Lithium citrate buffer

Atkin, G. E., and Ferdinand, W., Accelerated amino acid analysis studies on the use of lithium citrate buffers and the effect of //-propanol, in the analysis of physiological fluids and protein hydrolyzates, Anal. Biochem., 38, 313,1970. 

The composition and pH of the buffer should be accurate to 0.001 mol l-1 and 0.01 pH units. Most methods rely on the sequential application of a series of buffer solutions of increasing pH and molarity, with the initial pH around 3.2. Sodium citrate or, preferably, lithium citrate buffers are used, which incorporate a detergent (BRH 35), an antioxidant (thiodiglycol) and a preservative... 

Figure 10.19 Amino acid analyser trace. Separation of a complex physiological standard mixture of amino acids in 3.5 hours using lithium citrate buffers and ninhydrin detection 10 nmol of each amino acid, including the internal standard, nor-leucine, were applied to the column (0.3 X 35 cm) in a total volume of 50 ml. 

A 0.05M NaOAc buffer (pH 6.85)71% MeOH/ 0.8% THE B 0.05M NaOAc buffer (pH 5.35)765% MeOH flow rate 1.6mL7min Lithium citrate buffer... 

Amino acids can be separated without prior derivatization on a cation-exchange resin column. The elution buffers are classically lithium citrate buffers with different pH values and salt concentrations, which are applied stepwise. There is usually a programmed increase in column temperature. Consequently, there are numerous variables affecting the separation of the individual amino acids [6]. For the detection of the amino acids, the column effluent is mixed with the ninhydrin reagent. Nowadays there are only very few manufacturers of AAAs left. The considerable cost of purchase and the operation costs are a potential threat to the widespread application of this technique, although it is still considered to be the definitive method for diagnosing disorders of amino acid metabolism. 

In brief, a 200-pl sample will be deproteinized with 20 pi 5% SSA containing internal standard. Following precipitation and centrifugation, 150 pi supernatant is diluted with 150 pi of the lithium citrate buffer and placed in the autosampler. The derivatization step takes 40 pi of sample and 20 pi of the OPA reagent, which are mixed and left to react for 30 s. A 20-pl aliquot is injected. 

Lithium citrate buffers (Li-A, Li-B, and Li-C) and a lithium cation exchange column (20 X 4.6 cm) were purchased from Beckman. The amino acid standard was prepared from the Beckman standard with the addition of Hyl/flZZo-Hyl (from Sigma). Optimization of the method with respect to temperature and buffer change times was required to adequately resolve Hyl from ammonia. Hyl is partially converted to allo-Hyl during hydrolysis. The peak areas for Hyl and allo-Hyl were combined for standards and samples. 

Filtrate (total amino acids) or supernatant (free amino acids) from above was diluted (20 1) with 0.2 N lithium citrate buffer, pH 2.8, prior to analysis using a Beckman 6380 amino acid analyzer (Beckman Coulter, Inc.) equipped with a 10 cm ion exchange column and lithium citrate buffer supplied by Beckman. Detection was via post column derivatization using ninhydrin. Calibration was via mixed external free amino acid standards. 

The determination of amino acids in physiological samples is one of the most demanding tasks in the field of amino acid analysis. Often, more than 40 components have to be quantified in complex biological matrices such as serum or urine. For such separations, lithium citrate buffers are used as the eluent The separation of asparagine, glutamic acid, and glutamine, important for... 

Figure 5.32 Separation of a standard of physiologically relevant amino acids on a totally sulfonated cation exchanger 0354675T. Column temperature three-step temperature program from 34 to 70 °C eluent 3 lithium citrate buffers pH 2.80, 3.65, and 3.70 flow rate 0.55 mlVmin detection see Figure 5.28 peaks 0.25 pmol/mL each of P-Ser (1), Tau (2), Pea (3), Urea (4), Gaa (5), Asp (6), OH-Pro...

Lithium citrate buffers have been successfully used in automatic amino acid chromatography on resin columns. (See for example. Perry et al., 1968 Peters et al., 1968). 

---