Flame Photometric Microdetermination of Boron in O r g a nobolron Cornpou nds TAMOTSU YOSHlZAKl Shionogi Research laboratory, Shionogi & Co., lid., Fukushima-ku, Osaka, Japan
b The flame spectrophotometric method was applied for determination of boron in organoborm compounds to eliminate the ambiguities which are inevitable in determination by other methods. During this investigation, it was observed that boron emission depends upon not only boron content but also the molecular structure or composition of the s'ample solution. This necessitated the complete decomposition of various boron compounds to boric acid-i.e., the standard substance. The flame spectrophotometric method i s characterized b y having no ambiguities in deyerminationsand i s composed of three steps-decomposition of weighed samples, preparations of standard-added solution and dilute sample solution, and measurement with a flame spectrophotometer. A micro amount of sample (0.5 to 1 mg. as boron) i s sufficient for analysis, while the relative error i s within 1% of the theory.
NI
analytical methods have becn reported for determining boron in organoboron compounds by :t technique in which t'oron is oxidized to horic acid, converted into mannitoboric acid, and tit,rated as u-ual. But there remain problems t,o be solvedfor example, danger thst a part of the sample may be lost by incomplete digeation or volatilization and trouhleame titration in the presence of amine, formed during the wet oxidatic'n of compounds containing nitrogen. The purpose of this investigat,ioniq t o i:xtend the flame photometric method tc the determination of boron in organoborori compounds containing nitrogen and t o standardize :t univcrval technique which is independent of nitrogen content and involves no ambiguities such as starting :ind end points of titratilin. The concentration-! ntensity (C-I) curve for boron in boi,ic acid was described as strictly linear up to a t least, 300 p.p.m. of boron ( S ) , and the present, author confirmed the linearity up to 500 p . p m a t 519.5 mp. When the C-I curve for boric acid wac used to convert the flame spectrometer readings into boron coneenbrations in sample solution,s or into boron contents in samples, boron values ivere too high :'or most boronnitrogen compounds, w e n when analyzed after complete digestion. This
was attributed t o the effect of amines formed from nitrogen in the compounds, as similar effects of enhancement due t o inorganic ions have been reported by Dean and Thompson ( 3 ) . The mode of interaction between boron emission and amine content was examined with aqueous solutions of boric acid and aniline hydrochloride. In this case, the emission increased continuously but not proportionally with the increment of amine concentration (Figure l), whereas the C-I curve for a fixed concentration of amine was strikingly linear up to 500 p.p.m. of boron. If this relation between boron emiasion and amine concentration holds for decomposition products of boronnitrogen compounds, it apparently indicates that the method of sbandard addition g i x s good results. This was
the case for some borazine derivatives but not for others, although discrepancies were decreased by adoption of the method of standard addition. Table I presents the results on a number of compounds. -4probable explanation for these discrepancies is that boron emission depends upon the species of atoms or groups attached to the boron atom in the sample solution. If this is true, the use of an oxidizing reagent such as nitric acid may lead to a better agreement, because boron in a sample is gradually converted to boric acid, which is identical to the standard subbtance. In the experiments with oxidizing acid, a set of values was given as the function of oxidation time for each sample examined. Satisfactory results were obtained after a period of oxidation, the length of ahich depended upon the specie. of sample. The procedure n as then rei ised to meet these requirements-i e., complete digestion. I n preparing the sample solutions, a constant amount of methanol was added to inereace the intensity of the boron emisaion (3); methanol is very suitable for this purpose because it i+highly miscible with nater. Buell ( 2 ) has recommended cleaner's naphtha and 4-methyl-2-pentanone for this purpoqe, but as the*e are inGoluble in water they are uniuited for the present inveatigation. Methanol seems to play an important role al-o in a\ oiding the clepoaition of iolid phase during or after the oxidation by disolving the oxidation products.
ANY
Aniline hydrochloride.
mg.1 l i t e r
Figure 1. Effect of aniline hydrochloride on boron emission
EXPERIMENTAL
Apparatus and Instrument Settings. Boron emission was measured with a Type EPU-SA spectrophotometer
In boric acid solutions containing 300 p.p.m. of boron
Table 1.
