Method of geochemical prospecting

A method of geochemical prospecting where samples of soil, sediment and rock in a geological area are collected and each sample is contacted with a leach solution of water, glucose and glucose oxidase. The oxidase acts upon the glucose to produce gluconic acid and hydrogen peroxide. The leach solution reacts with manganese dioxide carrying trace metal elements and compounds in the samples to produce a leach reaction solution. The leach reaction solution and the residue of the said sample are separated, and the leach reaction solution is analyzed to determine the trace metal content present. From the analysis, the mineral content of the geological area is predicted.

BACKGROUND OF THE INVENTION 
1. Field of the Invention 
This invention relates to geochemical prospecting and more particularly to 
geochemical prospecting using selective leaches and dissolution 
techniques. 
2. Description of the Prior Art 
By using a geochemical prospecting process, it is possible to infer the 
presence, location and magnitude of ore bodies below the surface. This 
process involves detecting surface areas containing excess localized 
anomalous concentrations of ore metals in comparison with their normal 
concentrations in the region. Such excess may result from diffusion or 
capillary transport of metal ions upward through overburden, or from 
physical transport of ore minerals, or from other hydromorphic processes 
which transport ground water and resulting stream flows. A detailed 
description of such prospecting may be found in "Geochemistry in Mineral 
Exploration" (2nd Ed.) by Rose, Hawkes and Webb, Academic Press, New York, 
1979. 
In an illustrative hydromorphic process, manganese and iron ions, together 
with associated ore metal compounds, are transported by water from below 
ground to the surface, where they precipitate as oxides. The fresh, 
initially largely amorphous, oxides deposit as coatings upon available 
solid bodies, ranging from clay particles to rocks and boulders. These 
oxides in turn scavenge heavy metal ions and metal compounds dissolved in 
the water. Depending upon the deposit conditions, metals such as silver 
(Ag), cobalt (Co) and copper (Cu) will associate predominantly with 
manganese oxides. Lead (Pb) will associate with iron oxide, while zinc 
(Zn) and nickel (Ni) will associate with either. Several forms of 
manganese dioxide occur in geological materials. Most of the manganese 
dioxide (MnO.sub.2) in typical soils or sediments is present in 
crystalline phases. Amorphous and semi-amorphous (partially crystalline) 
manganese dioxide usually are a portion of the MnO.sub.2 in superficial 
geological materials. The amorphous form of this compound is the most 
reactive form, and it is a very effective trap for many trace elements due 
to its complex surface and larger surface area. 
Because the concentration of ore metals is higher in the oxide coatings 
than in the underlying solid bodies, it is advantageous to strip these 
coatings off the substrate before testing them for their metal content. 
This step enhances the contrast between the composition of anomalous 
samples and that of the normal background in the region, and permits a 
more sensitive and extensive delineation of the anomalous area. It is 
further advantageous to be able to strip separately the manganese oxide 
and iron oxide metal-containing materials. 
For this purpose, numerous selective leach solutions have been developed 
which, when applied separately or in carefully-ordered sequences in 
well-known partial dissolution techniques, can strip off one or more of 
the separate components of the coating and resolve the sample into useful 
fractions. Most processes dissolve all of the manganese oxide. See, T. T. 
Chao, "Use of Partial Dissolution Techniques in Geochemical Exploration," 
Journal of Geochemical Exploration, Vol. 20 (1984) pp. 101-135, Elsevier 
Science Publishers, Amsterdam, and the bibliography attached thereto. 
Among these known leach solutions are hydroxylamine hydrochloride, oxalic 
acid and ascorbic acid. Hydroxylamine hydrochloride contains chloride 
ions, which can produce serious analytical interferences, and is not a 
viable leaching agent when seeking many low-level trace-element 
signatures. Ascorbic acid and oxalic acid leaches, on the other hand, are 
not selective for certain oxides. 
Oxalic acid leaching of rock, soil, and stream sediment samples as an 
anomaly-accentuation technique is described by H. V. Alminas and E. M. 
Mosier in U.S. Geological Survey Open File Report 76-275, 25 pp, 1976. A 
rapid partial leach and organic separation for the sensitive determination 
of Ag, Bi, Cd, Cu, Mo, Pb, Sb, and Zn in surface geologic materials by 
flame atomic absorption is described by J. G. Viets, J. R. Clark and W. L. 
Campbell in Journal of Geochemical Exploration, Vol. 20, p. 355-366, 1984. 
While concentrated hydrogen peroxide has been widely used as an oxidant in 
selective leaching processes, particularly for the destruction of organic 
portions of the sample, hydrogen peroxide can also function as a reducing 
agent for several metallic oxides. In an aqueous solution, it will react 
with manganese dioxide, consuming hydrogen ions, resulting in the 
manganese being reduced to the divalent state, which is soluble, thus: 
EQU MnO.sub.2(s) +H.sub.2 O.sub.2 +2H.sup.+ .fwdarw.Mn.sup.2+ +O.sub.2(aq) 
+2H.sub.2 O 
In the process, all trace elements trapped in the manganese dioxide are 
released. See, for example, U.S. Pat. No. 4,872,909, issued Oct. 10, 1989 
to J. P. Allen, et al. for "Process for Acid Leaching of Manganese Oxide 
Ores Aided by Hydrogen Peroxide." It is known, however, that concentrated 
H.sub.2 O.sub.2 will only slowly dissolve some crystalline forms of 
manganese oxide. 
