SynopsisErrors diie to light scat t.eriiig are R feat lire of spectrophotometric met,hods of protein determiriation. The experimental miidit ions may he chosen to allow calculation of the scatter coiitribrition, which is obtained eiserltially from the linear relationship between log turbidity and log incident wavelength, together with knowledge of the slope of such fruictions Using this iriformation, the scatter cwrrections may be obtained by varioiis simple and rapid methods, applicable to single or mult icomponent protein solut,ions.The apparent absorbance of protein so1ut.ions embraces a component due to light, scattering, and wit,h lipoproteins, for example, this scattered component, may greatsly exceed that, of t,he true absorbance. Protein concent,rat,ions may be estimated spectrophotometrically from t'he absorbance at. 280 rim or occasionally 254 nm in flow-cell syst,ems, from the 215/225 nm absorbance difference'z2 or by the Lowry m e t h~d ,~ giving color development estimated at 650 nm. The turbidity contribution with the Lowry method may be reduced by heating or pre-incubat'ing at high pH4J arid with the related but, less specific biuret method by incorporating det,ergent,G but. the errors are not generally severe, as the scatter decreases with increase in wavelength. In the ultraviolet range, the use of a close differ~n c e measurement reduces the scattering contributions, which are most severe when absorbance measurements are restricted to one wave1engt.h.The scat,teririg component in the ultraviolet can be determined from the \vell-knon.n Rayleigh relationship that states that, turbidity or optical density is inversely proport,ional to the wavelength of the incident light raised to a power Ti., this exponent, having a value of between 4 and commonly 2 , dcperidirig on the relshonship between t,he incident, light wavelength and the size, shape and riat,ure of interface of the dissolved macrorn~lecules.~ Thus, det.ermination of t.urbidit,y values at. a series of wavelengths in the visible range allows extrapolation to the ult,raviolet and subtraction of the scnt.t,ercd component, to give t,rue absorbance. This ext,rapolation may be derived from a calculation of the Rayleigh scattering constarit and the wivelength or more empirically from graphical plots of log (t,urbidit,y) versus log (incident. wavelength). lo, l 1 These latter extrapolat.ions are very close to line:wl2 and t,Iw slopes :ire t,he \v:ivelengt,h cxponmts 12448 0
