The surfactant Bio-Solv-BBS-3 as a scintillator in liquid scintillation counting

ANALYTICAL

56, 313-323

BIOCHEMISTRY

The Surfactant

Bio-Sob-BBS-3

in Liquid

of Metabolic

Received

mtl

as a Scintillator

Scintillation

S. E. SHARPE Division

(1973)

III

AND

Counting

E. D. BRANSOME,

Endocrine Diseases, Medicnl Augusta, Georgia 30.%X?

January

15, 1973;

accepted

June

College

JR.l of Georgia,

4, 1973

When the surfactant mixture Bio-Solv-BBS-3 is added to a scintillation solvent it acts as a primary scintillator in response to /3 emissions (and Compton electrons from ys). The fluorescence excitation threshold is higher and fluorescence yield is lower than those of the primary scintillators usually employed in scintillation counting. Presence of a surfactant in a sample containing 14C or more energetic /3s will be counted at higher efficiency than would be indicated by a quench correction curve (efficiency vs sample channels ratios or external standard channels ratios) derived from standards not containing surfactant.

Bio-Solv-BBS-3, a proprietary mixture of a nonionic alkylphenylet~her and an anionic surfactant, distributed by Beckman Instruments Inc., has been used to make samples in aqueous solution miscible with organic solvents used in liquid scintillat’ion “cocktails” (1,21. BBS-3 and other quenching but are acsolubilizers contribute to impurity or “chemical” ceptable because with good miscibility, it is possible to count more labelled sample in a homogenous preparation. Figures of merit are thus increased. While investigating the effects of different scintillators and solubilizers on the wavelength of photofluorescence emitted by various scintillation counting cockt.ails, we discovered that the BBS-3 preparation slightly increased the scintillat’ion yield of quenched samples and could itself act as an efficient scintillator. This property of surfactants may be of future value in attempts to develop new sample preparations for liquid scintillat,ion counting of ,8s and y s. Our findings should also serve as an indication that quench correction curves must bc constructed using samples containing similar amounts of surfactant, rather than commercially available sealed standards. MATERIALS

AND

METHODS

Standards of 14C-labelled toluene, cytosine-2- [‘“C 1 dissolved in sterile water, 3H-labelled toluene, and 3H-labclled water were purchased from 1 To whom Copyright All rights

requests

for reprints

should

be addressed.

313 @ 1973 by Academic Press, Inc. of reproduction in any form reserved.

314

SHARPE

AND

BRANSOME

New England Nuclear Corp. and were diluted with different solvents to give calibrated standards of [‘“Cl and [3H]. Calibrated standards of [““S] -Li,SO, dissolved in toluene and in water were donated by the Amersham Searle Co. Other radioisotopes were obtained from Dr. Mark Brown, Radiology Department, Medical College of Georgia,, Augusta, Georgia and Dr. Menard Ihnen, Nuclear Medicine Department of University Hospital, Augusta, Georgia. Samples o’f 0.1 ml or 0.2 ml were pipetted into glass scintillation vials containing 10.0 ‘ml counting solution. We used a Beckman LS 150 liquid scintillation counter (employing pseudologarithmic amplification) with external standard channels ratios (ESR) from a [137Cs] y source. An ESR reading of 0.750 represented unquenched samples; lesser readings reflected various degrees of attenuation of the Compton electron spectrum. The counting efficiency of a sample

Effects

of BBS-3 Counting

TABLE 1 on the Efficiency of [3H] and in Toluene With or Without Additives toluene

Isotope =H,o~

'HTOI

PPOb

[‘%I Liquid Scintillation the Scintillator

to a lo-ml solution BBS-3~

ESRa

Absolute

efficiency

0 + 0 +

0 0 + +

,660 .076 .603

0.04 1.9 2.5 40.5

0 + 0 +

0 0 + +

. 000 ,657 .088 ,618


0 + 0 +

0 0 + +

. 000 ,657 .076 ,583

.09 8.9 22.9 86.0

0 + 0 +

0 0 + +

,000 ,655 ,080 ,590

0.1 89.0 21.8 85.4

,000

Q ESR: channels ratio of the Compton spectrum of an external r3rCs Materials and Methods. b 0.07 g PPO added to each lO.O-ml solution of toluene. c 1.0 ml BBS-3 added to each lO.O-ml solution of toluene. d Original sample of isotope in 0.1 ml Hz0 or 0.2 ml toluene. Subscript denotes whether the sample is water or toluene based.

y source.