Dilute hydrogen peroxide is a poor oxidizer of metallic gold and sulfide 
minerals. It is known, however, that H.sub.2 O.sub.2 will act in 
combination with other reagents to aid in the leaching of these 
substances. The presence of H.sub.2 O.sub.2 raises the fugacity of oxygen 
in a solution and helps increase the efficiency of dissolution of gold by 
cyanide. Hydrogen peroxide will oxidize halide ions, such as chloride, 
bromide, and iodide, to chlorine, bromine, and iodine. Aqueous solutions 
of these halogen elements can effectively oxidize and dissolve precious 
metals such as gold and many sulfide minerals. 
It is well known in the art of beekeeping, that natural raw honey contains 
a very low concentration of the heat-labile enzyme, glucose oxidase, 
which, acting upon the dextrose in the honey, has the interesting property 
of maintaining a very low, 33 parts per million, concentration of hydrogen 
peroxide in the honey. This peroxide, among other factors, prevents growth 
of pathogenic and degradative microorganisms in the honey. In brief the 
reaction is: 
##STR1## 
OBJECTS OF THE INVENTION 
It is the principal object of the present invention to provide an improved 
method of geochemical prospecting. 
Another object of the present invention is to provide an improved leaching 
solution capable of leaching trace elements and minerals from soils, 
sediments, and rocks without leaching the underlying strata. A related 
object is to provide an improved leaching method utilizing the leaching 
solution, which method can be utilized for geochemical prospecting, 
pollution studies, agricultural studies and the like. 
Another object of the present invention is to provide an improved method of 
geochemical prospecting which utilizes a selective leach capable of 
enhanced selection and separation of manganese oxides and iron oxides from 
the substrate on which they are present. 
A further object of the present invention is to provide an improved method 
of geochemical prospecting utilizing a selective leach which not only is 
able to select and separate manganese oxide and iron oxide components, but 
also does not add materials which interfere with or confound the operation 
of analytical equipment used to analyze low concentrations of metals in 
the leachate. 
A related object of the present invention is to provide an improved 
selective leaching solution for use in the separation and analysis of 
manganese oxides and iron oxides and components carried thereby. 
Still a further object of the present invention is to provide an improved 
leaching solution capable of enhanced separation of amorphous and 
semi-amorphous manganese oxides. 
Still another object of the present invention is to provide an improved 
selective leaching solution capable of separating manganese oxides and 
components carried thereby from iron oxides and components carried 
thereby. 
Another related object of the present invention is to provide an improved 
method of selectively leaching geological materials to dissolve and 
separate manganese oxides therefrom. 
A further object of the present invention is to provide an improved method 
of selectively leaching geological materials wherein manganese oxides and 
components carried thereby are separated from iron oxides and components 
carried thereby. 
SUMMARY OF THE INVENTION 
The present invention is embodied in an improved method of leaching soils, 
sediments, and rocks to remove surface minerals therefrom. The leaching 
solution is then analyzed and the results utilized for geochemical 
prospecting functions, pollution studies, agricultural studies and the 
like. The process utilizes a leaching solution containing highly dilute 
concentrations of hydrogen peroxide, either added to or generated within 
the solution. The preferred procedure is to generate the hydrogen peroxide 
within the solution utilizing a sugar and sugar enzyme reaction. More 
specifically, the dilute concentrations of hydrogen peroxide are formed by 
a leach solution comprising water, glucose, and glucose oxidase which acts 
upon the glucose to produce gluconic acid and hydrogen peroxide. Soil 
samples, sediment samples, rock samples and the like are then subjected to 
the leach solution, the leachate is separated from the solid materials and 
analyzed. The analytical results are utilized for a variety of purposes as 
indicated. For geochemical prospecting, for example, selected soil, 
sediment, and rock samples are contacted with a leach solution comprising 
water, glucose and glucose oxidase. The oxidase acts upon the glucose to 
produce gluconic acid and hydrogen peroxide which in turn leaches reactive 
manganese oxide phases, such as amorphous and semi-amorphous manganese 
dioxide, from the samples. The leach reaction solution is then separated 
from the materials and analyzed to determine quantitatively the mineral 
content thereof. The analytical results are used to predict the presence 
of an underground body of minerals. 
In certain situations, it is desirable to analyze for iron and this can be 
accomplished by adding the enzyme catalase to the leaching solution. The 
catalase reacts with the hydrogen peroxide to produce water and oxygen. 
The destruction of hydrogen peroxide and the consumption of oxygen to 
produce gluconic acid together sufficiently lower the fugacity of oxygen 
of the leach solution to additionally dissolve iron oxides. A similar 
result can be obtained utilizing ascorbic acid. 
For precious metals such as gold and metal sulfides, cyanide and halide 
ions can be added to the leaching solution. After the leaching reaction, 
the leaching reaction solution can be analyzed for precious metals and 
metals originally present as metal sulfides.

DESCRIPTION OF THE PREFERRED EMBODIMENT 
The present invention is broadly embodied in a leaching solution and 
process capable of selectively leaching reactive materials from soils, 
sediments, rocks, and the like, and finds utility in a wide variety of 
applications including geochemical prospecting, heap leaching of precious 
metals, mineral exploration, pollution studies and agricultural studies. 
