windows (3x3 mm) were created on the enamel surfaces of bovine incisor teeth. A control window was covered with nail varnish, and WSLs were created on the other windows (after demineralization, first week and fourth week) in acidified gel system. The test material (MI Varnish) was applied on the demineralized areas, and the treated enamel samples were stored in artificial saliva. At the fourth week, the enamel surfaces were tested by surface microhardness (SMH), quantitative light-induced fluorescence-digital (QLF-D), energy-dispersive spectroscopy (EDS) and laser fluorescence (LF pen). The data were statistically analyzed (α=0.05). Results While the LF pen measurements showed significant differences at baseline, after demineralization, and after the one-week remineralization period (p<0.05), the difference between the 1- and 4-week was not significant (p>0.05). With regards to the SMH and QLF-D analyses, statistically significant differences were found among all the phases (p<0.05). After the 1- and 4-week treatment periods, the calcium (Ca) and phosphate (P) concentrations and Ca/P ratio were higher compared to those of the demineralization surfaces (p<0.05). Conclusion CPP-ACP containing fluoride varnish provides remineralization of WSLs after a single application and seems suitable for clinical use.

Efeito in situ da utilização de uma goma de mascar com caseína fosfopeptídea fosfato de cálcio amorfo (CPP-ACP) previamente ao desafio erosivo inicial / Effect of CPP-ACP chewing gum previously to erosive demineralization

O efeito protetor da caseína fosfopeptídea fosfato de cálcio amorfo (CPP-ACP) contra a erosão dentária é controverso. Este estudo in situ teve como objetivo investigar a capacidade de uma goma de mascar com CPP-ACP em prevenir uma única desmineralização erosiva. Blocos de esmalte bovino (120) selecionados pela dureza superficial inicial foram divididos aleatoriamente entre os grupos: GI - goma de mascar com CPP-ACP, GII - goma de mascar sem CPP-ACP e GIII - controle negativo para avaliação do efeito protetor sem estimulação salivar (sem goma de mascar). Dezenove voluntários participaram do estudo durante três fases cruzadas de 2 h cada. Nas fases de GI e GII os voluntários usaram dispositivos intrabucais palatinos contendo 2 blocos de esmalte, durante 120 minutos e mascaram uma unidade da goma de mascar correspondente ao grupo nos últimos 30 minutos. No grupo controle os voluntários usaram o dispositivo intrabucal por 2h, sem uso de goma de mascar. Em cada fase, imediatamente após a utilização, os dispositivos intrabucais foram imersos em refrigerante tipo cola durante 5 minutos para promover a desmineralização erosiva. A dureza superficial final foi mensurada e os valores foram utilizados para o cálculo do percentual de perda de dureza. Os dados foram analisados por ANOVA de medidas repetidas e teste Tukey (α = 5%). Menor perda de dureza do esmalte foi encontrada após a utilização de goma de mascar com (GI - 32,7%) e sem (GII - 33,5%) CPP-ACP relação ao efeito salivar sem estimulação (GIII - 39,8%) (p <0,05). Não houve diferença entre GI e GII (p> 0,05). Os resultados sugerem que a utilização de goma de mascar imediatamente antes de uma desmineralização erosiva é capaz de diminuir a perda de dureza do esmalte. No entanto, a presença de CPP-ACP na goma de mascar não foi capaz de melhorar este efeito.

The erosion-protective effect of CPP-ACP is controversial. This in situ study aimed to investigate the ability of CPP-ACP chewing gum to prevent a single event of erosive demineralization. Bovine enamel blocks (120), after selection (initial surface hardness) were randomly assigned to groups: GI-chewing gum with CPP-ACP, GIIchewing gum without CPP-ACP and Control group-salivary effect without stimulation (no gum). Nineteen volunteers participated on this study during 3 crossover phases of 2 h. On GI and GII, the volunteers wore intraoral palatal devices for 120 min and chewed a unit of the corresponding chewing gum on the last 30 min. On Control group the volunteer wore the appliance for 2 h, without chewing gum. On each phase immediately after the intraoral use, devices were extra orally immersed in cola drink for 5 minutes to promote erosive demineralization. The percentage of surface hardness loss was calculated. The data were analyzed by Repeated Measures ANOVA and Turkeys test. Less enamel hardness loss was found after the use of chewing gum with (GI-32.7 %) and without (GII-33.5%) CPP-ACP when compared to salivary effect without stimulation (control- 39.8%) (p<0.05). There was no difference between GI and GII (p>0.05). The results suggest that the use of chewing gum immediately before an erosive demineralization is able to diminish the enamel hardness loss. However, the presence of CPP-ACP in the chewing gum cannot enhance this protective effect.

