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United States Biological
 
Newsletter
Biological Buffers:
virion visualized by TEMBiological reactions depend on the pH of the solution being kept within a specific range. This is accomplished with buffers which maintain the required pH parameters. Buffers are an everyday but crucial part of your lab supplies. You go through a lot of them and need to be sure that they are top-quality. United States Biological sells a wide variety of premium buffers and buffer components, and since we are using the same prices that have been in effect since 1995 for most of our biochemicals, you can afford to stock up.

 


Choosing the right buffer:

 

Common Buffer Table (at 20°C)
Buffer pKa pH range MW
HEPES 7.55 6.8-8.2 238.3
MOPS 7.28 6.5-7.9 209.3
MES 6.15 5.8-6.5 195.2
PIPES 6.8 6.1-7.5 302.4
TAPS 8.49 7.7-9.1 243.3
Tricine 8.15 7.4-8.8 179.2
Tris 8.2 7.5-9.0 121.1


This month's Special:
10% off selected Good's Buffers

 

H2010 HEPES, M3000 MES, P4200 PIPES

 

HEPES, MES, and PIPES are three of the biological buffers (zwitterionic) designed by Good, et al., and typically referred to as Good’s buffers. They are useful in cell culture media formulations. Selection of biological buffer systems should include the following criteria: exclusion by biological membranes, low absorption between 240 and 700nm, chemical stability, and stable to temperature and concentration changes.

 

MES

MES Structure


Biological Buffers
Good's buffers are good buffers

Good buffers (Good’s buffers) are twelve buffering agents selected and described by Norman E. Good and colleagues in 1966.  Good developed buffers based on several criteria that made them useful candidates for biochemical research. He sought to identify buffering compounds which met several criteria likely to be of value in biological research.

 

1.  pKa.  pKa values in the regions between 6-8 were selected because most biological reactions take place at near-neutral pH.
2.  Solubility.  For ease in handling aqueous biological systems, good solubility in water is required.  Poor solubility in nonpolar solvents (fats, oils, organic solvents) was also required because this would tend to prevent the buffer from accumulating in cell membranes and other nonpolar compartments of the biological system.
3.  Membrane impermeability.  Buffers were considered that did not readily pass through cell membranes, thus reducing the accumulation of the buffer compound within cells.
4.  Minimal salt effects. Highly ionic buffers may cause problems or complications in some biological systems
5.  Minimal temperature and ionic strength effects.  There should be a minimum influence of buffer concentration, temperature, and ionic strength of the medium on the pKa of the buffer.  For example, Tris (a frequently used, but not a Good buffer) has one of the largest pH temperature effects (ΔpKa/°C= -0.031 ) of all biological buffers.
6.  Well-behaved cation interactions.  If the buffers form complexes with cationic ligands, the complexes formed should remain soluble.  Ideally, at least some of the buffering compounds will not form complexes.
7.  Stability.  The buffers should be chemically stable, resisting enzymatic and non-enzymatic degradation.
8.  Optical absorbance.  Buffers should not absorb in the visible or ultraviolet at wavelengths longer than 230nm so as not to interfere with commonly used spectrophotometric assays.
9.  Ease of preparation.  Buffers should be easily prepared and purified from inexpensive materials.

 

The twelve buffers selected by Good are tabulated below.

 

Buffer                           pK at 20°C           DpKa/°C
MES                             6.15                     -0.011
ADA                             6.6                       -0.011
PIPES                          6.8                       -0.0085
ACES                                    6.9                       -0.020
Cholamine chloride         7.1                       -0.027
BES                             7.15                     -0.016
TES                              7.5                       -0.020
HEPES                         7.55                     -0.014
Acetamidoglycine          7.7                          --
Tricine                          8.15                     -0.021
Glycinamide                           8.2                       -0.029
Bicine                                    8.35                     -0.018

 

N. E. Good, G. D. Winget, W. Winter, T. N. Connolly, S. Izawa and R. M. M. Singh (1966). "Hydrogen Ion Buffers for Biological Research".
Biochemistry 5:467-477.

 

We can get a flavor of Norm Good’s own words related to his biological buffers from the Citation Classic interview in Current Contents (1983) 40:23.

 

Norman E. Good
Department of Botany and Plant Pathology
Michigan State University
East Lansing, MI 48824

 

July 5, 1983

 

“The idea of tailoring molecules for specific buffering purposes came from a game I used to play with one of my colleagues, Richard O’Brien, at the University of Western Ontario.  He was interested in the effects of ionization on the activity of nerve poisons and related drugs.  We developed between us a game of guessing pKa’s on the basis of oftentimes rather complex structures of the drugs he was studying.  Meanwhile I had been investigating electron transport and ATP synthesis in chloroplast lamellar preparations, and I had discovered that various anions commonly used in buffers could uncouple electron transport from phosphorylation.  How then to avoid the anion uncoupling and still have essential control of hydrogen ion concentration?  Since the inner salt glycine had absolutely no uncoupling effect, it occurred to me that I might modify glycine in such a manner that it’s new pKa would bring it into an appropriate buffering range (glycine itself buffers at far too high a pH for chloroplast research).  It seemed ovious that replacing the amino group of glycine with tris(hydroxymethyl)-aminomethane (tris) would produce an amino acid with a pKa rather similar to that of Tris itself.  Thus, Tricine was born.