The concept behind the present invention is the utilization of a leaching 
solution containing highly dilute amounts of hydrogen peroxide, either 
added to, or more preferably, generated directly within the leaching 
solution. One procedure for solution generation of hydrogen peroxide 
involves the use of sugars and sugar enzymes which react with the sugar to 
produce an organic acid or related compound such as an aldehyde, 
corresponding to the type of sugar and dilute concentrations of hydrogen 
peroxide. Also, there are enzymes that react with specific lipids, 
alcohols, cellulose or other organic substrates to produce hydrogen 
peroxide and corresponding byproduct compounds. The hydrogen peroxide 
range runs from 0.000001 molar to 1.0 molar hydrogen peroxide and more 
specifically 0.00001 molar to 0.1 molar peroxide, with a preferred range 
of 0.0001 molar to 0.01 molar peroxide. The solution can be further 
modified by utilizing cyanide ions, such as the alkaline metal cyanides, 
potassium cyanide and sodium cyanide as well as the halide ions, chloride, 
bromide, and iodide. These solutions find utility for leaching precious 
metals such gold and sulfide minerals. The broad range of cyanide and 
halide ions ranges from 0.000002 molar to 2.0 molar, more specifically 
from 0.00002 molar to 0.2 molar, with a preferred content of 0.002 molar 
cyanide or halide ion. 
For some applications, a further enzyme can be added which acts upon the 
hydrogen peroxide to produce water and oxygen and thereby results in a 
leaching solution capable of leaching certain metal oxides such as iron 
oxides. Ascorbic acid when added in small amounts to the leaching 
solution, affords a similar function. 
The present invention finds particular but not necessarily exclusive 
utility in geochemical prospecting applications, particularly for the 
location of various mineral deposits and trace metals carried thereby. The 
process is particularly selective to reactive amorphous and semi-amorphous 
manganese dioxide and trace elements trapped within those phases. This is 
accomplished by utilizing the leaching solution to leach geological 
materials with the intent of locating surface anomalies. 
In the geochemical prospecting process embodying the present invention, 
soil, sediment, or rock samples are collected from a selected area. The 
sample collection sites are carefully mapped and the samples indexed. The 
samples are then analyzed by using an enzyme leach process capable of 
selectively leaching amorphous and semi-amorphous manganese dioxide and 
the trace elements trapped and retained thereby. The enzyme leach 
selectively dissolves relative large amounts of the specific reactive 
forms of manganese oxides desired to form a leachate. The leachate is then 
analyzed to reveal anomalies in the presence of trace minerals. The 
anomalies are then analyzed and studied, and an intelligent prediction is 
made as to the presence of underground mineral deposits. 
The enzyme leaching solution utilized is a weak aqueous sugar solution, to 
which has been added a small amount of an enzyme capable of converting 
sugar to provide a low concentration of hydrogen peroxide and some organic 
acid. This solution is an effective selective leach solution capable of 
selectively dissolving relative large amounts of amorphous manganese 
oxides on the particles in a particular soil, sediment or rock sample, 
resulting in the release of the specific ore metal content, without 
appreciable attack on the iron oxides or the substrates of the particles. 
A similar leach solution, to which has been added an enzyme capable of 
converting hydrogen peroxide to water and oxygen, or to which has been 
added some ascorbic acid, has a sufficiently lowered oxidation potential 
to dissolve iron oxides on the particles and release their ore metal 
content, without attacking the substrates of the particles. The hydrogen 
peroxide which is slowly generated in the enzyme-sugar leach solution, 
reacts with the reactive manganese dioxide which is present. The weak 
organic acid simultaneously produced is an excellent complexer of metals 
and helps to hold them in solution. It also contributes hydrogen ions to 
the reaction between hydrogen peroxide and manganese dioxide. Since the 
hydrogen peroxide is slowly generated, a negative-feedback effect results 
which limits its maximum concentration, and the hydrogen peroxide does not 
reach high concentrations at which it would effectively attack organic 
matter present in the sample. Nor would there be extensive oxidation and 
consequent possible dissolution of reduced mineral substances. The slow 
rate of reaction further allows the user to alter rates of sample leaching 
by adjusting concentrations of the reactants. 
In the following description, reactant concentrations are given in 
weight/volume times 100 or volume/volume.times.100, e.g., 100.times.gm/ml, 
expressed as "percent." For example, 1% represents a concentration of 0.01 
grams (or ml) of reactant per milliliter of solution. Further, amounts of 
enzyme reactants are expressed in terms of weight of the commercially 
prepared powders or volumes of commercially prepared aqueous suspensions 
in which they are obtained, which may typically contain a very small 
percentage of actual enzyme content. 
It is very difficult to determine the weight of many enzymes in the 
commercial preparations in which they are sold. This is the case with 
glucose oxidase. Therefore, the concentration of many enzymes cannot be 
stated in terms such as mg/g or g/ml. There are standardized methods for 
measuring the activity of many enzymes, in which the rate of a reaction 
gives a measure of the activity of the enzyme in "units." Commercial 
preparations of enzymes are assayed by the manufacturer, and the customer 
can duplicate the assay procedure. Commercial enzyme preparations 
generally do not have a constant enzyme activity from lot to lot. The 
manufacturer lists the activity of the enzyme in each lot of product, and 
the customer can adjust the quantity of the product to be used. 