(H20

See

or Tol)

SCINTILLATION

FROM

315

SURFACTANT

could therefore be determined by matching the ESR to a quench correction curve of a series of quenched standards as long as the “cocktail” was the same (3). The standard counting solution was 7 g PPO/liter toluene (Fisher Scintaaalyzed) . Other LLcocktails” were 7 G. PPO/liter 10% BBS-3, 90% toluene (v/v) and 100% toluene with no additives. RESULTS

BBS-S: a Primary Scintillator. Aliquots of labelled water were added to scintillation vials containing 10.0 ml of the different solutions. The samples were counted for at least 10 min in a wide channel (o-1000 window). For Table 1 counting efficiencies were derived of a number of ,8 and 7 emitting isotopes in toluene-BBS-3 with their efficiencies in toluene alone or in the presence of PPO, a primary scintillator. Aliquots (0.1 ml) of labelled water were added to scintillation vials containing 10.0 ml of the different solutions. The samples were counted for at least 10 min in a wide channel (O-1000 window). For Table 1,

Effects

TABLE 2 of BBS-3 on the Relative Scintillation Counting and y Sources in 0.1 ml Water, Introduced

Eficiency of Various into Toluene

p

Additives Isotope

PPO”

BBS-3

y0 Relative efficiencya

ESR

1261

0 + 0 +

0 0 + +

,000 ,669 ,076 ,593

0.1 11.2 24.0 100.0

1311

0 + 0 +

0 0 + +

,000 ,669 ,080 ,599

s.5 32.5 86.5 100.0

wo

0 + 0 +

0 0 + +

,000 ,661 ,084 .603

0.2 0.3 0.6 100.0

lQiHg

0 + 0 +

0 0 + +

,000 ,667 ,072 ,599

5.4 22.0 74.5 100.0

0 The most efficient scintillation mixture is regarded as having efficiency. b 0.07 g PPO added to each lO.O-ml vial of toluene. c 1.0 ml BBS-3 added to each lO.O-ml vial of toluene.

100%

relative

counting

316

SHARPE

AND

BRANSOME

counting efficiencies were derived from cpm of accurately calibrated standards. For Table 2, inasmuch as calibrated standards were unavailable, samples having the highest count rate were used as reference standards of 100.0% relative efficiency for the three other samples. Water-based isotopes counted with the organic scintillator PPO required solubilizer; toluene-based isotopes required no such solubilizer (Table 1). Toluene has been noted to be an excellent scintillat,ion solvent but is known to be a poor scintillant (4). In every case, no matter what the nature of the sample, counting efficiency in toluene-BBS-3 was significantly higher than in toluene alone. Figure 1 illustrates t’he reason for the lower efficiencies when BBS-3 was the only scintillator: a downward shift in energy of the photoelectron spectrum. The relationship of ,0 energy to the absolute counting efficiency in a wide channel with and without PPO and BBS-3 is shown in Fig. 2. Since the ESRs of toluene-PPO-BBS-3 solutions were almost equal to ESRs in toluene-PPO solutions (Tables 1 and 2)) improvement of t,he solvent properties of toluene could not account for the differentials in scintillation efficiency when BBS-3 was added. BBS-3 must function as a primary scintillator. The relatively poor performance of toluene-BBS-3 in counting [3H] (Table 1, Fig. 2) suggests that the surfactant has a higher excitation

100

200

WINDOW

300

I 400

(2%)

FIG. 1. Percent of total cpm in 2% channels of a pseudo-logarithmic ‘42 spectrum of O-1000 discriminator units. Each sample of ‘T-toluene contained 10,000 dpm. (Beckman LS-150 scintillation counter.) (A) 7 g PPO/liter toluene : 9006 cpm; (B) 7g PPO/liter toluene 9O%:BBS3 IO%, 884.3 cpm: (C) toluene 9O%:BBS3 10%, 1692 cpm.