 

Subsequently, it also occurred to me that I could use sulfonic acids instead of carboxylic acids and thereby avoid much of the binding of polyvalent metal ions.  Consequently, when I moved to Michigan State University, I ordered all of the simple primary and secondary amines commercially available and started making the appropriate N-substituted taurines and glycines, drawing on the intuition developed in the above-mentioned pKa guessing game.  Hence the origin of the buffers listed in the paper.

 

Almost all experimental biologists and not a few analytical chemists were interested in the new buffers and consequently the paper received instant attention.  The fact the buffers often proved superior to anything before available made their wide use inevitable.  It is a measure of their acceptance that the biological and chemical journals now allow the use of  the trivial names of several – e.g., Hepes, Tricine – without definition and without reference.

 

In a desultory way, I have continued to make and introduce more buffers along the same lines and I have occasionally enlisted the assistance of real chemists.  A series of N-substituted 3-aminopropanesulfonic acids was prepared using propane sultone and, more recently, W.J. Ferguson and I (1) have introduced a series of N-substituted 3-amino-2hydroxypropanesulfonic acids, all excellent buffers.

 

A summary of available biological buffers was compiled in 1972 by me and my colleague, S. Izawa (2).”

 

1. Ferguson, W.J., Braumschweiger, K.L., Braumscheiger, W.R., Smith, J.R., McCormick, J.J., Wasman, C.C., Jarvis, N.P., Bell, D.H., and Good, N. E. Hydrogen Ion buffers for biological research.  Anal. Biochem. 104:300-310 (1980).

 

2. Good, N.E. and Izawa, S. Hydrogen Ion Buffers, Meth. Enzymology 24: 53-68 (1972).

 

 

 

The picture shows an image created in live coleus leaf by light-induced chloroplast movements. The person appearing in the leaf is Norman E. Good (1917-1992), a photosynthesis researcher and inventor of the "Good" buffers.
From: Discoveries in Photosynthesis, By Govindjee, J.T. Beatty, H. Gest, J.F. Allen, Eds. Springer, 2006 p.703.

Hangarter,R. P. and Ort, D.R. Photosynthesis Research. 34: 245-247, 1992.

 

 

 

 

US Biological Good Buffers

 

B2102                       Bis-Tris
C1076            CAPS, Free Acid
G8225            Glycylglycine (Gly-Gly, Diglycine)
H2030                        HEPES Sodium Salt                                  
H2010                        HEPES Free Acid                           
M3000           MES Free Acid                                
M3005           MES, Sodium Salt                          
M4610           MOPS Free Acid                             
M4615           MOPS Sodium Salt                                    
M4620           MOPSO Free Acid                          
P4200                        PIPES Free Acid                             
P4205                        PIPES, Sodium Salt                                   
T1000                        TAPS Free Acid                              
T1005                        TAPS, Sodium Salt                                     
T1010                        TAPSO Free Acid                           
T1015                        TAPSO, Sodium Salt                                  
T2900                        TES Free Acid                                 
T2905                        TES, Sodium Salt               
T8400                        Tricine                                   
T8600                        Tris Base Ultrapure
T8650                        Tris Hydrochloride (Tris HCl)
T8601                        Tris Base USP
T8600-20      Tris Hydrochloride Buffer (Tris HCl), 1M, pH 7.5
T8600-21      Tris Hydrochloride Buffer (Tris-HCl), 1M, pH 8.0
T8600-05      Tris Acetate  

 

 

 

 

coleus leaf; www.bio.indiana.edu/facultyresearch/faculty/Hangarter.htmlThe picture shows an image created in live coleus leaf by light-induced chloroplast movements. The person appearing in the leaf is Norman E. Good (1917-1992), a photosynthesis researcher and inventor of the "Good" buffers
(From: Discoveries in Photosynthesis, By Govindjee, J.T. Beatty, H. Gest, J.F. Allen, Eds. Springer, 2006 p.703/ Hangarter Lab)

Potassium Acetate
Purity: ≥ 99.0%
Exceeds ACS Specifications

Potassium Phosphate
Dibasic Anhydrous

Purity: ≥ 98.5%
Appearance: White crystalline powder
Solubility: Colorless, clear, complete.

Potassium Phosphate Monobasic Anhydrous
Purity: ≥ 98%
Appearance: White granular powder
Solubility: Colorless, clear, complete.