The commercial preparation of glucose oxidase that has been used most often 
in enzyme leach is Sigma Chemical Company product number G-6125. The 
activity of glucose oxidase in that product ranges from 15,000 to 25,000 
units per gram of solid. One unit of glucose oxidase will oxidize 1.0 
micromole of beta-D-glucose to gluconic acid and H.sub.2 O.sub.2 per 
minute at pH 5.1 at 35.degree. C. The resulting amount of H.sub.2 O.sub.2 
produced in the leaching solution may range broadly from 0.000001 molar to 
1.0 molar H.sub.2 O.sub.2. Preferably, the amount of H.sub.2 O.sub.2 
should fall in the range 0.0001 molar to 0.01 molar. 
The commercial preparation of catalase used in leaching has been Sigma 
Chemical product number C-3515, an aqueous suspension. It nominally 
contains 5,000 units catalase per milligram of protein, and it is sold as 
an aqueous suspension. Each individual lot assay lists the units of enzyme 
per milligram protein and the milligrams of protein per ml of suspension. 
One unit of catalase will decompose 1.0 micromole of H.sub.2 O.sub.2 per 
minute at pH 7.0 at 25.degree. C., while the H.sub.2 O.sub.2 
concentration falls from 10.3 to 9.2 micromoles. 
The lots of glucose oxidase that have been used in the present invention 
for the enzyme leach range from 18,000 to 23,800 units per gram of solid. 
Typically, 1.25 g of product G-6125 was dissolved in 25 ml water to make a 
stock enzyme solution containing 0.05 g/ml of G-6125. Then 0.1 ml of this 
solution was mixed with 15 ml of dilute glucose solution to leach a 
sample. This yielded 0.00033 g of G-6125/ml in the leach solution. With 
the possible range of glucose oxidase activity for that product, this 
yields a range of glucose oxidase activity for the leach solution of 5.0 
to 8.3 units/ml of leach solution. 
The selective leach solution must have a glucose concentration of at least 
0.0001%, or 0.000001 g/ml, in order to be effective in leaching a sample 
at a solution-to-sample ratio of 50 to 1, and proportionately more for 
smaller ratios, and no more than 20%, or 0.2 g/ml, in order to avoid 
sticky solutions. The range 0.1% to 5.0%, or 0.001 g/ml to 0.05 g/ml is 
preferred, with the further note that, above about 5.0%, or 0.05 g/ml, 
carbon can deposit on the torch of an ICP/MS analytical instrument, if 
that instrument is used to analyze the leached metals. A 1%, or 0.01 g/ml, 
glucose concentration is preferred. 
The glucose oxidase concentration should be at least 0.0001%, or 0.015 
units/ml, in order to avoid too slow a reaction, and not more than 5.0%, 
or 1250 units/ml, in order to avoid excessive cost. The preferred range of 
glucose oxidase concentration is broadly 0.001% to 0.10%, or more 
narrowly, 0.15 units/ml to 25.0 units/ml. A 0.03%, or 7 units/ml glucose 
oxidase concentration is more specifically preferred. 
For a reasonably rapid reaction to occur, the ratio of the amount of 
glucose oxidase to the amount of glucose should be at least 0.21 units/gm. 
To balance the amounts of glucose and glucose oxidase, the ratio of 
glucose oxidase to glucose should be in the range 0.001 to 1.0, or 21 
units/gm to 21,000 units/gm. A ratio of 0.03, or 700 units/gm is 
preferred. 
Commercial preparations of glucose oxidase frequently contain large amounts 
of another enzyme, catalase, which decomposes hydrogen peroxide into water 
and oxygen, lowering the oxidative potential of the solution. Too much 
catalase in the selective leach solution can lower that potential 
sufficiently that significant amounts of iron oxides can be reduced to the 
ferrous state and thus rendered soluble, along with the manganese oxides. 
This is undesirable when the intent is to analyze the two oxide suites 
separately. There are purified grades of glucose oxidase powder which 
contain only moderate amounts of catalase, of the order of one-tenth of 
their glucose oxidase content, and these can be used for leaches which are 
selective for manganese oxides alone. 
But when the object is to dissolve the iron oxides, either in a second and 
separate treatment of the sample after the manganese oxide components have 
been stripped or in a combined single treatment to strip both the 
manganese and iron oxides, a selective leach comprising catalase, as well 
as glucose and glucose oxidase, can be effective. For this purpose, the 
concentration of catalase can be about equal to that of the glucose 
oxidase. The catalase concentrate should be at least 0.015 units/ml, in 
order to insure a sufficient rate of decomposition of hydrogen peroxides, 
and no more than 1500 units/ml, in order to avoid excessive cost. The 
preferred concentration of catalase in the solution is broadly in the 
range 0.0001% to 0.10%, or more narrowly 0.15 units/ml to 50 units/ml. A 
0.0033%, or 6 units/ml, catalase concentration is preferred. 
Catalase is obtained commercially from microbial and fungal sources and 
also from mammalian livers. Catalase from livers should be avoided, since 
it is frequently contaminated with a wide spectrum of heavy metals. 
Various other commercial enzyme preparations may contain other unwanted 
components, such as unacceptable amounts of sodium chloride, which can 
complicate the task of analyzing the resulting leachate. 
The oxidative potential of a glucose-glucose oxidase selective leach 
solution can be lowered by adding a reducing organic acid such as ascorbic 
acid. The ascorbic acid concentration should be at least 0.0001 g/ml, in 
order to insure sufficient rates of decomposition of hydrogen peroxide, 
and no more than 0.05 g/ml, in order to prevent problems with instrumental 
determinations of elements in the leach solution. The preferred 
concentration of ascorbic acid in the solution is in the range 0.05% to 
1%, or 0.0005 g/ml to 0.01 g/ml, and a 0.1% or 0.001 g/ml, concentration 
is preferred as it performs well and does not deposit excessive carbon in 
analytical instrumentation. 
