Carbohydrate Metabolism

The metabolism of several carbohydrates by sourdough LAB strains was determined by using API 50 CHL galleries (BioMérieux, Marcy-l’Etoile, France) according to the manufacturer instructions. Gas production was detected by Durham tube method [37 ] in MRS broth (Biolife, Milan, Italy) for 24 h at 30 °C. The growth performance of strains was monitored at 10, 30, 37 and 45 °C for 24 h in an MRS broth using a Thermo Bioscreen C automatic turbidometer (Labsystems, Helsinki, Finland). The viability of the isolated strains to grow in acidic environments was evaluated in MRS broth acidified to a final pH of 2.5 with HCl (Biolife, Milan, Italy) in tubes, according to Lee et al. [38 (link)]. Total viable counts were determined by using standard plate count techniques [39 ]. The results were expressed as log of colony-forming units (CFU) per milliliter. All phenotype analyses were carried out in triplicate.

In the present study, the oxidation reaction of glucose, palmitic acid, and an average of 20 amino acids was chosen to represent the metabolism of carbohydrates, lipids, and protein, respectively, and calculations were performed on the basis of reactions Equations (2)–(4). All the calculations for NAFLD patients with different Child–Pugh Scores and healthy people were performed by using the data given in Table 4 based on the planned diet plans shown in Table 5. According to the procedure described by Öngel et al. [10 (link)], the entropy generation rate and lifespan estimation of the patients were calculated. It is assumed that oxidation of protein, fat, and carbohydrate is 28%, 46%, and 88% for the patient in Child–Pugh Score A, 31%, 55%, and 86% for the patients in Child–Pugh Score B, and 24%, 59%, and 88% for the patient with Child–Pugh Score C, respectively [56 (link)]. However, it was considered that during the disease, the patients in Child–Pugh Scores A, B, and C lost 5 kg, 10 kg, and 20 kg weight, respectively, due to complications. For a healthy person, these rates are 92%, 95%, and 99% for protein, fat, and carbohydrate, respectively [54 (link)]. On the basis of the entropy balance equation (Equation (7)) by Özilgen and Sorgüven [57 ] and Kuddusi [45 (link)], the entropy generation rate and lifespan estimation were found. It was assumed that $\frac{d{\left[m s\right]}_{system}}{dt}$ equals zero because the thermodynamic system, the body, is in a quasi-steady-state condition. To calculate the entropy generation rate during NAFLD (S_cirrhotic), it was assumed that patients maintained a healthy life until they reached age 40 and consumed the diet of a healthy person, and then they became NAFLD patients; therefore, they started consuming a special diet for each Child–Pugh Score (Table 5). The lifespan entropy generation limit was 11,404 kJ/K kg in the calculations of Kuddusi [45 (link)]. The one-year survival rate is approximately 95%, 80%, and 44% for the patients with Child–Pugh Scores of A, B, and C, respectively [57 ]. Öngel et al. [10 (link)] estimated the disease-related entropy generation for 19 different varieties of cancer as:
The same expression is employed in this study for the NAFLD, and the remaining average lifetime (t_avg) was estimated for each Child–Pugh Score patient and listed in Table 7. If persons are diagnosed with Child–Pugh Score C NAFLD at the age of 40, they would have already generated 4796 kJ/kg K entropy and may generate 6604 kJ/kg K of more entropy during the rest of their lifespan. Pinter et al. [58 (link)] indicated that 95%, 80%, and 44% of the NAFLD patients with Child–Pugh Score A, B, and C will survive the first year of the disease; therefore, when compared with the 119.9 kJ/kg K year of entropy generation rate of an obese (otherwise healthy) person, we may estimate that, on average, a Child–Pugh Score A patient may generate (119.9 kJ/kg K) [1/(0.95)]= 126.2 kJ/kg K of entropy annually. Similarly, Child–Pugh Score B and C patients may generate 149.9 and 272.5 kJ/kg K of entropy annually, respectively (Table 8).

A total of 240 1-day-old male Arbor Acre broilers (body weight, 42.62 ± 0.82 g) were randomly allocated to three groups. Each group consisted of eight replicates (pens) with 10 broilers per pen. Two phase non-medicated basal diets in mashed form were formulated based on the nutrient requirements of the National Research Council (1994) ; (Table 1). The three groups included basal diet (CT, n = 8), and basal diet with a dose of 1.0 × 10⁸ CFU/kg (BCG1, n = 8) and 1.0 × 10⁹ CFU/kg (BCG2, n = 8) B. licheniformis BCG, respectively (Wang et al., 2017 (link); Kan et al., 2021 (link); Xu et al., 2021 (link)). All broilers were feed in wire-floored cages in a one-level battery on their respective diets. The study lasted 42 days, during which time broilers had ad libitum to feed and fresh water. Broilers were housed in an environmentally controlled room and temperature was gradually reduced from 35°C on day 1 to 26°C at day 21 and then kept roughly constant. A 20 h light-4 h dark cycle was carried out throughout the experimental period. B. licheniformis BCG was isolated from humus soil in the northeast forest area and preserved in the Key Laboratory of Feed Biotechnology of Ministry of Agriculture and Rural Affairs. It presents great biological characteristics in carbohydrate metabolism enzymes and stress tolerance through the whole genome sequencing and in vitro evaluation. B. licheniformis BCG with viable count = 1.08 × 10¹⁰ CFU/g was used and mixed in the basal diet, which was prepared in bacterial mashed form after processed in activation, culture, centrifugation, freeze-drying, and grinding.