Laboratory evaluation of anti-plaque and remineralization efficacy of sugarless probiotic jelly candy supplemented with natural nano prebiotic additive

Abstract

We evaluated the anti-cariogenic effect of an experimental synbiotic compound containing probiotic Lacticaseibacillus rhamnosus (NRRL B-442)-based jelly candy supplemented with natural prebiotic grape seed extract (GSE) in a nanoemulsion formula on the colonization and establishment of Streptococcus mutans (ATCC 25175) and Actinomyces viscosus (ATTCC 19246) biofilms through counting colony forming units, scanning electron microscopy (SEM), and transmission electron microscopy (TEM). We were then analysing the remineralizing effect of synbiotic jelly candy on human enamel surface lesions using Vickers microhardness testers, atomic force microscopy (AFM), SEM, energy-dispersive X-ray spectroscopy (EDAX), and confocal laser scanning microscopy (CLSM) at three stages (sound, after demineralization, and after pH cycling). We found after 21 days of treatment of the pH-cycled enamel discs with jelly candy for 10 min twice daily, a 68% decrease in S. mutans colony formation, reducing biofilm development, trapping S. mutans visualized in jelly candy under SEM examination, and significantly altering the morphological structure of these bacteria under TEM analysis. For remineralization measurements, statistically significant differences in microhardness integrated mineral loss, and lesion depth through CLSM between demineralization and treatment stages. These findings provide an effective anti-cariogenic synbiotic compound of grape seed extract and probiotic jelly candy with potential remineralizing activity.

Effect of flavonoids from grape seed and cranberry extracts on the microbiological activity of Streptococcus mutans: a systematic review of in vitro studies

“…Bias evaluation was performed according to the parameters established in Table 2 . The main risk of bias was found in that study limitations were either not reported in the study report [ 24 – 26 , 29 , 36 , 37 , 39 , 41 – 43 ] or were described briefly [ 27 , 28 , 31 – 34 , 40 , 44 ]. Secondly, there was a failure to include relevant ethical aspects such as institutional ethics committee approval, or patient informed consent for saliva sample collection was not clearly detailed in the article text, or a declaration of project financing and conflict of interest was absent [ 27 – 29 , 31 – 37 , 39 – 42 , 44 ].…”

“…Of the included studies, three were conducted in Japan [ 24 , 36 , 42 ], seven in the United States [ 25 , 28 , 32 , 33 , 35 , 37 , 39 ], three in India [ 26 , 27 , 29 ], two in Australia [ 7 , 38 ] one in Saudi Arabia [ 30 ], one in Italy [ 31 ], one in France [ 34 ], one in Israel [ 40 ], one in England [ 41 ], one in Poland [ 43 ], and one in Egypt [ 44 ] sixteen were conducted at universities [ 7 , 24 – 26 , 28 , 30 – 35 , 37 – 41 ], and six in other types of institutions [ 27 , 29 , 36 , 42 – 44 ].…”

“…For their part, Duarte et al [ 32 ] and Gregoire et al [ 39 ] used S. mutans WHB 410. Some studies do not specifically report the strain used [ 29 , 43 ] or biofilm formation was performed by collecting saliva from healthy donors [ 7 , 30 ].…”

“…The evaluation of biofilm formation was conducted using various methods and culture media, including Todd Hewitt broth in 96-well cell culture plates [ 24 ]; brain–heart infusion (BHI) broth in 96-well cell culture plates [ 36 ]; BHI broth + 1% glucose in 96-well cell culture plates [ 25 ]; BHI broth + sucrose in 24-well tissue culture plates [ 27 , 38 ], or in 96-well cell culture plates [ 31 , 40 ], or in 6-well tissue culture plates [ 33 ]; Mitis Salivarius agar + tellurite [ 34 ]; tryptone-yeast extract broth [ 44 ] by the addition of sucrose on saliva-coated hydroxyapatite disks [ 28 , 32 , 37 ]; trypticase soy broth (TSB) in 96-well titre plates [ 29 , 42 ], or TSB + 20% glycerol [ 39 ]. In one study [ 43 ], a biofilm growth medium containing a mixture of pasteurized saliva, amino acids, vitamins, nucleotides, inorganic salts, trace elements, and glucose, supplemented with fluid universal medium, and enriched with sucrose and glucose was used. The antibacterial activity against S. mutans was also evaluated by different methods, including inoculation on Mitis salivarius bacitracin agar medium in Petri dishes [ 26 ], using the disc diffusion method (Kirby–Bauer) on agar plates, and the bacterial strains were grown in Mutans-Sanguis agar (Hi media) [ 30 ], or the cultures of S. mutans were inoculated into Mueller–Hinton broth [ 41 ].…”

“…The antibacterial activity against S. mutans was also evaluated by different methods, including inoculation on Mitis salivarius bacitracin agar medium in Petri dishes [ 26 ], using the disc diffusion method (Kirby–Bauer) on agar plates, and the bacterial strains were grown in Mutans-Sanguis agar (Hi media) [ 30 ], or the cultures of S. mutans were inoculated into Mueller–Hinton broth [ 41 ]. Sample storage time ranged between 20 and 48 h; in ten studies the sample was incubated for 24 h [ 24 , 27 , 29 , 32 – 35 , 38 , 40 , 42 ], in nine studies, incubation was done for 48 h [ 7 , 25 , 26 , 30 , 31 , 36 , 41 , 43 , 44 ], and in one for 20 h [ 37 ]. The temperature reported in twenty studies was 37 °C [ 7 , 24 – 26 , 28 – 38 , 40 – 44 ], in one was 35 °C [ 27 ], and another study did not report temperature data [ 39 ].…”