**4. Safety evaluation of bamboo vinegars**

#### **4.1. Cytotoxicity of bamboo vinegars**

One of the cytotoxicity results for the bamboo vinegars collected at temperatures of 90–92°C with the original vinegar (no diluting) and a range of diluting percent contents of 50.00, 33.33, 25.00, 20.00, 13.33 and 10.00% for *S. typhimurium* TA98 and TA100 without S9 mix, as well as for with S9 mix with diluting percent contents of 33.33, 25.00, 20.00, 13.33 and 10.00% is shown in **Table 1**.

The residual bacterial count of the control group (Blank) toward *S. typhimurium* TA98 and TA100 is 1838 and 1500, respectively. For bamboo vinegars collected at temperatures of


1 Survival (%) = (the bacterial count of test group/the bacterial count of control group) × 100. 2 Blank (the control group) was added without bamboo vinegars.

3 Mean.

mutagenicity. Therefore, the bamboo charcoals have no mutagenicity toward *S. typhimurium* TA98 and TA100. Furthermore, Weng reports that the maximum percent weight of Moso bamboo at RH 40% is 6.7% and at RH 90% is 9.9% [39]. The water activity of the bamboo charcoal (0.57), compared to Aw for the growth of a microorganism environment, is below 6.0. The heavy metal contents (as Pb ppm base) of charcoal are below 40 ppm and meet the Sanitation Standard for Edible Natural Colorants, Food Sanitation Standards as well. For the Ames test, the bamboo charcoal has no toxicity and mutagenicity toward *S. typhimurium* TA98 and TA100. In the scope of this report, the above results indicate that the bamboo charcoal can

One of the cytotoxicity results for the bamboo vinegars collected at temperatures of 90–92°C with the original vinegar (no diluting) and a range of diluting percent contents of 50.00, 33.33, 25.00, 20.00, 13.33 and 10.00% for *S. typhimurium* TA98 and TA100 without S9 mix, as well as for with S9 mix with diluting percent contents of 33.33, 25.00, 20.00, 13.33 and 10.00% is shown

**Figure 2.** Mutagenicity of Moso bamboo charcoal toward *S. tyhpimurium* TA98 and TA100 without or with S9 mix.

The residual bacterial count of the control group (Blank) toward *S. typhimurium* TA98 and TA100 is 1838 and 1500, respectively. For bamboo vinegars collected at temperatures of

be expected to be the same as the materials of edible natural colorants.

54 Bamboo - Current and Future Prospects

**4. Safety evaluation of bamboo vinegars**

**4.1. Cytotoxicity of bamboo vinegars**

in **Table 1**.

**Table 1.** Cytotoxicity of bamboo vinegar collected at the temperature of 90–92oC toward *Salmonella typhimurium* TA98 and TA100 without or with S9 mix.

90–92°C, the residual bacterial count without S9 for diverse diluting percent contents is 0–1830 for TA98 and 87–1641 for TA100; for those with S9, it is 1605–2000 for TA98 and 1736–2249 for TA100. Waleh *et al*. indicate that the residual bacteria rate of *S. typhimurium* must be over 80% of the control group to determine that the test group has no cytotoxicity for *S. typhimurium* [36]. The residual bacteria rate (Survival, %) toward TA98 and TA100 for bamboo vinegar without S9 mix at a diluting percent content 33.3% or less, and with S9 mix at 25.00% or less is greater than 80%. In other words, the bamboo vinegars collected at temperatures of 90–92°C has cytotoxicity for *S. typhimurium* at a diluting 33.3% less. For vinegars collected at all different temperatures, the Survivals are shown in **Table 2**.

The Survival of all bamboo vinegars collected from 80 to 150, over 80, 90–92, 99–102, 120–123, and 145–150°C without S9 mix at a diluting percent content of 33.33% or less is all higher than those for Blank by more than 80%. However, the Survival of the bamboo vinegar collected at temperatures of 99–102°C at a diluting percent content of 25% shows that the cytotoxicity toward *S. typhimurium* TA98 and TA100 with S9 mix is lower than 80%, indicating "with toxicity." It is asserted that the Survival of bamboo vinegars collected at all different temperatures at a diluting percent content of 20.00% less for either with or without S9 mix is no cytotoxicity absolutely, and the dose for the mutagenicity test can be selected according to the results (**Table 2**).


and 140–206 for TA100. The results on the same **Table 3** also show that the bamboo vinegars in the test range do not exceed spontaneous revertants by more than two times for TA98 and TA100 without/with S9 mix; that is, the MR is less than 2. The spontaneous revertants of the other bamboo vinegars collect at diverse temperatures, 80–150, over 80, 99–102, 120–123 and 145–150°C, at the ranges of the diluting percent content are smaller than those of Blank by

**Bamboo vinegar (90–92°C)**

−S9 25.00 53 0.81 129 0.98

S9 Blank 76 1.00 140 1.00

MR (mutagenicity ratio) = induced revertants per plate/spontaneous revertants per plate (control).

