Caesium-137 labelled algae for filtration studies

International Journal of Applied Radiation and Isotopes, 1960, Vol. 9, pp. 49-53. Pergamon Press Ltd. Printed in Northern Ireland

Caesium- 13 7 Labelled Algae for Filtration Studies K.j. IVES D e p a r t m e n t of Civil a n d M u n i c i p a l E n g i n e e r i n g U n i v e r s i t y College, L o n d o n

(First received 30 October 1959 and in final form 16 February 1960) A m e t h o d is described w h e r e b y the green algae Chlorella a n d Scenedesmus were cultured in a g r o w t h m e d i u m c o n t a i n i n g Gs x37. T h e s e radioactive algae were used as a suspension in w a t e r passing t h r o u g h a c o l u m n of filter sand. T h e distributions of the algal cells r e t a i n e d in the filter were m e a s u r e d w i t h a scintillation counter m o u n t e d externally to the column. Calibrations of the shielded scintillation counter for the a m o u n t of activity p e r algal cell, a n d for the g e o m e t r y of the filter c o l u m n are described. LES A L G A E M A R Q U I S E S D E CI~SIUM-137 P O U R SUR LA FILTRATION

LES I~TUDES

O n ddcrit u n e m d t h o d e p a r laquelle les algae vertes Chlorella et Scenedesmus furent cultivdes d a m u n milieu c o n t e n a n t d u Cs 137. Ces algae radio-actives f u r e n t employdes e n suspension d a m l ' e a u q u i passait ~ travers u n e colonne de sable de filtre. L a mesure des distributions des cellules algales retenues dans le filtre se fit a u m o y e n d ' u n c o m p t e u r ~t scintillement m o n t 6 e x t ~ r i e u r e m e n t ~t la colonne. O n d~crit l'6talonnage d u c o m p t e u r ~ scintillement abritd p o u r la q u a n t i t d d'activit6 p a r cellule algale, et p o u r la gdom6trie de la colonne-filtre. IIPHMEHEHHE B O ~ O P O C S I E F I , M E q E H B I X Cs 137, B H C C J I E ~ O B A H H X IIO tDH3IBTPAIAHH Onl4caH MeTO~, B HOTOpOM aeaeHble Bo~opoc:II4 Chlorella ~I Scenedesmus I~y,'IbTl4BI4pOBaJmcb Ha cpe~e, coAepma~e~t CstaL OTH pa~lnOaKTHHHble BoAop0cJ~H B BO~HO~ cycI]eH3HI4 lxponycgaal4cl~ qepe3 KO~OHny, Hal]o~Helmym IleCKOM. Pacnpe~eaeHne R~IeTOK B0~0pocaefi, OCTaIOIIDfXC~I Ha qbl4~l~Tpe, 143Mep~l~I¢ CI~I,IHTI4JI$IJ:II~I4OHHIaIMcqeTqi4HOM, pacttoJIoH~eHHI,IM BHe HOaOl4ttl4. ~)HpaHI4pOBaHHblit CI][HHTHYIJIFIIii4OHHbII';~CqeTqHH 61aIJI r p a ~ y ~ p o n a n B eAl4nm~ax aUTii4BItOCVI4 Ha I~JTeTRy Boaopocan. OnncaHa reoMeTpn~ KOaOI4Rti JI,~1~ qbm'abTpatml4. GsxaT-MARKIERTE ALGEN FUR FILTERUNTERSUCHUNGEN Eine M e t h o d e ftir K u l t u r e n yon grtinen Algen Chlorclla u n d Scenedesmus in Cs ~a7 enthalt e n d e m M e d i u m wird beschrlcben. Diese radioaktiven m a r k i e r t e n Algcn, suspendiert i n Wasser, w u r d e n d u r c h eine Sands~iule filtriert. Die V e r t c i l u n g der Algenzellen die i m Filter zuriickblieben, w u r d e mittcls eines Szintillationsz~ihlers ausscn a n dcr Siiule gemessen. Die K a l i b k r i e r u n g des aggeschirmtcn Szintillationsz~ihlcrs beziiglich der M c n g c d e r Aktivit~it p r o Algenzelle u n d d e r G e o m e t r i c d e r Filters~iule w i r d bcschrieben.

INTRODUCTION The particles had to be of a uniform size, discrete, of definable shape, denser than the fluid, strong enough to resist the fluid shear and one or two orders of magnitude smaller than the grains of the filter medium. For this purpose two species of green algae were

T o investigate experimentally the laws governing the filtration of particles suspended in a fluid passing through a porous granular medium, a method was required to determine the distribution of the arrested particles in the column. 49

50

K. J. Ires

chosen~ Chlorella and Scenedesmus, suspended in water filtering through sand columns with a grain size of approximately 0.5 mm. It was decided to measure the distributions of the algae in the sand column by labelling them with a radioisotope and detecting them by an instrument mounted externally to the column. Therefore an isotope was required which: (1) would be taken up by the algal cells during growth, (2) would not be returned by the algae to solution in non-radioactive water,

(3) could be detected through the sand, water and glass walls of the filter column. As culture of the algae would take a few weeks, and the harvesting, washing and experimentation an extra few days, a half-life of less than 100 days would have been undesirable. After consideration of several alternative isotopes caesium-137 was chosen. It also had the advantage of being immediately available where the experiments were conducted.

