31.2.04 AOAC Of fi cial Method 968.10 Spi ral Ves sel Count of Ca ...

31.2.04
A. Ap pa ra tus
AOAC Of fi cial Method 968.10
Spi ral Ves sel Count
of Ca cao Prod ucts
First Ac tion 1968
Fi nal Ac tion 1974
Co dex-Adopted–AOAC Method*
(a) Sieve.—No. 230, 5 in. (13 cm) di am e ter, stain less steel.
(b) Grinding equip ment.—(1) Coarse grind ing (cut ting
ac tion).—Hobart Food Cut ter No. 84181-D (Hobart Man u fac turing
Co., 711 Penn syl va nia Ave, Troy, OH 45374, USA), or equiv a lent.
(2) Fine grind ing.—13 cm (5 in.) glass mor tar and pes tle or Tor sion
Bal ance Co. elec tric mor tar grind ers MG1 or MG2. Ad just MG2 so
that pes tle and shaft are not un der ten sion by loos en ing top knob and
lock nut by 3 turns and ad just clos ing spring con trol to 1 2 ten sion.
(c) Alu mi num dish.—Di am e ter ca 77 mm, height ca 33 mm; with
cover.
(d) Brush.—No. 10, ny lon, rub ber set, oval sash paint brush with
bris tles cut to 4–4.5 cm (avail able from Sherwin-Williams Co.,
Pros pect Ave, Cleve land, OH 44101, USA, or dis trib u tors).
B. Prep a ra tion of Test Sam ple
(a) Choc o late li quor, choc o late.—Pre pare as in 970.20(b) (see
31.1.01).
(b) Expeller cake.—Crush with mor tar and pes tle and grind to
pass No. 30 sieve in mill, (b)(1), ca 1 2 tea spoon ful at time. Mix well
and store in tightly stoppered jar.
(c) Co coa press cake.—Pre pare and store as in (b). (Many test
sam ples can be eas ily pul ver ized af ter dry ing 2–3 h at 60�–70°C.)
(d) Co coa.—Use as is. Store as in (b).
C. Defatting and Grinding
Set up in hood No. 230, 5 in. (12.7 cm) sieve in 15 cm (6 in.) glass
fun nel with tip dip ping ca 2 cm into 500 mL flat-bottom Py rex
cen tri fuge bot tle.
Place 15 g co coa, coarsely ground (30–40 mesh) co coa press
cake, or expeller cake, or 25–30 g choc o late or choc o late li quor in
250 mL cen tri fuge bot tle. Add 100 mL ether, stop per, shake
thor oughly to dis solve fat, and pour onto sieve. Wash ma te rial on
sieve well with ether. Wash lower rim and both sides of sieve and
in side of fun nel with ether. Let ma te rial on sieve stand un til dry (ca
15 min).
Cen tri fuge mix ture in 500 mL cen tri fuge bot tle 10 min at
2000 rpm. De cant and dis card supernate. Re place cen tri fuge bot tle
un der fun nel.
Place sieve with dried co coa ma te rial in re ceiver (sieve bot tom
pan). Brush ma te rial through sieve with No. 10 sash paint brush.
Trans fer re tains, us ing brush, to 12.7 cm glass mor tar and grind ca
45 s with glass pes tle, or grind 2 min in mo tor-driven mor tar grinder.
Trans fer to sieve and rebrush. Re peat grind ings and brushings un til
vir tu ally all ma te rial passes through sieve. Quan ti ta tively trans fer
ma te rial, in clud ing small amount on sieve (83.4)
re peat de ter mi na tion through out, but weigh 0.200 g test portion and
� 2006 AOAC IN TER NA TIONAL

Table 968.10. Stan dard spi ral ves sel count values
Shell in choc o late
com po nent, %
di lute to 100 mL with H 2O in 100 mL glass-stoppered grad u ate
con tain ing 50 mL glyc erol. Count at 200�. Use col umn for counts
listed un der “>15% shell” for pre par ing stan dard curve.
F. Counting In struc tions
Spi ral ves sels vary greatly in size. No dis tinc tion is made in
count ing be cause of size dif fer en ti a tion. In ap pear ance spi ral
ves sels have par al lel walls of even in ten sity with clear cen ters. On
oc ca sional piece, walls may be frayed due to grind ing. Walls of very
small ves sels do not ap pear as sharp as those of the larger ones at 200�.
Some spi rals are closely knit to gether. Pho to mi cro graphs of spi ral
ves sels and pos i tive sec tions of them are shown in Fig ure 968.10.
� 2006 AOAC IN TER NA TIONAL
�15% Shell
(0.350 g/50 mL)
Spi ral ves sel counts
>15% Shell
(0.200 g/100 mL)
0 4.5 1.5
1 15 5.8
2 24.4 9.7
3 32.8 13.2
4 40 16.6
5 47 19.7
8 62.2 27.7
11 72.9 34.8
15 83.4 42.4
20 91.1 50.1
30 98.2 62.1
60 80.0
100 86.8
In count ing spi ral ves sels, most pos i tive fields counted will have
eas ily rec og nized pos i tive spi ral ves sel fig ures such as long or short
mass of spi ral ves sels, large bro ken sec tions of these, or some times
tan gled mass of spi ral ves sels. Some sine wave-like fig ures and
some full “S”- or “Z”-like fig ures will be found. There will,
how ever, be some smaller fig ures and some poorly formed “S”- or
“Z”-like fig ures pres ent in some fields. For these fig ures, the
fol low ing ap plies:
“S”- or “Z”-like fig ures or mir ror im ages of these should have 1 3 or
more of top and bot tom nor mal lin ear dis tance for such fig ure and
not just stubs. One-third of cen ter sec tion is es ti mate of this dis tance.
If “2”-like fig ure is found, lower por tion or “V” part of fig ure should
be well extended and suf fi cient top curve should be pres ent so that
fig ure does not ap pear es sen tially like a “V.” Fig ures may be
stretched out. Spi ral ves sels in break ing some times break so that
there will be joined sec tions of half cir cles or less. Count as pos i tive
any such sec tion con sist ing of �3 nearly 1 2 cir cles joined. Two spi ral
cir cles joined to gether are counted pos i tive. Cir cles show ing no
spi ral ing are not counted. A “W” fig ure is pos i tive; a “V” or “C” is
not. In view ing small sec tion of spi rals per pen dic u lar to axis, 2nd
and 3rd spi ral, etc., may be just faintly seen, but the sec tion should
be counted as pos i tive.
There is some fine cell wall struc ture pres ent which when bro ken
may frac ture into “Z”-like char ac ters sim i lar in ap pear ance to “Z”
formed from small thin spi ral ves sels. Care should be ex er cised in
dis crim i nat ing be tween the two.
Ref er ence: JAOAC 51, 725(1968).
Re vised: March 1996
* Adopted as a Co dex De fining Method (Type I) for spi ral ves sel and
stone cell count of co coa shell in co coa (ca cao) nib, co coa (ca cao) mass,
co coa press cake and co coa dust (co coa fines) when man u fac tur ing
co coa and choc o late prod ucts.
Fig ure 968.10. Spi ral ves sel sec tions ca 340�. (A) Mass of spi ral ves sels; note size dif fer en tial and nar row ing
ef fects. (B) Spi ral ves sels stretched out; note dif fer ence in size. (C) Bro ken sec tion; count pos i tive if 3 joined rods are
pres ent. (D) Pos i tive “S”-shaped sec tion.