SCINTILLATION

FROM

0 100 Me” E MAX

317

SURFACTANT

I OOMev ,9

solutions of toluencFIG. 2. Relationship of detection efficiency to E mBx in various based P-emitting isotopes. (A) “H, (B) ‘“C, (C) 9, (D) %a, (E) “Cl, (F) “P. PPO-BBS : 7 g PPO/liter toluene 90% :BBS-3 10% ; BBS : toluene 90%:BBS-3 10% ; H,O: This latter curve represents the Cerenkov efficiency of these isotopes in a system without anomalous refractive dispersion (5,6).

threshold as well as less overall scintillation efficiency than PPO. The fact that ESR values of toluene-BBS-3 solutions were disproportionately low when compared to counting efficiencies of all the isotopes in Tables 1 and 2 (except [“HI ) is further evidence of a threshold of activation in the lower energy range between [3H] and [l”C]. Figure 3 shows that this dichotomy extends over the entire range of BBS-3 concentration in counting solution. The Compt’on efficiency (and therefore ESR) with a [lsiCs] source is moreover so low that ESRs cannot be used as indices of the efficiency of ,8 detection in BBS-3-toluene, even though the shape of the Compton spectrum is similar to that observed when PPO is added. We have not investigated more energetic y sources for the Compton spectra of external standards. The background count rate of toluene-BBS-3 mixtures was lower (4050 cpm in a maximum O-1000 wide channel) than with the reference counting with toluene-PPO (70-75 cpm). The optimum concentrat,ion of BBS-3 for isotope efficiency, 5-15s (v/v), shown for [‘“Cl in Fig. 3 pertains to all the isotopes we have counted in toluene-BBS-3 systems. BBS-3 with PPO. The implications omfour findings are also important for accurate quench correction of isotopes counted in combined solutions of efficient primary scintillators and surfactanta. Wit.h an isotope below the effective threshold of BBS-3, the addition of surfactant has essentially

318

SHARPE

AND

BRANSOME

..600

-.500

E w .400

.300

..200

,.I00

IO

% BBS-3

20

30

IN TOLUENE

FIG. 3. Effect of BBS-3 on the absolute counting efficiency of “C-toluene, with and without PPO added. Series of duplicate 10 ml “cocktails.” (A) Counting efficiency in PPO 7 g/liter toluene 90%: BBS-3 10%; (B) (‘Ts) ESRs of the samples of series A; (C) counting efficiency in toluene 90% : BBS-3 10% ; (D) (“‘Cs) ESRs of the samples of series C.

the same effect as chloroform [CHCI,], another impurity quencher. In Fig. 4, the effects of both substances on [3H] efficiency and ESR are seen to be quite similar. Plots of both measurements vs % BBS-3 are almost parallel, so that there is no deviation of samples containing BBS-3 on a quench correction curve for PPO-toluene (Fig. 5). The extreme sensitivity of counting efficiency to impurity “chemical” quenching by chloroform is an additional indication that BBS-3 acts as a scintillator with a significantly higher threshold than PPO. With more energetic isotopes however, such “standard” quench correction curves based either on ESRs or channels ratios of sample radioactivity (7) are not valid for samples containing surfactank Figure 4 provides an example: BBS-3 seemsto protect [‘“Cl radioactivity from quenching, so that the “standard” quench correction curve (Fig. 6) cannot be used in samples containing surfact.ant; indeed addition of BBS-3 can result in significantly increased counting efficiency.