Blank (the control group) was added without either bamboo or wood vinegars.

**Blank2 653 1.00 132 1.00**

20.00 42 0.64 121 0.91 13.33 53 0.81 109 0.83 10.00 41 0.63 115 0.87

20.00 69 0.90 195 1.39 13.33 73 0.96 204 1.45 10.00 74 0.97 206 1.47

**TA98 MR1 TA100 MR**

Preliminary Safety Evaluation of Bamboo Pyrolysis Products: Charcoal and Vinegar

http://dx.doi.org/10.5772/intechopen.68542

57

The MR of the bamboo vinegars collected at all different temperatures toward *S. typhimurium* TA98 and TA100 without S9 mix for a diluting percent content of 25.00, 20.00, 13.33 and 10.00% or with S9 mix for a diluting percent content of 20.00, 13.33 and 10.00% is shown in **Table 4**. No matter what the vinegars are for all collected temperatures and *S. typhimurium* TA98 and TA100 without/with S9 mix, the MR is less than 2. Neither mutagenicity nor toxicity is observed for bamboo vinegars collected at diverse temperatures toward *S. typhimurium* TA98 or TA100 without/with S9 mix. The bamboo vinegars, therefore, have no mutagenicity

**Table 3.** Mutagenicity of diverse bamboo vinegar collected at the temperature of 90–92°C toward *Salmonella typhimurium*

If mutagenicity occurred in a treated material, the results of the antimutagenic assay would be confused due to increased or decreased numbers of revertants of TA98 and TA100 [42]. Hence, without S9 mix a diluting percent content of 25.00–10.00% and with S9 mix at 20.00–10.00% a

as direct mutagens requiring metabolic activation and indirect acting mutagen, respectively

are used

diluting percent content are selected for the antimutagenic activity. NQNO and AFB<sup>1</sup>

toward *S. typhimurium* TA98 at 25% less and TA100 at 20% less.

**4.3. Antimutagenic activity of bamboo vinegars**

more than two times [38].

**S9 mixture Diluting percent** 

TA98 and TA100 without or with S9 mix.

1

2

3 Mean. **content (%)**

1 Survival (%) = (the bacterial count of test group/the bacterial count of control group) × 100.

**Table 2.** Survival of diverse bamboo vinegars collected at different temperatures toward *Salmonella typhimurium* TA98 and TA100 without or with S9 mix.

#### **4.2. Mutagenicity of bamboo vinegars**

Ames *et al*. report that the number of spontaneous revertants induced by the specimen is less than that for the control group by more than two times; the specimen has no mutagenicity [32]. **Table 3** shows the mutagenicity results for the bamboo vinegar collected at temperatures of 90–92°C with a diluting percent content of 25.00, 20.00, 13.33 and 10.00% for *S. typhimurium* TA98 and TA100 without S9 mix and with S9 for a diluting percent content of 20.00, 13.33 and 10.00%. The spontaneous revertants of Blank are 65 for TA98 and 132 for TA100 and of bamboo vinegars without S9 mix are from 41 to 53 for TA98 and from 109 to 129 for TA100. With S9 mix, the spontaneous revertants of Blank and bamboo vinegars are 69–76 for TA98 and 140–206 for TA100. The results on the same **Table 3** also show that the bamboo vinegars in the test range do not exceed spontaneous revertants by more than two times for TA98 and TA100 without/with S9 mix; that is, the MR is less than 2. The spontaneous revertants of the other bamboo vinegars collect at diverse temperatures, 80–150, over 80, 99–102, 120–123 and 145–150°C, at the ranges of the diluting percent content are smaller than those of Blank by more than two times [38].


1 MR (mutagenicity ratio) = induced revertants per plate/spontaneous revertants per plate (control). 2 Blank (the control group) was added without either bamboo or wood vinegars. 3 Mean.

**Table 3.** Mutagenicity of diverse bamboo vinegar collected at the temperature of 90–92°C toward *Salmonella typhimurium* TA98 and TA100 without or with S9 mix.

The MR of the bamboo vinegars collected at all different temperatures toward *S. typhimurium* TA98 and TA100 without S9 mix for a diluting percent content of 25.00, 20.00, 13.33 and 10.00% or with S9 mix for a diluting percent content of 20.00, 13.33 and 10.00% is shown in **Table 4**. No matter what the vinegars are for all collected temperatures and *S. typhimurium* TA98 and TA100 without/with S9 mix, the MR is less than 2. Neither mutagenicity nor toxicity is observed for bamboo vinegars collected at diverse temperatures toward *S. typhimurium* TA98 or TA100 without/with S9 mix. The bamboo vinegars, therefore, have no mutagenicity toward *S. typhimurium* TA98 at 25% less and TA100 at 20% less.