After a sample has been leached and the supernatant leachate drawn off, 
enough ultrapure nitric acid may be added to make the leachate 1% by 
volume HNO.sub.3, to stabilize it against reprecipitation of some 
elements. Also, some formaldehyde should be added to prevent spoilage by 
microorganisms. 
Many variations of the foregoing method can be devised. For one example, 
addition of a small amount of halide salt to the glucose-glucose oxidase 
leach solution produces an aqueous solution of the halogen, which attacks 
gold, platinum group metals and some sulfide minerals. Addition of 
glucose-glucose oxidase leach solution to a leach containing a 
pseudo-halide such as cyanide or thiocyanate, by raising the oxidation 
potential, increases the rate of leaching of gold, silver, and platinum 
group metals or sulfide minerals. Such variations in the leach have 
potential metallurgical applications for not only mineral exploration but 
also heap leaching ores and mine tailings to recover precious metal 
values. 
The trace amounts of hydrogen peroxide produced by the glucose 
oxidase/glucose reaction will rapidly dissolve a relatively large amount 
(0.024 gram) of amorphous MnO.sub.2 (Example X, below). Refferring to FIG. 
1, the typical manganese content of B-horizon soils in northern Minnesota 
ranges between 400 and 4,000 micrograms/gram (parts per million or ppm). 
Of a set of more than 1600 soil samples from that region the mean 
leachable Mn content with three leaches was as follows: water leach=0.3 
ppm; Enzyme leach=1.5 ppm; Enzyme plus Ascorbic Acid leach=15 ppm (FIG. 
1). Thus, the Enzyme leach, sometimes referred to as ENZ leach, typically 
dissolves less than 1% of the MnO.sub.2 in these soil samples, and the 
Enzyme plus Ascorbic Acid leach, sometimes referred to as ENZ+ASC leach, 
removes less than 10%. This contrasts with the MnO.sub.2 -specific 
hydroxylamine hydrochloride leach which rapidly dissolves more than 60% of 
the total Mn in typical soil samples. 
The present invention is highly selective for the most reactive form(s) of 
manganese dioxide, amorphous and semi-amorphous MnO.sub.2. Because 
amorphous manganese dioxide is such an effective trap for many trace 
elements, the enzyme leach can be used to reveal anomaly contrast in 
samples that show no contrast with stronger partial leaches. 
EXAMPLE I 
Soil samples were collected on Little American Island, located about ten 
miles east of International Falls, Minn., in Rainy Lake. Little American 
Island is highly mineralized. All the material comprising the A-horizon of 
the soil (decaying leaf litter, humus, and organic-rich mineral layers) 
were excavated to reveal reddish-brown B-horizon. Approximately one pound 
of B-horizon material was collected at each site. The sample sites were 
marked as accurately as possible on the 1:24,000-scale Island View, Minn., 
71/2 minute quadrangle map that is published by the U.S. Geological 
Survey. Later, the latitude and longitude of the sample sites were 
determined from the field map using a digitizing pad attached to an 
IBM-compatible personal computer, running the GSMAP.EXE program published 
by the U.S. Geological Survey. For comparative purposes a set of B-horizon 
soil samples was collected from an unmineralized area south of 
International Falls, in the southern part of the Ranier, Minn., 71/2 
minute quadrangle. The sample locations are given in Table 1. 
TABLE 1 
______________________________________ 
Latitudes and Longitudes of sample sites near 
International Falls, MN, used in example of 
analytical results produced with Enzyme leach. 
Site Number Latitude (.degree.N) 
Longitude (.degree.W) 
______________________________________ 
Little American Island set: 
INL 0007 48.602194 93.167530 
INL 0008 48.602211 93.167365 
INL 0009 48.602224 93.167217 
Background set: 
INL 0034 48.509631 93.344618 
INL 0035 48.508923 93.344665 
INL 0036 48.508264 93.344629 
______________________________________ 
The samples were dried at 35.degree. C. for approximately 48 hours and were 
sieved for the minus-0.25-mm (minus-60-mesh) fraction of the sample 
particles. The grain sizes in the sieved samples comprised the fine sand, 
very-fine sand, silt, and clay of these B-horizon soil samples. The 
preferred size fraction may vary with the sample media and location from 
which it was collected. 
All the laboratory procedures described herein were conducted at ambient 
temperature and pressure. All reagent and standard solutions were prepared 
in water that had been purified to a minimum resistance of 16.7 megohms/cm 
using a Barnstead NANOpure II system. For most analyses, distilled water 
would be of sufficient purity. A 0.01 gram/ml (0.0555 molar) glucose 
solution was prepared by dissolving 30 grams of glucose (Sigma Chemical 
Co. product number G-5767) in water and diluting to three liters. An 
enzyme solution was prepared by dissolving 1.25 grams of glucose oxidase 
commercially available powder (Sigma Chemical Co. product number G-6125, 
prepared from Aspergillus niger) in 25 ml of water. The commercial powder 
containing the enzyme was weighed into a clean 25 ml volumetric flask, 
water was added, the contents were gently swirled until the enzyme was 
dissolved, and the solution was diluted to volume and mixed. The glucose 
oxidase activity in this stock solution ranges between 750 and 1,250 units 
per ml of solution. The assay of the particular lot of commercial enzyme 
powder used in this test was 23,800 units of glucose oxidase per gram of 
solid (Lot 46F-9655), yielding an activity in the stock solution of 
approximately 1,190 units/ml in the solution. As long as the glucose stock 
solution is not contaminated with microorganisms, it is stable for an 
indefinite period of time. A fresh glucose oxidase stock solution was 
prepared every 24 hours. 