Efeito in situ da utilização de uma goma de mascar com caseína fosfopeptídea fosfato de cálcio amorfo (CPP-ACP) previamente ao desafio erosivo inicial / Effect of CPP-ACP chewing gum previously to erosive demineralization

O efeito protetor da caseína fosfopeptídea fosfato de cálcio amorfo (CPP-ACP) contra a erosão dentária é controverso. Este estudo in situ teve como objetivo investigar a capacidade de uma goma de mascar com CPP-ACP em prevenir uma única desmineralização erosiva. Blocos de esmalte bovino (120) selecionados pela dureza superficial inicial foram divididos aleatoriamente entre os grupos: GI - goma de mascar com CPP-ACP, GII - goma de mascar sem CPP-ACP e GIII - controle negativo para avaliação do efeito protetor sem estimulação salivar (sem goma de mascar). Dezenove voluntários participaram do estudo durante três fases cruzadas de 2 h cada. Nas fases de GI e GII os voluntários usaram dispositivos intrabucais palatinos contendo 2 blocos de esmalte, durante 120 minutos e mascaram uma unidade da goma de mascar correspondente ao grupo nos últimos 30 minutos. No grupo controle os voluntários usaram o dispositivo intrabucal por 2h, sem uso de goma de mascar. Em cada fase, imediatamente após a utilização, os dispositivos intrabucais foram imersos em refrigerante tipo cola durante 5 minutos para promover a desmineralização erosiva. A dureza superficial final foi mensurada e os valores foram utilizados para o cálculo do percentual de perda de dureza. Os dados foram analisados por ANOVA de medidas repetidas e teste Tukey (α = 5%). Menor perda de dureza do esmalte foi encontrada após a utilização de goma de mascar com (GI - 32,7%) e sem (GII - 33,5%) CPP-ACP relação ao efeito salivar sem estimulação (GIII - 39,8%) (p <0,05). Não houve diferença entre GI e GII (p> 0,05). Os resultados sugerem que a utilização de goma de mascar imediatamente antes de uma desmineralização erosiva é capaz de diminuir a perda de dureza do esmalte. No entanto, a presença de CPP-ACP na goma de mascar não foi capaz de melhorar este efeito...

The erosion-protective effect of CPP-ACP is controversial. This in situ study aimed to investigate the ability of CPP-ACP chewing gum to prevent a single event of erosive demineralization. Bovine enamel blocks (120), after selection (initial surface hardness) were randomly assigned to groups: GI-chewing gum with CPP-ACP, GIIchewing gum without CPP-ACP and Control group-salivary effect without stimulation (no gum). Nineteen volunteers participated on this study during 3 crossover phases of 2 h. On GI and GII, the volunteers wore intraoral palatal devices for 120 min and chewed a unit of the corresponding chewing gum on the last 30 min. On Control group the volunteer wore the appliance for 2 h, without chewing gum. On each phase immediately after the intraoral use, devices were extra orally immersed in cola drink for 5 minutes to promote erosive demineralization. The percentage of surface hardness loss was calculated. The data were analyzed by Repeated Measures ANOVA and Turkeys test. Less enamel hardness loss was found after the use of chewing gum with (GI-32.7 %) and without (GII-33.5%) CPP-ACP when compared to salivary effect without stimulation (control- 39.8%) (p<0.05). There was no difference between GI and GII (p>0.05). The results suggest that the use of chewing gum immediately before an erosive demineralization is able to diminish the enamel hardness loss. However, the presence of CPP-ACP in the chewing gum cannot enhance this protective effect...

