Phosphate buffer is very commonly used in the laboratory. Its applications range widely, from enzymatic assays to histology. Wherever it is necessary to maintain a neutral pH under variable conditions, phosphate buffer is ideal. Another major advantage is the ability to control the buffer\u2019s pH and adjust it to specific requirements (e.g. the optimal pH for the activity of particular enzymes).<\/p>\n\n\n\n
There are two types of phosphate buffer. The first is based on sodium salts, the second on potassium salts. Both buffers contain salts of dihydrogen phosphate and hydrogen phosphate.<\/p>\n\n\n\n Oba bufory posiadaj\u0105 podobn\u0105 obj\u0119to\u015b\u0107 buforow\u0105, jednak ich unikalne w\u0142a\u015bciwo\u015bci mog\u0105 wp\u0142ywa\u0107 na ich zastosowanie.<\/p>\n\n\n\n Both the sodium phosphate buffer and the potassium phosphate buffer have similar pH-maintaining capabilities. Under certain specific conditions, their differing properties may significantly affect the test result or the stability of the system.<\/p>\n\n\n\n To prepare a 0.1 M sodium phosphate buffer, you must first prepare 0.2 M solutions of sodium dihydrogen phosphate (V) and sodium hydrogen phosphate (V).<\/p>\n\n\n\n Solution A (0.2 M sodium dihydrogen phosphate, NaH\u2082PO\u2084)<\/strong><\/p>\n\n\n\n Dissolve 27.8 g of NaH\u2082PO\u2084 in 800 ml of water, transfer the solution volumetrically to a 1000 ml measuring flask and make up to the mark.<\/p>\n\n\n\n Solution B (0.2 M sodium hydrogen phosphate, Na\u2082HPO\u2084)<\/strong><\/p>\n\n\n\n Dissolve 53.65 g of Na\u2082HPO\u2084\u00b77H\u2082O or 71.7 g of Na\u2082HPO\u2084\u00b712H\u2082O in 800 ml of water. Transfer to a 1000 ml volumetric flask and make up to the mark.<\/p>\n\n\n\n To obtain a buffer with the correct pH, mix solutions A and B as shown in the table below and make up to a final volume of 200 ml with water.<\/p>\n\n\n\n I recommend preparing the buffer fresh and using it within two weeks. Solutions A and B can be stored for longer and used as starting solutions for preparing the working buffer.<\/p>\n\n\n\n To prepare a 0.1 M potassium phosphate buffer, you must first prepare 0.2 M solutions of potassium dihydrogen phosphate and potassium hydrogen phosphate.<\/p>\n\n\n\n Solution A (0.2 M potassium dihydrogen phosphate, KH\u2082PO\u2084)<\/strong><\/p>\n\n\n\n Dissolve 34.84 g of KH\u2082PO\u2084 in 800 ml of water, transfer the solution by volume to a 1000 ml volumetric flask and make up to the mark.<\/p>\n\n\n\n Solution B (0.2 M potassium hydrogen phosphate, K\u2082HPO\u2084)<\/strong><\/p>\n\n\n\n Dissolve 34.84 g of K\u2082HPO\u2084\u00b73H\u2082O in 800 ml of water. Transfer to a 1000 ml volumetric flask and make up to the mark.<\/p>\n\n\n\n To obtain a buffer with the correct pH, mix solutions A and B as shown in the table below and make up to a final volume of 200 ml with water.<\/p>\n\n\n\n I recommend preparing the buffer fresh and using it within two weeks. Solutions A and B can be stored for longer and used as starting solutions for preparing the working buffer.<\/p>\n\n\n\n Solution calculator: https:\/\/www.sigmaaldrich.com\/PL\/pl\/support\/calculators-and-apps\/buffer-calculator<\/a><\/p>\n\n\n\nSodium-phosphate buffer<\/strong><\/td> Potassium-phosphate buffer<\/strong><\/td><\/tr> Disodium phosphate