One gram of each sample was weighed into a separate 16mm.times.150mm 
disposable culture tube, 15.0 ml of the glucose stock solution was added, 
and 0.10 ml (119 units glucose oxidase) of the enzyme solution was added 
to each tube. The final enzyme concentration in the leach solution was 
about 7.9 units/ml. Immediately, each tube was capped with a new plastic 
cap (Fisher brand TainerTop, part number 02-706-33), shaken and vortexed 
to insure complete mixing of the contents. After 60 minutes each culture 
tube was vortexed again and centrifuged for 10 minutes at 2000 rpm. Ten 
(10.00) ml of the supernatant leach solution was carefully drawn off each 
leach tube (using an Eppendorf Maxipettor and Maxitip L) and transferred 
to new 16mm.times.100mm disposable culture tubes. Care was taken to 
prevent transfer of sediment and flotsam from the original culture tube. 
In order to prevent precipitation of trace metals in the leach solutions, 
0.100 ml of concentrated nitric acid (Merck Suprapur brand) was added to 
each tube. In order to prevent the growth of microorganisms, 0.1 ml of 
concentrated formaldehyde (Fisher brand, analyzed reagent grade) also was 
added to each culture tube. At this point internal standard spikes of 
scandium and terbium were added for reasons outlined below. The additions 
of these liquids to the tubes was accomplished using a 100 microliter 
Eppendorf micropipet. Then the tube was sealed with a new cap (Fisher 
brand TainerTop), and the contents were thoroughly agitated to insure 
complete mixing. 
The sample solutions can be analyzed by a variety of instrumental 
techniques, depending on which analytes are to be determined and the 
anticipated concentrations of those analytes in the leach solutions. 
Graphite furnace atomic absorption spectroscopy (GFAAS) or inductively 
coupled plasma/mass spectrometry (ICP/MS) would be appropriate 
instrumental methods for determining the low nanogram/ml 
(part-per-billion) concentrations of trace elements, such as cobalt, zinc, 
and arsenic, which would be expected in Enzyme leach solutions of typical 
soil samples. Inductively coupled plasma/atomic emission spectroscopy can 
be used to determine more geochemically abundant elements, such as 
magnesium, calcium, manganese and sodium. ICP/MS was used to determine 
cobalt, zinc, and arsenic in the soil samples described herein. Since it 
is advisable to use an internal standard for ICP/MS determinations, in 
order to correct for instrumental drift, spikes of one or more elements 
should be added to normalize the ion counts between the sample solutions 
and standard solutions. 
A spike solution was prepared by adding 10 ml each of 1000 microgram/ml Sc 
and Tb, Spex Industries brand, standards to a 1 liter volumetric flask 
(Spex item numbers AQSC2-500 and AATB2-500, respectively, each of which 
was in a 2% (vol./vol.) nitric acid/water matrix). Ten ml of Merck 
Suprapur brand nitric acid also was added to the volumetric flask, the 
contents were diluted to 1.000 liter and mixed. The resulting spike 
solution was 10 micrograms/ml each of scandium and terbium in an 
approximately 0.01 ml/1.0 ml nitric acid/water solution. Indium and 
lutecium were found to be equally effective as internal standards, and the 
stock solution would be prepared in a similar manner. At the time that the 
nitric acid and formaldehyde were added to each leach solution, 0,100 ml 
of this spike solution also was added to the leach solution. 
ICP/MS standards for cobalt, zinc, and arsenic were also prepared from Spex 
brand 1000 microgram/ml standards (Spex item numbers AQCO2-500, AQZN1-500, 
and AAAS2-500, respectively). The commercial cobalt and arsenic standards 
were in 2% (vol./vol.) nitric acid/water solutions, while the zinc 
standard was in a 2% (vol./vol.) hydrochloric acid/water solution. A 
multielement stock standard was prepared by transferring 10.00 ml of each 
of these Spex standards to a 1 liter volumetric flask. Ten ml of Merck 
Suprapur brand nitric acid was added, and the contents were diluted to 
1.000 liter. This yielded a stock standard containing 10 micrograms/ml of 
each cobalt, zinc, and arsenic in a 0.01 ml/1.0 ml nitric acid/water 
matrix. 
A stock blank solution was prepared by dissolving 30 grams glucose (Sigma 
number G-5767) and 1.00 gram glucose oxidase (Sigma number G-6125) in 
water and diluting to 3.0 liters. The solution was transferred to a clean 
brown-glass bottle. Then 30 ml of Merck Suprapur nitric acid and 30 ml 
Fisher brand reagent formaldehyde were added an mixed with the contents of 
the bottle. Ten (10.00) ml of the Sc-Tb spike solution was diluted to 
1.000 liter in a volumetric flask with this blank solution, the contents 
were mixed, and the solution was transferred to a new Nalgene plastic 
bottle. This yielded an instrumental analytical blank. Next, 2.00 ml of 
the 10 microgram/ml Co-Zn-As standard and 10.00 ml of the Sc-Tb spike 
solution were transferred to a 1.000 liter volumetric flask and diluted to 
volume with the stock blank solution. The solution was mixed and 
transferred to a new Nalgene plastic bottle and was used as a 20 
nanograms/ml combined Co-Zn-As instrumental standard. Next, 10.00 ml of 
the 10 microgram/ml Co-Zn-As standard and 10.00 ml of the Sc-Tb spike 
solution were transferred to a 1.000 liter volumetric flask and diluted to 
volume with the stock blank solution. The solution was mixed and 
transferred to a new Nalgene plastic bottle and was used as a 100 
nanograms/ml combined Co-Zn-As instrumental standard. 