ALGAL CULTURE AND HARVESTING Zinc sulphate 0.4 mg ] The principal algal culture was grown in a Manganese sulphate 0.3 mg L Micrometabolic 101. (nominal capacity) aspirator bottle, Boric acid 0.3 mg [ constituents illuminated by circular fluorescent lamps. Cupric chloride 0.003 mg J Subsidiary cultures were grown in 500 ml measuring cylinders arranged round the 10 1. Caesium-137 was added as carrier-free bottle. Air containing 5-10 per cent CO~ caesium chloride, at a concentration of 100 was continuously bubbled through all the /~c/1. and sodium hydroxide was added to cultures. This was sufficient to keep all the bring the pH between 5.0 and 6.0. It generally took cultures 15-20 days to algae in the measuring cylinders in suspension, but a magnetic stirrer had to be become a dark opaque green suspension approximately 2 × 108 algal cells/ml. provided to agitate the 10 1. culture. The culture medium had to support the Usually a 20-30-day culture was used, and autotrophic growth of the algae and suppress the original culture was centrifuged to heterotrophic organisms such as moulds, separate the algae. These centrifuged algae bacteria and protozoa. It has been shown by were washed in distilled water, recentrifuged WmmAMS and SWANSON~1) that algae take up and then the process repeated. Finally the caesium and potassium in a similar manner, algae were dispersed in a tank of tap water and in order to have a significant amount of supplying the filter. To test that the Cs 137 taken up by the caesium taken up by the algae the potassium in the growth medium must be reduced to a algae during growth was not passed back minimum. After Several trials of different into the water (which was an indispensable media the following was found to be satis- requirement of the experiment) such a triple-washed suspension was shaken in factory: distilled water in a flask for a week. The Per litre of solution algae were then separated from the water Sodium nitrate 2-5 g with a millipore filter. The algae on the Magnesium sulphate 4.9 g millipore membrane exhibited considerable Potassium dihydrogen radioactivity, whereas the water had no phosphate 0.25 g Ferric citrate 3.0 mg detectable activity. EXPERIMENTAL APPARATUS

The filter consisted of a glass column containing sand, with connexions to the column for maintaining constant water flow

rate, and for measuring the pressure at various points in the filter. The radioactivity distributions were

Caesium- 137 labelled algae for filtration studies

measured with a lead shielded scintillation counter coupled to a timing and scaling unit. The lead shield had a horizontal slot 1 cm × 7 cm through to admit radiation from the filter to the scintillation crystal. The

51

shielded scintillation detector and photomultiplier was mounted on a vertical rack and pinion to enable it to be set against any level of the filter. The complete assembly is shown in Fig. 1.

M E A S U R E M E N T OF RADIOACTIVITY

To determine the distribution of the algal cells in the depth of the filter column two calibrations were required. The first of these was to determine to what extent the count rate recorded by the scintillation counter was affected by radioactive material not exactly opposite the collimating slit i n the lead shield. This characteristic of the shield and source geometry was determined by use of a plane source of Cs13~ (CsCl solution as a thin layer in a glass container fitting inside the filter column) moving this source relative to the scintillation counter and recording the count rate. The resulting "smear" of count rate, expressed as a percentage of the m a x i m u m count rate (immediately opposite the slot) is shown on Fig. 2. The second calibration was to determine what count rate was recorded from a known number of algal cells in the filter pores. This was achieved by taking a radioactive algal suspension in water, which had been counted in a haemocytometer, placing it in the empty filter column and then adding sand and vibrating to remove air and to consolidate the sand column. Algal cells were then trapped uniformly in the sand pores. Care was taken to leave no column of algal suspension above the sand, which might have settled on the sand surface giving a locally higher concentration of activity. By locating the counter at mid-depth, recording the count rate and correcting it for the "smear" characteristic the count rate for a known cell concentration in the pores was determined. This had to be done for each species of algae as the mean amount of activity per algal cell was different. It was assumed that the radioactivity was distributed statistically uniformly on all the algal

cells, as enumerated in the haemocytometer. This microscope count also enabled an examination to be made as to whether the culture contained m a n y dead cells or was significantly contaminated with fungi or 36 32. 28 24 20

16. 12 8 4

~o

I0 2O 5O 4O

4

~,.o....,~50-

60

70

80

90

I00

12 16 2O 24 28 32

4o', 44 FIC. 2. Smear from plane source of Cs T M scintillation counter with slotted lead shield.

bacteria, which might have taken up a significant fraction o f the Cs 187. Using the largely inorganic medium already described no significant contamination was seen and very few dead (colourless) algal cells detected. This was not true for the medium suggested by WILLrAMS and S w ~ s o N ~1) which supported dense bacterial populations, hence its use was abandoned.