Sodium hydrophosphate
pH 5,7 \u2013 8,0<\/td>Potassium dihydrophosphate
Potassium hydrophosphate
pH 5,8 \u2013 8,0<\/td><\/tr><\/tbody><\/table><\/figure>\n\n\n\nWhich phosphate buffer should I choose?<\/h2>\n\n\n\n
Property<\/strong><\/td> Sodium-phosphate buffer<\/strong><\/td> Potassium-phosphate buffer<\/strong><\/td><\/tr> Solubility<\/strong><\/td> At low temperatures and during freezing, it may precipitate, causing cloudiness and the formation of aggregates.<\/td> Better solubility at low temperatures. Does not cause cloudiness or precipitation during freezing.<\/td><\/tr> Interaction with SDS<\/strong><\/td> Does not react with SDS<\/td> Potassium ions react with SDS, leading to precipitation. Many people recommend avoiding this buffer in SDS-PAGE.<\/td><\/tr> pH range<\/strong><\/td> Effective buffering within the pH range of 5.7\u20138.0. Maximum buffering capacity at pH 6.8.<\/td> Effective buffering within the pH range of 5.8\u20138.0. Maximum buffering capacity at pH 6.8.<\/td><\/tr> Mobility (CZE)<\/strong><\/td> It may cause molecules to migrate more quickly<\/td> Slower migration<\/td><\/tr><\/tbody><\/table><\/figure>\n\n\n\n How to prepare a sodium-phosphate buffer?<\/h2>\n\n\n\n
pH<\/td> ml solution A (0,2 M NaH2PO4)<\/td> ml solution B (0,2 M Na2HPO4)<\/td><\/tr> 5,7<\/td> 93,5<\/td> 6,5<\/td><\/tr> 5,8<\/td> 92,0<\/td> 8,0<\/td><\/tr> 5,9<\/td> 90,0<\/td> 10,0<\/td><\/tr> 6,0<\/td> 87,7<\/td> 12,3<\/td><\/tr> 6,1<\/td> 85,0<\/td> 15,0<\/td><\/tr> 6,2<\/td> 81,5<\/td> 18,5<\/td><\/tr> 6,3<\/td> 77,5<\/td> 22,5<\/td><\/tr> 6,4<\/td> 73,5<\/td> 26,5<\/td><\/tr> 6,5<\/td> 68,5<\/td> 31,5<\/td><\/tr> 6,6<\/td> 62,5<\/td> 37,5<\/td><\/tr> 6,7<\/td> 56,5<\/td> 43,5<\/td><\/tr> 6,8<\/td> 51,0<\/td> 49,0<\/td><\/tr> 6,9<\/td> 45,0<\/td> 55,0<\/td><\/tr> 7,0<\/td> 39,0<\/td> 61,0<\/td><\/tr> 7,1<\/td> 33,0<\/td> 67,0<\/td><\/tr> 7,2<\/td> 28,0<\/td> 72,0<\/td><\/tr> 7,3<\/td> 23,0<\/td> 77,0<\/td><\/tr> 7,4<\/td> 19,0<\/td> 81,0<\/td><\/tr> 7,5<\/td> 16,0<\/td> 84,0<\/td><\/tr> 7,6<\/td> 13,0<\/td> 87,0<\/td><\/tr> 7,7<\/td> 10,5<\/td> 90,5<\/td><\/tr> 7,8<\/td> 8,5<\/td> 91,5<\/td><\/tr> 7,9<\/td> 7,0<\/td> 93,0<\/td><\/tr> 8,0<\/td> 5,3<\/td> 94,7<\/td><\/tr><\/tbody><\/table><\/figure>\n\n\n\n How to prepare a potassium-phosphate buffer?<\/h2>\n\n\n\n
pH<\/td> ml solution A (0,2 M KH2<\/sub>PO4<\/sub>)<\/td> ml solution B (0,2 M K2<\/sub>HPO4<\/sub>)<\/td><\/tr> 5.8<\/td> 8.5<\/td> 91.5<\/td><\/tr> 6.0<\/td> 13.2<\/td> 86.8<\/td><\/tr> 6.2<\/td> 19.2<\/td> 80.8<\/td><\/tr> 6.4<\/td> 27.8<\/td> 72.2<\/td><\/tr> 6.6<\/td> 38.1<\/td> 61.9<\/td><\/tr> 6.8<\/td> 49.7<\/td> 50.3<\/td><\/tr> 7.0<\/td> 61.5<\/td> 38.5<\/td><\/tr> 7.2<\/td> 71.7<\/td> 28.3<\/td><\/tr> 7.4<\/td> 80.2<\/td> 19.8<\/td><\/tr> 7.6<\/td> 86.6<\/td> 13.4<\/td><\/tr> 7.8<\/td> 90.8<\/td> 9.2<\/td><\/tr> 8.0<\/td> 94.0<\/td> 6.0<\/td><\/tr><\/tbody><\/table><\/figure>\n\n\n\n Literature:<\/strong><\/h2>\n\n\n\n