A Sciex Elan model 250 instrument was used to make the ICP/MS 
determinations. This instrument uses an inductively-coupled argon plasma 
torch as an ion source for a quadrupole mass spectrometer. Solutions 
containing the analytes are introduced into the plasma by means of a spray 
chamber, from where a flow of argon carries microscopic droplets of the 
solutions through an injector tube into the plasma. Instrumental settings 
were optimized using a 100 nanogram/ml cobalt standard. Readings for 
cobalt, zinc, and arsenic were collected at mass numbers 59, 66, and 75 
respectively. A potential isobaric interference from titanium oxide was 
not a significant problem at mass 66, since the Enzyme leach does not 
effectively dissolve titanium. Simultaneously, readings were collected for 
Sc and Tb at mass numbers 45 and 159. All readings were measured in 
ion-counts per second, and the counts for cobalt, zinc, and arsenic were 
normalized against those for Sc and Tb. The instrumental blank and 
standard solutions were used to calibrate the ICP/MS instrument, and the 
sample solutions were compared to the standards in order to determine the 
amount of each element in the leach solutions. Since the dilution factor 
for the sample weight to leach-solution volume was 15, the analyses for 
the leach solutions were multiplied by 15 to provide the leachable 
concentrations of these elements in the soil samples. 
Enzyme leachable cobalt, zinc and arsenic are substantially higher in the 
Little American Island soil samples than in the background samples (Table 
2). This geochemical anomaly indicates the presence of mineralized bedrock 
on the island. 
TABLE 2 
______________________________________ 
Comparative Enzyme leach data from B-horizon soil 
samples collected near International Falls, Minnesota. Values 
are in nanograms/gram (parts-per-billion) of soil material. 
Sample No. Co Zn As 
______________________________________ 
Little American Island Set (mineralized area): 
INL 0007 79 560 51 
INL 0008 150 2000 1600 
INL 0009 41 240 71 
Background set: 
INL 0034 30 110 9 
INL 0035 38 160 14 
INL 0036 9 150 14 
______________________________________ 
EXAMPLE II 
Enzymes other than glucose oxidase will react with organic substrates to 
produce hydrogen peroxide. Some of these enzymes are available 
commercially, although they are quite expensive relative to glucose 
oxidase. One of these is galactose oxidase, which reacts with galactose 
and oxygen to produce galacto-hexodialdose and hydrogen peroxide. Leaching 
with this enzyme was performed in the same manner as described above. A 
galactose stock solution was prepared that contains 0.01 grams/ml (0.0555 
molar) galactose instead of glucose in water. The galactose oxidase 
solution was prepared by dissolving 5.3 milligrams of Sigma Chemical 
product G-3385 in 0.80 ml water. This product is prepared from Dactylium 
dendroides (Sigma Chemical Co. catalogue). Lot number 50H6813 of this 
product contained 86 units/milligram solid (one unit produces a change in 
absorption at 425 nanometers of 1.0/min. at pH=6 and 25.degree. C. with a 
1 cm light path in a peroxidase and o-tolidine system). One gram of 
geological sample material was leached for one hour with 15 ml of the 
galactose solution and 0.2 ml (114 units enzyme) of the galactose oxidase 
solution. In all other respects this leach was conducted identically as 
described above. The galactose oxidase version of the enzyme leach is 
sometimes referred to herein as the GAOX leach. 
EXAMPLE III 
A stronger version of the enzyme leach uses the enzyme catalase in addition 
to glucose oxidase. Catalase decomposes hydrogen peroxide. The only 
commercial preparation that was found to be acceptable for this process 
was Sigma Chemical Co. product C-3515, which is a suspension prepared from 
Aspergillus niger. The steps in performing this version of the leach were 
the same as for the ENZ leach, except for the following changes. A 
catalase solution was prepared by diluting 0.075 ml of catalase product 
solution (Lot 116F-3828) in 15 ml water. The lot of catalase that was used 
contained 248,000 units of enzyme per ml. The resulting solution contained 
about 1240 units catalase per ml. One gram of each sample material was 
leached with a mixture of 15 ml glucose solution, 0.100 ml of the glucose 
oxidase solution, and 0.100 ml (124 units) of the catalase solution. The 
catalase concentration in the final leach solution was about 8.3 units/ml. 
The leach with catalase was allowed to proceed for 24 hours. In all other 
respects the procedure was the same as with the enzyme leach process 
described above. This version of the enzyme leach process is sometimes 
referred to herein as the ENZ+CAT leach. 
EXAMPLE IV 
Still another version of the enzyme leach uses ascorbic acid in place of 
catalase to achieve a more vigorous leach of the sample. In this procedure 
the ascorbic acid was included in the stock glucose solution, making a 
0.01 gram/ml glucose and 0.001 gram/ml ascorbic acid solution. This was 
done by dissolving 30 grams glucose (Sigma Chemical product number G-5767) 
and 3 grams ascorbic acid (Baker brand product number 7-B581) in water and 
diluting to 3 liters. As with the catalase version of the leach, the 
leaching process was allowed to proceed for 24 hours. Three grams ascorbic 
acid was also included in the three-liter stock blank solution that was 
prepared for making instrumental standards, and a separate set of 
standards was used. All other steps were the same as with the ENZ leach 
process described above. This version of the enzyme leach is referred to 
herein as the ENZ+ASC leach. 