SCINTILLATION

FROM

319

SURFACTANT

IOC * 90

B C

I8Oj 701

$

0.600-

60-

_w

E

SO-

w 0.500-

2% w ae 40 30

D

20 +

0.400-

E

‘OL----10

PO

0.300 1 30

IO

20

% BBS-3

30

% BBS-3

FIG. 4. Effect of BBS-3 on impurity quenching by CHCl, of “H- and “C-toluene. Each 10 ml contained 0.07 g PPO with the proportions of toluene and BBS-3 varying as shown. (A) “C-air quenched; (B) “C-, CHCl, 0.1 ml; (C) ‘“C-, CHCl, 0.2 ml; (D) 3H-air quenched; (E) “H-CHCI, 0.05 ml,

I

700

I

,600

I

I

.500

,400

ESR FIG. 5. Effect of varying the concentration of BBS-3 on the quench correction curve of “H in toluene, PPO 7g/L. Air quenched: 0, 0.05 ml CHCla added: 0. The points were obtained from the D and E series of air and CHCh-quenched C3H1 described in Fig. 4.

320

SHARPE

‘“\

,700

AND

BRANSOME

,600

500

300

400

ESR

h’rc. 6. Effect of BBS-3 on a 14C-toluene quench correction curve. The data are derived from Fig. 4, but. are plotted without consideration for the concentration of BBS-3. Samples with increasing BBS-3 concentrations of course had lower ESRs inasmuch as BBS-3 itself contributes to impurity quenching. BBS-3 containing samples (indicated for each condition of impurity quenching by CHCL) fail to conform to the ESR standard curve.

Effect

of BBS-3

TABLE 3 on the Counting Efficiency Various Solvents Absolute

of [3H] and

counting

[i%]

efficiency

in

To

additives Isotope0 ‘HH~o

=TOI

%H,O

WTOI

BBS-3b

CHCL

Toluene PPO 1.5

Benzene .O

-

-

+ +

+

42.0

29.2

.l

+ +

+

45.0

.l ‘2 0 .l

-

-

+ +

+

-

39.1 28.0 2.7

2.2

.l

Toluene .O 2.6

.l .O 2.4

Xylene

Dioxane

.l .l

.l .I .l

.O .O .O

.l

.o

.I

.2

1 :o

2.3

2.1

.l

,l

.l

4.6 .3

87.0 84.4

44.5 4.7

.2 48.5 6.3

.3 45.2 6.3

-

89.0

+

87.5 84.1

2.7 51.4 7.0

4.2 54.3 8.3

8.8 52.2 8.1

.5

2.4

11.2 3.0

a Calibrated isotopes in 0.1 ml Hz0 or 0.1 ml toluene introduced into various b 1.0 ml BBS-3 added to each vial of 10.0 ml counting solution (7 g PPO/liter c 0.1 ml CHClz added to each vial of 10.0 ml counting solution.

Water

.l .l .l 6.8 2.4 1.8

solvents. toluene).

SCIKTILLATIOK

FROM

321

SURFACTANT

Further Investigations. To show that the scintillator propert,ies of BBS-3 were not dependent on toluene, the effects of chemical quenching with different organic solvents are shown in Table 3. There was a similar increase in efficiency in benzene and xylene for each isotope when BBS-3 was added. Figure 7 shows that no matter how energetic the isotope, the concentration dependent fluorescence of BBS-3 will not increase above 20% BBS-3 by volume. Energetic ,8s or ys may however, be counted at high efficiencies when dissolved in 100% BBS-3, significantly above Cerenkov efficiencies (see Fig. 2). The threshold of ,8 detection was more than an order of magnitude below that observed in Cerenkov counting of the same isotopes in water alone, too great a difference to be explained by anomalous refractive dispersion (6). Impurity quenching moreover, is not observed in Cerenkov counting (5). Further evidence of the behavior of BBS-3 as a primary scintillator was provided by the fluorescence excitation experiments represented in Fig. 8. Photon emission by toluene alone was inconsequential. The addition of 10% BBS-3 to toluene resulted in a more than several hundred fold increase in fluorescence yield at wavelengths wit’hin the range of efficient liquid scintillation counter phototube response. This behavior, characteristic of the deactivation of primary scintillators (8), was also ob-