The effect of CPP-ACP and Nd:YAG laser on the bond strength of softened dentin

The purpose of this study was to investigate the effect of CPP-ACP treatment and Nd:YAG laser on microtensile bond strength (µTBS) of softened dentin. Sixty samples were obtained from thirty sound third molars. All samples were submitted to dentin softening procedure, by the immersion of the specimens in 30 mL of Sprite Zero for 30min. Afterwards, the samples were randomly divided according to the CPP-ACP treatment: CG-Control group; MP-treated with CPP-ACP paste (MI Paste); MPP-treated with CPP-ACP+900 ppm NaF paste (MI Paste Plus). Each group was further divided according to bonding procedure: NL-No laser; L–Laser irradiation after adhesive application and before polymerization. The laser parameters used were 1.4 W, 10 Hz, 140 mJ/pulse, with an optic fiber of 320 µm, generating energy of 174 J/cm2 per pulse. All samples were restored with Clearfil SE Bond/Filtek Z350 XT. After 24 h, the restored samples were cut into beams (± 1 mm2adhesive interface area) and subjected to a µTBS test. Data were analyzed by two-way ANOVA test and Holm-Sidak post-hoc method (α = 0.05). The treatment with CPP-ACP pastes did not significantly affect softened dentin µTBS (p = 0.070). Statistic revealed significant reduction on µTBS values for CG/L, leading to the rejection of the second null hypothesis (p < 0.001). Both CPP-ACP based pastes did not affect µTBS of softened dentin for the adhesive system utilized. The Nd:YAG laser irradiation after application of adhesive system did affect µTBS values of softened dentin samples untreated with CPP-ACP based pastes.

A preliminary study of enamel remineralization by dentifrices based on RecaldentTM (CPP-ACP) and NovaMin® (calcium-sodium-phosphosilicate)

The purpose of this study was to investigate the enamel remineralization potential of two toothpastes, one of which was based on RecaldentTM (CPP- ACP) and the other on NovaMin® (Calcium-sodium-phosphosilicate). Human permanent molar teeth were subjected to three consecutive demineralization cycles. These cycles were followed by remineralization of the experimental groups by toothpastes containing RecaldentTM and NovaMin® respectively. The samples were analyzed by Scanning Electron Microscope, (SEM) and energy-dispersive X-ray spectroscopy analysis (EDX). Extensive demineralization was noted in the control group (without remineralization) while the groups treated with the dentifrices demonstrated various degrees of remineralization, as shown by formation of different types of deposits on the enamel surface. The EDX analysis showed increased amounts of Ca, P, Si and Zn in the enamel of the experimental groups, compared to the control one. Toothpastes containing RecaldentTM and especially NovaMin® have the potential to remineralize enamel, a property which might be important in finding a substitute to pit and fissure sealing.

El objetivo del trabajo fue investigar el potencial de remineralizacion del esmalte de dos pastas dentifricos, una de ella con formulacion basada en RecaldentTM (CPP- ACP) y la otra en NovaMin® (phosphosilicato de calcio y sodio). Se realizaron tres ciclos cosecutivos de desmineralizacion en molares permanentes humanos, seguidos de remineralizacion, en los grupos experimentales con los denti fricos que contenian RecaldentTM y NovaMin® respectivamente. Se analizaron las muestras con microscopia electronica de barrido (SEM) y analisis espectroscopico por dispersion de rayos X (EDX). En el grupo control (sin remineralizacion) se observo una extensa demineralizacion mientras que los grupos tratados con los dentifricos mostraron varios grados de remineralizacion, evidenciados por la formacion de diferentes tipos de depositos sobre la superficie del esmalte. El analisis EDX mostro cantidades aumentadas de Ca, P, Si y Zn en los grupos tratados en comparacion con el grupo control. Los dentifricos conteniendo RecaldentTM y especialmente Nova- Min®, tienen potencial de reminalizacion del esmalte, una propiedad que puede resultar importante como substituto del sellado de fosas y fisuras.

Properties of alkaline protease genetically engineered on cell surface of the yeast Yarrowia lipolytica.

ALP2 gene encoding alkaline protease cloned from Aureobasidium pullulans HN2-3 was ligated into the surface display plasmid and expressed in the cells of the yeast Yarrowia lipolytica. The expressed alkaline protease was immobilized on the yeast cells. The activity of the immobilized enzyme with 6 His tag was found to be significantly higher than that of without 6 His tag. The immobilized enzyme showed lower optimal temperature and a lower affinity for azocasein than the free enzyme purified from A. pullulans HN2-3. The thermal stability of the immobilized enzyme enhanced and the pH stability decreased, compared to that of the free enzyme.

Response surface optimization of effective medium constituents for the production of alkaline protease from a newly isolated strain of Pseudomonas aeruginosa.

Optimization of the fermentation medium for maximum alkaline protease production was carried out with a new strain of Pseudomonas aeruginosa (B-2). Replacing the protein source/inducer (albumin in place of casein) brought about significant increase in yield after 48 hr of inoculation. Three most effective medium constituents identified by initial screening method of Plackett-Burman were albumin, (NH4)2SO4 and glucose. Central Composite Design (CCD) and Response Surface Methodology (RSM) were used in the design of the experiment and in the analysis of the results. Optimum levels of the effective medium constituents were albumin (6.586%); (NH4)2SO4, 0.164%; and glucose, 6.72%. The alkaline protease production increased from 533460 to 793492 Ul(-1).