Results b y Method of Standard Addition for Boron
s o . of
Compound Phenylboronic anhydride
results
Boron found, 70 Max. Nin. Av.
Boron theory, C"
/O
Av.theory,
5
5
10.89
10.31
10.64
10.41
0.23
borazine
2
21.02
21.02
21.02
19.70
1.32
borazine
9
10.23
9.67
9.91
9.25
0,6(i
borazine
10
S.32
8.00
8.15
8.26
borazine
3
7.10
6.95
7.03
6.80
B-Trimethyl-LV-trimethyl-
B-Trimethyl-S-triphenylH-Triethyl-A'-triphenylB-Tributyl-A--triphenyl-
VOL. 35, NO. 13, DECEMBER 1963
-0.11
0.23
2177
I
t
c L
0
m +
8.0
-
c L
O
n
4
7.0-
6.0L 0
"
"
I
'
"
1 2 3 DeCOmpOSitiOn time.
Figure 2.
4 hr.
Boron emission of methyl-N-triphenylborazine as tion of decomposition time
" 5
B-trifunc-
Upper and lower dotted lines represent tentatively permitted limiis of error (&2% of theory)
with a Type H-2 flame attachment (Hitachi, Ltd., Tokyo), employing oxygen-hydrogen fuel. The spectrophotometer was adjusted for 300 p.p.m. of boron solution to give a net reading of 80 to 90 in full scale of 110 a t 519.5 mp. Flame height, mirror position, and fuel pressure were adjusted in the usual way (1, 6) to obtain maximum emission.
Recommended Procedure. -4sample containing 0.5 to 1 mg. of boron is weighed into a 4-ml. ampoule, and 0.5 ml. of acetone is added to dissolve the sample. Although the decomposition rate is expected t o increase with the solubility in acetone, it is not necessary to dissolve the sample completely. For the sample which is soluble in 17N HYOa aqueous solution, the addition of acetone is not necessary. After 1 ml. of 17iV HNOI aqueous solution is added, the ampoule is sealed and kept in a water bath a t 80"C. for a time estimated to be sufficient for complete decomposition, by analogy or bs. appropriate preliminary work. After cooling, the ampoule is opened and the content is made up to 2.5 ml. with methanol to give solution I. Two solutions are then prepared by mixing 2 ml. of methanol and 1 ml. of I in each of two 5 m l . measuring flasks. To one flask, 2 ml. of standard solution of boric acid containing 500 p.p.m. of boron and pure water are added to make 5 ml. of solution 11. To the other flask is added pure water to make 5 ml. of solution 111. A blank solution (IV) is prepared in the same manner as the sample solution. The emissions of solutions 11, 111, and I V are measured a t 519.5 mp on a spectrophotometer to give the readings of Isa, Is, and Io, respectively, in the following equation, which gives the boron concentration, C, in sample solution 111.
c = 500 x
2
x Isa
- Is
(p.p.m.1
Slit, mm.
0.035 t o 0.045 1.25 Oxygen, kg./sq. cm. Hydrogen, kg./sq. cm. 0.20 Sample. Samples used in this investigation were synthesized in our laboratory and shown to be of good purity by their physical properties as well as by the determination of carbon, hydrogen, and nitrogen.
where 2 is the dilution factor-Le., in this case.
2/5
RESULTS
Table I1 lists the results of determinations of boron in a number of organo boron compounds by this procedure.