FIG. 7. Effect efficiency

of various

of varying isotopes

the concentration in toluene.

of

BBS-3

on

the

relative

counting

322

SHARPE

300

400

AND

500 EMISSION

BRANSOME

600 WAVELENGTH

700

800

FIG. 8. Fluorescence yield of scintillation solvents excited at 265 nM. Emission spectra were recorded with an Aminco-Bowman Spectrofluorimeter. (A) Toluene 10% BBS3; (B) 79 PPO/liter toluene; (C) 7 g PPO/liter 90% : T BBS-3 10%; (D) Toluene. For ease of comparison the intensity scale for sample D has been multiplied by lo*.

served when BBS-3 was added to standard counting solution. As predicted by Fig. 6, fluorescence yield at 375 nM was increased 27%. The absorption maximum of BBS-3 in toluene (260-280 nM) was consistent with the alkylphenol constituent of the surfactant (9). SUMMARY

AND

CONCLUSIONS

There is compelling evidence that Bio-Solv-BBS-3 (Beckman Inc.) a proprietary surfactant widely used to render labelled aqueous samples miscible with toluene or other aromatic organic scintillation solvents, can itself function as a primary scintillator. Although inquiries to Sent01 Associates Inc. and Beckman Inc. concerning exact composition of this solubilizer have been unsuccessful, we have obtained data (to be reported in the near future) suggesting that other surfactant. mixtures also possess

SCIh-TILLATION

FROM

SCRFACTAKT

323

properties of primary scintillators. Knowledge of the relationship between physicochemical structure and thresholds of fluorescence should lead to development of more efficient solubilizers with low excitation thresholds, and to scintillation “cocktails” with much less liability to impurity quenching than toluene-BBS-3. Although fluorescence by surfactants in response to ,f3 and 7 emissions is as far as we know unreported, Wirth and associates have found that the addition of nlkoxy groups to oligophenylene scintillators increased their eolubilitp in nonpolar solvents without compromising fluorescence yield (10). They ended up with compounds somewhat, similar in structure to surfactant alkylphcnyl ethers. Further definition of the mechanisms and structure-activity relationships of the surfactant fluorescence inducible by radioactivity thus appears to be a realistic goal. R.EFEREKCES 1. Technical Bulletin 012767 Sent01 Assoc. Inc., Littleton, Col. 1967. 2. BRAY, G. A. (1970) in The Current Status of Liquid Scintillation Counting (E. D. Bransome, Jr., ed.), pp. 170-180. Grune and Stratton, New York. 3. CaVaNAuGH, R. (1970) in The Current Status of Liquid Scintillation Counting, pp. 170-180. 4. YANARI, S. S.. BOVEY, F. A.. AND LUMRY, R. (1963) Nature 200, 242. 5. PARKER, R. P., AND ELRICK, R. H. (1970) in Current Status of Liquid Scintillation Counting, pp. 110-122. 6. Ross, H. H. (1970) in Current Status of Liquid Scintillation Counting, pp. 1233126. 7. KOBAYASHI, Y., AND MAUDSLEY. D. V. (1970) in Current Status of Liquid Scintillation Counting, pp. 7685. 8. CRUM, F., COSTA, L. F., MILLER, R. S.. AND PRZYBYLOWICZ, E. P. (1971) in Organic Scintillators and Liquid Scintillation Counting (D. L. Horrocks and C. T. Peng, eds.), pp. 1005-1029. Academic Press New York. 9. GILL~~I, A. E., AND STERN, E. S. (1954) Electronic Absorption Spectroscopy, Arnold, London. 10. WIRTII, H. 0.. HERRMANN. F. V., HERRMANN, G., .~ND KERN, W. (1968) Mol. Cry&

4. 321.