*2.7. Statistical Analysis*

Data were expressed as the mean ± standard deviation (SD). Data were subjected to one-way ANOVA, and significance differences were analyzed using SPSS version 19.0 (IBM Company, Armonk, NY, USA).

#### **3. Results and Discussion**

#### *3.1. Cell Viability Assay*

Antibiotics easily cause some side effects in the treatment of diseases, such as antibioticassociated diarrhea [23]. Even though it is the bacteriocin approved by JECFA, nisin could also bring some adverse effects to the gut microbiota of the body [24]. As shown in Figure 1, with the experimental concentrations of GCP from 100 μg/mL to 600 μg/mL, cell viability increased from 101 ± 0.8% to 115 ± 1.3% (*p* < 0.05), suggesting that GCP was not toxic to Caco-2 cells. This toxicity result was similar to that of *Ganoderma lucidum* polysaccharide reported in our earlier work [21] and in works by other researchers. For example, Zhang et al. [25] found that the alkali-soluble polysaccharides from *Arctium lappa* L. had no toxicity to RAW264.7 cells. Caillot et al. [26] reported that the blackberry wine polysaccharides had no toxicity to RAW 264.7 macrophages. Meanwhile, during the whole experiment, there were no signs of disease or death in mice, indicating the security of GCP to mice. Furthermore, He et al. [27] demonstrated that a novel polysaccharide produced by *Streptomyces Virginia* H03 was safe to mice when administered at doses of 500 mg/kg/day. Therefore, the concentrations of GCP used in the following tests were 100 μg/mL, 200 μg/mL and 400 μg/mL, respectively.

**Figure 1.** Cell viability assay of GCP to Caco-2 cells (*n* = 5). Significance was determined through ANOVA, \* *p* < 0.05, \*\* *p* < 0.01.

#### *3.2. Effect of GCP on the Body Weight of Normal Mice*

As can be seen from Table 1, the administration of GCP decreased the average body weight of mice, and the body weight gain of mice significantly (*p* < 0.05) decreased from 10.5 ± 1.21 g to 8.4 ± 1.17 g. Chen et al. [23] reported that *Pueraria lobata* polysaccharide could effectively reduce the average body weight of mice. Meanwhile, Tian et al. [28] and Yin et al. [6] demonstrated that *Lycium ruthenicum* and resveratrol had similar weight loss effects as those seen in this work. However, Yang et al. [29] and Wei et al. [30] found that flaxseed polysaccharides and *Musa basjoo* had almost no effects on the weight loss of mice, which was inconsistent with the results of this work. The weight gain control of GCP might relate to the metabolism of acetate, alanine aminotransferase and aspartate aminotransferase [6,28], and this will be analyzed in the following work.


**Table 1.** Effect of GCP on the body weight of mice (*n* = 10).

Different letters (a–e) represent significant differences, *p* < 0.05.

#### *3.3. Effect of GCP on the Serum Biochemistry of Normal Mice*

The effects of GCP on physiological parameters in the serum of mice are shown in Table 2. There was a decreasing trend (*p* < 0.05) in the concentration of aspartate transaminase and alanine aminotransferase in the mice serum after the administration of GCP; the aspartate transaminase concentration decreased from 38.5 ± 3.37 U/L to 35.4 ± 1.87 U/L, and the alanine aminotransferase concentration reduced from 121.5 ± 6.43 U/L to 105.4 ± 10.91 U/L. Meanwhile, Guo et al. [31] and Yin et al. [6] reported that the weight loss effects of nanobubble water and resveratrol were partly related to the reduction inaspartate transaminase and alanine aminotransferase in the body, which could partly explain why the GCP reduced the weight of the mice in Table 1. However, the activities of total protein, albumin, globulin, urea, high-density lipoprotein, low-density lipoprotein and the glucose in the mice serum among the four groups underwent little change, suggesting that GCP had no toxicity to mice.

**Table 2.** Effect of GCP on the serum biochemistry of mice (*n* = 10).


Different letters (a–d) represent significant differences, *p* < 0.05.