II. Results for Boron b y Recommended Method Recovery Oxidation Boron, % Found - % of time (80' C.), Found Theory theory theory hours Compound 10.45 Phenylboronic anhydride 0.04 100.4 6 Table
10.21 10.41 -0.20 98.1 6.08 99.0 -0.06 6.27 6.14 0.13 102.1 Triethylaminoboron trichloride 4.85 -0.10 98.0 4.83 4.95 -0.12 97.6 19.45 -0.25 98.7 B-Trimethyl-i2'-trimethylborazine 20.06 19.70 0 . 3 6 101.8 9.30 B-Trimethyl-X-triphenylborazine 0.05 100.5 9.34 9.25 0.09 101.0 B-Triethyl-A/-triphenylborazine -0.21 8.05 97.5 8.16 8.26 -0.10 98.8 6.40 B-Triethyl-N-tris(p-chloropheny1)-0.14 97.9 6.54 6.54 borazine 0.00 100.0 B-Tributyl-N-triphenylborazine 6.89 0.09 101.3 6.77 6.80 -0.03 99.6 6.21 B-Triphenyl-A'-triphenylborazine 0.17 102.86.07 6.04 0.03 100.5B-Triphenyl-A'-tris(p-N,N-dimethyl- 4.78 -0.09 98.2 aminopheny1)borazine 4.78 4.87 -0.09 98.2 B-Tris(p-N,N-dimethylaminopheny1)- 4 . 7 5 -0.12 97.5 12'-triphenylborazine 4.75 4.87 -0.12 97.5
Trimethylaminoborontrichloride
2178
ANALYTICAL CHEMISTRY
5 2
3
2 3 2 2 2 2 2 2 4.5 4.5 2 2
3 s'O 0.0
i
: I 4
c
;7 0 c
4
5.0
4.01
2
3
4
Decomposition time.
Figure 3.
5 hr.
Boron emission of
B-tri-
methyl-N-tris(p-N,N-dimethy laminophenyl) borazine as function of decomposition time Upper and lower dotted lines represent fentatively permitted limits of error ( & 2 % of theory)
DISCUSSION
Two examples of the dependence of emission on decomposition time are given in Figures 2 and 3, where apparent boron contents in the solid samples are plotted against decomposition time. In this experiment, nitric acid was employed as the oxidizing reagent, and the digestion was carried out in a water bath kept a t 80" A 3" C. Satisfactory results were obtained after a period of oxidation, the length of which depended upon the species of the sample. These preliminary observations led to the above-mentioned recommended procedure. Nost of the samples involved in this investigation could be satisfactorily analyzed by this procedure, but occasionally slight modifications were heeded, as described below. When a sample is difficultly soluble in both acetone-acid mixture and 17N HNOa a t 80" C., it is heated with nitric acid (without acetone) in a sealed ampoule to an elevated temperature-e.g., 130" C.-until all the solid disappears. Then the ampoule is transferred to the water bath a t 80" C. to assure complete decomposition. In a few cases, a rapid depression of emission intensity was observed during the period of measurement, because of vaporization of organic solvent, but this trouble is removed by the use of triethylene glycol dimethyl ether or preferably pure water in place of volatile solvent-Le., methanol or acetonein preparation of the sample solutions. The use of hard glass ampoules is preferable, to avoid an explosion due to the elevated pressure in the ampoule during or after the decomposition. Care must be taken to minimize the loss of solution attached to the cutoff tip of the ampoule on opening. I t is
desirable to duplicate thi: determination a t different periods of decomposition time to ensure complete decomposition. Advantages of Method. -4 small amount of sample containing 0.5 to 1 mg. of boron suffices for analysisless than in the titrimetric method. The ambiguities in the determination, such as the choice of pH range in the titrimetric method, have been eliminated by converting boron into boric acid, which is identical to the standard substance, so that no calibration is needed against any interfering effect. In the gravimetric d1:termination of carbon and nitrogen in organoboron compounds low values occasionally were obtained (4, 6), probably because of the formation of boron carbide, boron nitride, or both, in the course of combustion. If boron is determined simultaneously, the value m u d also be low, because of incomplete rligestion. The
present method of analysis does not have such handicaps. While slight modifications are needed in the procedure for preparing sample solutions for a few compounds, the general principle that boron is converted to boric acid is not altered, and naturally the method is applicable to compounds which do not contain nitrogen. The analysis is somewhat time-consuming, because of the comparatively long period for decomposing treatment, but that time can be utilized for other work. The time of manual operation is less than 1 hour for one run. To exhibit the accuracy and precision of the method, recoveries (per cent of required) are listed in Table 11. The mean error is 0.6 (per cent of required) and the relative standard deviation is 0.6i% with 22 results in Table 11. Thus the method gives satisfactory values for ordinary demands. If a larger amount of sample is available, a higher
precision can be obtained by using two or more additions of the standard boron solution. ACKNOWLEDGMENT
The author expresses appreciation to Haruyuki Watanabe, Toshio Nakagawa, and Ken’ichi Takeda for their helpful advice and encouragement. LITERATURE CITED (1) Buell, B. E., ANAL.CHEM.30, 1514 11968). ( 2 ) Ibzd., 34, 635 (1962). (3) Dean, J. .4., Thompson, C., Ibid., 27, 42 (1955). ( 4 ) Gerrard, W., Hudson, H. R., Mooney, E. F., J . Chem. SOC.1962,113.
