The influence of metal salt on sugar consumption by suspension cells in food models constructed by a sugar and salt aqueous solution was investigated based on mid-infrared spectroscopic analysis. The contaminated suspension cells in the food model could be detected using the spectral feature change that measured the present spectrum subtracted in the initial spectrum. The cells were prepared for growth and although the cell did not grow under the induction period, the cell activation (start of sugar metabolism) was detected on the subtracted spectral behavior before the cell growth. The rough grasp of the spectral change behavior is useful for the high-throughput spectroscopic method to detect the contaminated cell activation. Furthermore, the detailed sugar consumption kinetics of the cells was also investigated based on the spectroscopic method. The kind of added salt in the food model influenced the cell activation and the potassium ions play an important role in the plant cells. The living cells activity in fresh food may act to prevent microbial contamination and to suppress the growth of the contaminated microorganism. Both the simple and detailed analyses based on the spectroscopic method presented in this study might be useful for risk management of food.
Microbial contamination is one of the serious problems for risk management of food products. In general, sugar-rich, salt-rich, or fermented foods are processed to prevent microbial contamination. However, consumers tend to like low-salt (sodium-restricted) and low-sugar foods for health care [
The application of spectroscopy, especially in the infrared region, to these measurements has a high potential. Incidentally, in a parallel study using Fourier transform infrared (FT-IR) spectrometers and attenuated total reflection (ATR) techniques [
The sugar aqueous solution with salts is one of the simple models used as a food product. However, it is important that the food models are prepared by including the living cells because most of the flesh food will include a living cell in the body, or the exogenous cells will invade the food. We also studied the quantitative analysis and kinetic analysis of sugar metabolism for plant cells in the basal MS salts media [
The objective of the present study was to investigate the salt influence on the sugar consumption by suspension cells based on spectroscopic analysis. In the study, we supposed the MS cultivation containing the TBY-2 suspension cells to be a contaminated food model and development of the detection method for the activated cells before the cell growth phase (induction period) was investigated based on the spectral feature change for application of the high-throughput spectroscopy. In addition, more detailed analysis of sugar consumption kinetics of the plant cells when the salt conditions changed was investigated based on the MIR spectral analysis. The activity of the exogenous cells that invade the food will be repressed by the living cells in the food. Evaluation of the salt influence on the sugar consumption of the TBY-2 cells as the plant cells composed in part of the food was investigated and discussed.
The Murashige-Skoog (MS) medium [
Composition of basal MS medium and modified MS media.
Components | Basal MS | MS (K→Na) | MS (Na→K) | MS (Na) | MS (K) |
---|---|---|---|---|---|
Ammonium nitrate | 1.65 × 103 | 1.65 × 103 | 1.65 × 103 | 1.65 × 103 | 1.65 × 103 |
Boric acid | 6.20 | 6.20 | 6.20 | 6.20 | 6.20 |
Potassium nitrate | 1.90 × 103 | 1.90 × 103 | 1.90 × 103 | ||
Sodium nitrate | 1.56 × 103 | 1.56 × 103 | |||
Potassium dihydrogen phosphate | 1.70 × 102 | 1.70 × 102 | 1.70 × 102 | ||
Sodium dihydrogen phosphate | 1.50 × 102 | 1.50 × 102 | |||
Potassium iodide | 8.30 × 10−1 | 8.30 × 10−1 | 8.30 × 10−1 | ||
Sodium iodide | 7.50 × 10−1 | 7.50 × 10−1 | |||
Disodium ethylenediaminetetraacetate | 3.73 × 101 | 3.73 × 101 | 3.73 × 101 | ||
Dipotassium ethylenediaminetetraacetate | 4.05 × 101 | 4.05 × 101 | |||
Disodium molybdate dihydrate | 2.50 × 10−1 | 2.50 × 10−1 | 2.50 × 10−1 | ||
Dipotassium molybdate dihydrate | 2.50 × 10−1 | 2.50 × 10−1 | |||
Calcium chloride dihydrate | 4.40 × 102 | ||||
Cobalt chloride hexahydrate | 2.50 × 10−2 | ||||
Sodium chloride | 1.75 × 102 | ||||
Potassium chloride | 2.23 × 102 | ||||
Magnesium sulfate heptahydrate | 3.70 × 102 | 3.70 × 102 | 3.70 × 102 | ||
Manganese sulfate monohydrate | 2.23 × 101 | 2.23 × 101 | 2.23 × 101 | ||
Zinc Sulfate heptahydrate | 8.60 | 8.60 | 8.60 | ||
Copper(II) sulfate pentahydrate | 2.50 × 10−2 | 2.50 × 10−2 | 2.50 × 10−2 | ||
Iron(II) sulfate heptahydrate | 2.78 × 101 | 2.78 × 101 | 2.78 × 101 | ||
Sodium sulfate | 2.47 × 102 | ||||
Potassium sulfate | 3.01 × 102 | ||||
Glycine | 2.00 | 2.00 | 2.00 | 2.00 | 2.00 |
Myoinositol | 1.00 × 102 | 1.00 × 102 | 1.00 × 102 | 1.00 × 102 | 1.00 × 102 |
Thiamin hydrochloride | 1.00 × 10−1 | 1.00 × 10−1 | 1.00 × 10−1 | 1.00 × 10−1 | 1.00 × 10−1 |
Pyridoxine hydrochloride | 5.00 × 10−1 | 5.00 × 10−1 | 5.00 × 10−1 | 5.00 × 10−1 | 5.00 × 10−1 |
Nicotinic acid | 5.00 × 10−1 | 5.00 × 10−1 | 5.00 × 10−1 | 5.00 × 10−1 | 5.00 × 10−1 |
2,4-Dichrorophenoxy acetic acid | 2.00 × 10−1 | 2.00 × 10−1 | 2.00 × 10−1 | 2.00 × 10−1 | 2.00 × 10−1 |
Sucrose | 3.00 × 104 | 3.00 × 104 | 3.00 × 104 | 3.00 × 104 | 3.00 × 104 |
Unit: mg dm−3.