Pressure-induced dissociation of casein micelles: size distribution and effect of temperature

Pressure-induced dissociation of a turbid solution of casein micelles was studied in situ in static and dynamic light scattering experiments. We show that at high pressure casein micelles decompose into small fragments comparable in size to casein monomers. At intermediate pressure we observe particles measuring 15 to 20 nm in diameter. The stability against pressure dissociation increased with temperature, suggesting enhanced hydrophobic contacts. The pressure transition curves are biphasic, compatible with a temperature (but not pressure)-dependent conformational equilibrium of two micelle species. Our thermodynamic model predicts an increase in structural entropy with temperature.

Obtenção e caracterização química e nutricional de diferentes concentrados de caseína / Obtention and chemical and nutritional characterization of different bovine casein concentrates

OBJETIVO: O objetivo deste trabalho foi avaliar a composição centesimal, o perfil de aminoácidos e as características nutricionais de três concentrados de caseína, obtidos do leite bovino por diferentes processos. MÉTODOS: Os concentrados de caseína foram analisados pelos seguintes processos: uma caseína comercial, obtida por precipitação ácida seguida de neutralização; caseína obtida pela coagulação enzimática; caseína micelar obtida, respectivamente, pelos processos de microfiltração e diafiltração em membrana. A composição centesimal foi determinada por meio de procedimentos descritos no manual Official Methods of Analysis. O perfil de aminoácidos foi determinado após hidrólise ácida da proteína (HCl 6N, 105ºC, 22h) em auto-analisador de aminoácidos, dotado de coluna de troca catiônica e reação pós-coluna com ninidrina. Os perfis de aminoácidos essenciais dos diferentes concentrados de caseína foram comparados e estão de acordo com o padrão Food and Agriculture Organization/World Health Organization, para crianças de 2 a 5 anos de idade. O valor nutritivo foi determinado em ratos da linhagem Wistar, recém desmamados, por meio dos índices, digestibilidade aparente da proteína, quociente de eficiência líquida da proteína e quociente de eficiência protéica operacional. RESULTADOS: A caseína comercial apresentou maior concentração de proteína (92,0 por cento), que a caseína micelar (86,0 por cento) e o coágulo de caseína (72,0 por cento). Os animais nas dietas com as diferentes fontes de proteína, não apresentaram diferencas significativas quanto ao ganho de peso e ingestão de dieta. Maior digestibilidade (93,8 por cento) foi verificada na caseína comercial, comparada à dos outros dois concentrados (91,0 por cento). CONCLUSÃO: Os concentrados de caseína apresentaram diferenças quanto à composicão centesimal, sendo a caseína comercial superior na concentração protéica. O coágulo de caseína apresentou resultados inferiores aos demais concentrados, quanto aos índices quociente de eficiência líquida da proteína e quociente de eficiência protéica operacional.

Yoghurt for athletic people

Buffaloe's concentrated yoghurt was made by using different levels of Na-caseinate or wheat germ [0.5%, 1.0% and 1.5%]. The obtained concentrated yoghurt of the all treatments had sufficient energy which is suitable to consume during training or sporting time. Concentrated yoghurt fortified with 1.0% Na-caseinate or 0.5% wheat germ gave the highest score for flavour, body and texture and appearance. ATB culture was used in this work

Purification and characterization of a small size protease from Bacillus sp. APR-4.

A thermostable extracellular protease of Bacillus sp. APR-4 was purified by size-exclusion and ion-exchange chromatographic methods and its properties were studied. The purified enzyme had a specific activity of 21,000 U/mg of protein and gave single band on SDS/PAGE with a molecular mass of 16.9 KDa. This protease had an optimal pH of 9 and exhibited its highest activity at 60 degrees C. The enzyme activity was inhibited by EDTA, suggesting the presence of metal residue at the active site. Ca2+ (5 mM) had stabilising effect on the activity of protease, but Cu2+ (5 mM) had inhibitory effect. The enzyme exhibited highest specificity towards casein (1%) and had a Km of 26.3 mg/ml and a Vmax of 47.6 U/mg with casein as a substrate. The stability of this enzyme was evaluated in the presence of some organic solvents and the enzyme was stable in methanol, petroleum ether and ethanol. Detergents (Wheel, Farishta) had stimulatory effect on the activity of this enzyme.