Quantitative method for the detection and quantification of galacto-oligosaccharides
A technology of galacto-oligosaccharides and buffer solutions, which is applied in measuring devices, immunoassays, and food testing, can solve problems such as inability to separate, and achieve the effect of quality assurance
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Embodiment 1
[0066] Example 1: PMP derivatives using C30 column - HPLC reverse phase chromatogram
[0067] In the same manner as in Comparative Example 1, a sample solution of prepared milk powder for childcare, a placebo sample solution, a galacto-oligosaccharide standard test solution, and a maltodextrin standard test solution were prepared. The galacto-oligosaccharide standard test solution was further prepared into dilutions diluted 10, 20, and 40 times with distilled water. The concentration of galacto-oligosaccharide liquid sugar in the diluent is 0.2g / 100ml, 0.1g / 100ml, 0.05g / 100ml respectively.
[0068] (1) Derivatization of PMP
[0069] Take 100 μl each of the sample solution, galacto-oligosaccharide standard test solution, and maltodextrin standard test solution in a screw-top test tube, add 100 μl of 0.6 M NaOH aqueous solution, and stir. Next, 200 μl of a 0.5 M PMP methanol solution was added and stirred. PMP derivatization was carried out by heating at 70°C for 30 minutes. ...
Embodiment 2
[0108] Example 2: Discussion of eluents in HPLC
[0109] Similar to Comparative Example 2, sample solutions (milk powder 101 (with GOS added), milk powder 102 (placebo)), galacto-oligosaccharide standard test solution and maltodextrin standard test solution were prepared, and each PMP was prepared in the same manner as in Comparative Example 2. Derivatized and analyzed using the following 6 HPLC conditions.
[0110] (1) HPLC under the condition of buffer pH4
[0111] Column: Develosil RPAQUEOUS (4.6×250mm)
[0112] Eluent: potassium phosphate buffer (pH4) / CH 3 CN(78 / 22)
[0113]Flow rate: 1.0ml / min
[0114] Detection (Detection): UV245nm
[0115] Column temperature (Column temp): 35°C
[0116] (2) HPLC under the condition of buffer pH5
[0117] Column: Develosil RPAQUEOUS (4.6×250mm)
[0118] Eluent: potassium phosphate buffer (pH5) / CH 3 CN(79 / 21)
[0119] Flow rate: 1.0ml / min
[0120] Detection (Detection): UV245nm
[0121] Column temperature (Column temp): 35°C ...
Embodiment 3
[0151] Example 3: PMP derivative-reversed-phase HPLC chromatogram (C30) in the case of nutritious food as a sample
[0152] The raw materials shown in Table 4 were mixed to prepare a nutritional food. After adding distilled water to 10 g of this nutritional food to make 50 ml, centrifugation was performed at 20000 g x 30 minutes, and further centrifugation was performed at 20000 g x 30 minutes. After centrifugation, the nutritious food was separated into three layers of the lower layer, the middle layer, and the upper layer. The intermediate layer was separated, and the separated sample was filtered through a 0.45 μm filter. On the other hand, a galacto-oligosaccharide standard test solution as a standard sample was prepared in the same manner as in Comparative Example 1 using OLIGOMATE 55 (Oligomet 55) (Yakult Pharmaceutical Co., Ltd.). Take 100 μl each of the filtered sample and galacto-oligosaccharide standard test solution in a screw-top test tube, and dry them under red...
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