K. J. h,es

52

COUNT RATE DISTRIBUTIONS 2O F D u r i n g a filtration experiment c o u n t rates were recorded at suitable depths of the filter c o l u m n every 30 min. D u e to the nature of the filtration process it was necessary to take count rates at every centimetre depth for the first 5 cm, then at 7 cm, 10 cm and below that at 5 cm intervals. Such a count rate distribution is shown as curve 1 on Fig. 3. This curve had to be corrected for the activity of algae collecting z" & Oar on the surface of the sand column, giving a corrected curve 3. This curve was the result 0 2 4 6 8 IO 12 14 16 8 20 22 24 ~ 28 30 of some true distribution, curve 4, being Depth, crn "smeared" by the characteristic (given on Fro. 3. C o u n t rate distribution curves. Fig. 2) of the counter assembly. There is no k n o w n analytical method of taking the smeared distribution curve 3 and applying obtained which produced the effect of curve the counter characteristic, Fig. 2, t o it to 5, almost identical with the corrected derive the true curve. T h e m e t h o d used by experimental curve 3. This indicated that the author was to assume a trial distribution curve 4 was the true distribution o f c o u n t and "smear" it with the counter character- rate through the filter depth. From a istics to see if the resulting distribution previous calibration the c o u n t rates o f curve matched curve 3. Successive approximation 4 were translated to algal cell concentrations enabled the true distribution to be found, in the filter pores. This procedure had to be but the numerical c o m p u t a t i o n of such a repeated for every" distribution recorded as method is extremely tedious. A sample the filter run proceeded. A m o r e effective analysis is shown in Table 1. shield to the counter with better collimation From a numerical analysis the curve 4 was w o u l d have reduced this burden of analysis.

TABLI~ 1. T a b u l a r m e t h o d of determining "smeared" curve of count rate, ti'om trial "true" distribution and k n o w n counter "smear" characteristics. ( A p p r o x i m a t e l y l / 7 t h of the full table shown). Depth (cm) I 2 3 4 5 6 7 8 9 10 11 12 13

0.21

20 15 11 8 6 5 4 3

/

0.28

35 27 20 15 11 8 6 5 / ..14~5

0.38

0-53

0.76

1.0

0.76

400 304 300 228 119 225 171 64 90 169 128 48 67 127 96 36 50 95 72 27 38 71 54/38 20 28 53 ~40 15 21 t 4 0 "I" 30 I1 16/23 I" 30 23 9 / 1 2 - - 1 7 23 6 ~ 9 13 17 7 10 13

Observed ("smeared") value is diagonal sum (e.g. 9 c m value),

228 171 128 96 72 54 40 30

"True" distribution.

0.53

0.38

212 159 119 90 67 50/36

152 112 84 114 84 63 85 63 47 64 47~.35/ 48/35 27 27 2O 27 20 15 20 15 11 15 11 8 11 8 6 9 6 5 6 5 4 5 4 3

28 21 16 12 9 7

0.28

0.21

Countel ,,smear,~

A n y value is product of "smear" value and "true" value e.g. 23 x 0-76 =~ 17.

Caesium-137 labelledalgaefor titration studies

53

STUDY OF FILTRATION With the algal deposit concentrations measured for known depth and time coordinates the mathematical and hydrodynamic laws of filtration were studied. Previously only one complete statement of ttie space-time distribution of deposited material in a homogeneous filter had been made. ~2~ This has been found to be inadequate in various respects and an alternative formulation has been proposed by Iv~s ~3~. This is ~ a 9-

Ot -- v ~ + ca

f-a

) C

(1)

where O" =

storage ratio =

volume of deposit unit volume of filter

time approach velocity of water to filter surface ;l, c, ~ = rate factor parameters, characteristic of the suspension, state of flow and filter grains f = filter initial porosity C = volume concentration of suspension in the flow. The volume concentration of suspension in the flow and the storage ratio are linked by the continuity equation t=

0 =

Oa o-7 +

OC = o

(2)

where x is the depth in the filter measured from the filter surface.

The algal deposit concentrations measured by the technique described enable graphs of a against t to be plotted for each interval of depth. Hence Oa/Ot is the slope of such a graph. Putting y=~ then from (1)

+ca

Y-

f--a

(3)

aa/at vC"

For the top layer of the filter C = Co, the inflow concentration, so y can be evaluated for various values of a from the a against t graph. A plot o f y against a will enable the constants 4, c and ff to be evaluated from equation (3). With these specified, it is possible to derive the deposit in the filter, and the concentration in the flow through the filter for any d e p t h and for any time, from equations (1) and (2). In fact no analytical solution is known for these equations, so they have been solved numerically by finite difference methods on an • electronic digital computer. Details of this, and the analysis of pressure loss through the filter layers is contained in the author's paper to the Institution of Civil Engineers.t3~ Acknowledgements--The work described in this paper was carried out in the Department of Sanitary Engineering, Harvard University, U.S.A., under the auspices of a Robert Blair Fellowship.

REFERENCES | . WILLIAMS L. G. a n d

SWANSOn H . D. Science 27,

(3291), 187 (1958). 2. MINTS D. M. Dokl. Akad. Nauk SSSR 78, 315 0951).

3. IvEs K. J . Rational Design of Filters.

Proc. Inst. Cir. Engrs. 16, 189 (1960).

(Abstract).