( 5 ) Gilbert, P. T., Jr., Hawes, R. C., Beckman, A. O., ANAL.CHEM.2 2 , 772 (1950). (6) Watanabe, H., Totitni, T., Nagasai+:i,
K., Yoshizaki, T., Shionogi Research Laboratory, Shionogi & Co., L t d , Osaka, Japan, unpublished data, 1962. RECEIVEDfor review April 9, 1963. Accepted September 9, 1963.
Spectrophotometric Determination of Nitrogen in Total Micro-Kjeldahl Digests Application of Phenol-Hypochlorite Reaction to Microgram Amounts of Ammonia in Total Digest of Biological Material LEWIS T. MANN, JR. laboratory o f Chemical Pathology, Department o f Pathology, Harvard Medical School, Boston 7 5, Mass.
b Modifications of putttished methods have permitted the direct estimation of ammonia in total micro-Kjeldahl digests by means of formation of “phenol-indophenol” from ammonia and phenol in the presence of base, sodium hypochlorite, and sodium ferrinitrosopentacyanide (“nitroprusside”). In the reported procediire, 1 to 15 pg. of nitrogen (as ammonia) can b e determined. The absorptivity of the colored solutions ranges from 1.1 to 1.4 X l o 3 cm.-l./gram ( E = ca. 15-20 X lo3)a t 630 mp, Absorptivity is constant with NH, concentration from 0.1 pg./mI. to 1.5 pg./mI. Once formed, colored solutions in this range obey the Beela-Lambert law, and those solutions, too dark for direct spectrophotometric obiiervation, may b e diluted to 2 to 5 volumes with an appropriate buffer without loss of linearity.
A
there are reports in the literature of the titrimetric or colorimetric determination of nitrogen as ammonia in total micro-Kjeldahl LTHOUGH
digests (3, 8, 9), each method has presented some particular difficulty when we have attempted to apply it to biological materials. The problems are principally those of available sample size and a wide variability of nitrogen content. In addition, we required a rapid method (24 hours or less) which did not involve the use of ultramicro equipment and techniques. Several recent reports (2, 4-6, 11, 14, i7,19)have been published in which the formation of “phenol-indophenol” from phenol and ammonia under basic oxidizing conditions has been used to determine ammoniacal nitrogen. The test is extraordinarily sensitive, so that under optimal conditions, amounts of nitrogen (as ammonia) in the microgram range should be determinable. Lubochinsky and Zalta (11) described uniquely a system in which the total Kjeldahl digests could be used in this colorimetric procedure. Because we experienced difficulty in repeating this procedure on our Kjeldahl digests, we undertook a more detailed investigation of the color reaction, under the conditions with which we had to contend,
namely, the use of a mercuric ion catalyzed Kjeldahl digest, which was adapted from Ogg (IS) and Steyermark (18). EXPERIMENTAL
Apparatus. A square aluminum block, 8 inches on a side, about 2.5 inches thick, was drilled with 36 evenly spaced holes 3//* inch in diameter, and 2 inches deep. To the underside of the block was fastened a 1000watt heating element (Calrod) which mas controlled by a variable transformer. Heat loss was cut by surrounding the block with glass wool. =i flat pan, filled with washed sand, placed atop a hot plate can also be used, but with less convenience. Reagents. All water was either distilled or passed over a sulfonic acid ion exchange column. Zinc dust, low X, finely divided, impalpable powder. Coarser grades (60- to 200-mesh) were not satisfactory. Sodium Phenolate Reagent. Phenol was distilled from aluminum turnings ( 7 ) and stored (tightly stoppered) in a refrigerator. Phenol (6.0 grams, 0.059 mole), was placed in a 100-ml. voluVOL. 35, NO. 13, DECEMBER 1963
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