The dry cell weight in the medium was compared by measuring the turbidity (OD 600 nm). The turbidity was measured by a UV-VIS-NIR scanning spectrophotometer (UV-3100PC; Shimadzu, Kyoto, Japan) using a cuvette with a light path length of 10 mm.
The sugar and ethanol concentrations in the culture medium were determined by the FT-IR/ATR method as described previously [
The logistic function expressed by (
Figure
Time behavior of the FT-IR/ATR spectra of the basal MS medium (a) and of the modified MS salt media represented by MS_Na (b), MS_K→Na (c), MS_K (d), and MS_Na→K (e) during cultivations.
During the cultivation, the behavior of the spectral changes was able to be classified in two patterns. The absorbance decreased and the spectral pattern was changed during the cultivation (Figures
Figure
Time courses of the pH, cell, and total sugar concentrations during cultivation using a basal MS medium (a) and modified MS media represented by MS_Na (b), MS_K→Na (c), MS_K (d), and MS_Na→K (e). The dot-dash and dash lines in (c) and (e) show the growth and total sugar profile from (a), results of the basal MS medium cultivation.
In the basal MS cultivation, the inoculum suspension cell was grown after a few days and the growth was stopped by 9 days of cultivation (Figure
Several models for different states of the contaminated cells were constructed by changing the salt condition as shown in Figure
Subtracted spectra at 2 days of cultivation which were obtained after spectral subtraction of 0 days of absorption and the standard deviation (SD) of the six spectra of the 0 days of cultivation using a basal MS medium.
Figure
Time courses of sugar and ethanol concentrations (a), amount of the glucose and fructose consumptions (b), consumption rate (c), and specific consumption rate (d) of basal MS cultivation.
Figure
Time courses of consumption and specific consumption rates of glucose, fructose, and total sugar in cultivations using basal MS ((a), (d)), MS_K→Na ((b), (e)), and MS_Na→K ((c), (f)) media.
Figure
Time behavior of nondimensional time of consumption and specific consumption rates of glucose, fructose, and total sugar in cultivations using basal MS ((a), (d)), MS_K→Na ((b), (e)), and MS_Na→K ((c), (f)) media.
In the MS_Na→K cultivation, the cell growth and the sugar consumption profiles were almost the same as those of the basal MS cultivation; however, drastic changes in the sugar kinetics were observed. In particular, the peak of the sugar specific consumption rate was shifted to the negative period of the nondimensional time more clearly. However, the cell growth was not suppressed in this situation. The reason for this phenomenon is not clear, but the behaviors of the consumption of the glucose and the fructose were slightly changed in comparison with the case of the basal MS cultivation. The sugar specific consumption rate of the MS_Na→K cultivation was faster than that of the basal MS cultivation. In general, most living cells are easy to utilize glucose than fructose. In all cases, the clear peak of the glucose consumption and specific consumption rates were observed. Therefore, the sugar metabolism of glucose will not be influenced under the changing salt conditions in medium. The peak shape of the fructose consumption rate depended on the kind of salt in the medium, accordingly it may have given the change for sugar metabolism behavior. In particular, cell growth in the early period of the cultivation was significantly suppressed in the MS_K→Na cultivation.
The living cells activity in fresh food may act to prevent microbial contamination and to suppress the growth of the contaminated microorganism. Sodium-restricted food (potassium chloride is used as a food additive for the replacement of sodium chloride) will be advantageous to keeping the living cells activity in the food. The kind of salt added in the food influenced the cell activation and the potassium ions play an important role in the living plant cells.
Physiological influences of the cations for plants were reported in several studies [
The activity of the cell which invaded the food model was changed by the added salt changing, using the modified MS medium. In addition, the cell activation (start of the sugar metabolism) could be detected by checking if the spectral feature changed before the start of the cell growth. The measurement and recode of the subtracted spectral data were useful to checking the microbial contamination and the evaluation of the cell activity in food for risk management of the contamination. The rough grasp of the behavior of the spectral change is important for the high-throughput spectroscopic method. The spectral information containing the metabolite components of the activated cells and detailed analysis of the sugar consumption kinetics of the suspension cells could be investigated by spectroscopic method. Both the simple and detailed analyses based on the spectroscopic method presented in this study might be useful for evaluation of the activity of living cells in the food. The rough grasp of the behavior of the spectral change is important for the high-throughput spectroscopic method. Checking the spectral feature changes is useful for the daily check of the activation of the contaminated cells to avoid the contamination. The spectroscopic analysis will be greatly useful if the detailed information of the contaminated cells is necessary. Thus, the proposed methods will be applied to the risk management of various situations of microbial contamination in food.
This work was partly supported by a Grant-in-Aid for Scientific Research (C) 20560729 from the JSPS KAKENHI and by Mie University Research Center of Science and Technology for Social Security.