For the purpose of producing comparative analyses, test samples were also 
analyzed using a water leach, where the leaching action of the traces of 
H.sub.2 O.sub.2 produced by the enzyme would not have an effect on the 
results. This leach was conducted in an identical manner to the leach 
described above, except 15 ml of water was used to leach the samples 
instead of the glucose solution, and no glucose oxidase was added. 
Standards for this leach were prepared in a similar manner, except water 
was used for the final dilution instead of the leach blank solution. 
Formaldehyde was not added to either standard or sample solutions. 
In examples discussed below, data is included for vanadium, cobalt, nickel, 
copper, zinc, arsenic, selenium, molybdenum, silver, antimony, tungsten, 
rhenium, gold, thorium, uranium, chlorine, bromine, iodine and manganese. 
Instrumental vanadium, nickel, copper, selenium, molybdenum, silver, 
antimony, tungsten, rhenium, gold, thorium, and uranium were prepared from 
Spex Industries 1000 microgram/ml standards (product numbers AQV2-500, 
AQNI2-500, AQCU2-500, AQSE2-500, PLMO9, AQAG2-500, AQSB5-500, AAW9-500, 
AQRE9-500, AQAU3-500, AQTH4-500, and AQU2-500 respectively). The standards 
were prepared in the same manner as the Co-Zn-As standards described 
above, except the highest concentration silver, tungsten, rhenium and gold 
standard was 20 nanograms/ml. The 10 microgram/ml silver stock standard 
was in a separate solution to prevent silver chloride from precipitating. 
These elements were included in combined standards along with cobalt, zinc 
and arsenic. A 1000 microgram/ml of each chlorine, bromine, and iodine 
stock combined standard was prepared by dissolving 1.51 grams ammonium 
chloride, 1.23 grams ammonium bromide, and 1.14 grams ammonium iodide in 
water and diluting to 1.000 liter in a volumetric flask. A combined 
halogen instrumental standard was prepared by diluting 0.200 ml of this 
stock standard to 1.000 liter with the appropriate blank leach solution, 
producing a 0.200 microgram/ml Cl-Br-I instrumental standard. A Spex 
Industries custom plasma 5.000% Mn standard in 20% nitric acid was used to 
make 2 microgram/ml standards in the appropriate blank matrix for ICP/AES 
determinations of Mn. 
Cobalt, zinc, and Arsenic determinations were performed as described above. 
Vanadium, nickel, copper, selenium, bromine, molybdenum, silver, antimony, 
iodine, tungsten, rhenium, gold, thorium, and rhenium were determined by 
ICP/MS at mass numbers 51, 58, 63, 78, 81, 98, 107, 121, 127, 184, 187, 
197, 234, and 238 (respectively). Chlorine was estimated indirectly by 
ICP/MS as chlorine oxide at mass number 53. Manganese determinations were 
performed by ICP/AES at 257.6 nanometers using an ARL brand model 3580 
fixed channel spectrometer. Generally, enzyme leach solutions are 
sufficiently dilute that interelement interferences are minimal with 
ICP/AES. 
Most elements in the leach solutions described above are stable, and 
determinations can safely be made as much as a month following leaching. 
the halogens are an exception. Dilute HNO.sub.3 and H.sub.2 O.sub.2 will 
combine to rapidly oxidize chloride, bromide, and iodide ions to aqueous 
Cl.sub.2, Br.sub.2, and I.sub.2. The elemental halogens will then diffuse 
into the atmosphere through the polyethylene caps on the culture tubes. 
This oxidation will occur much more slowly if nitric acid is not added to 
the leach solutions. If chlorine, bromine, or iodine are to be determined, 
the leach solutions should not be stabilized with nitric acid, and the 
instrumental determinations should be made within 24 hours after the 
completion of the leaching process. 
EXAMPLE V 
The U.S. Geological Survey exploration reference standard soils were 
developed for use in mineral exploration (U.S. Geological Survey, 
Open-File Report 78-163). These standard soils are known to be 
inhomogeneous from one bottle to another. The results shown here are 
included for comparing the relative results that can be produced by 
different versions of the Enzyme leach. These standard soils can be 
obtained from the U.S. Geological Survey, Branch of Geochemistry, M.S. 
973, Denver Federal Center, Denver, Colo. 80225. 
Standard GXR-2 is a soil sample from the Park City mining district, Summit 
County, Utah. Standard GXR-5 is a B-horizon soil sample collected in 
Somerset County, Me., in an area covered by thin glacial till, where the 
underlying bedrock was mineralized. Leachable cobalt and antimony 
concentrations in GXR-2 and cobalt and nickel in GXR-5 increased with 
increasing intensity of the leaching process (Table 3). The ENZ and GAOX 
leaches appear to be roughly equivalent. The ENZ+ASC leach produces the 
strongest leaching action, and the ENZ+CAT leach is intermediate in 
strength between the ENZ and ENZ+ASC leaches. 
TABLE 3 
______________________________________ 
Comparative leach results for U.S. Geological Survey 
exploration reference standards GXR-2 and GXR-5. 
The leachable concentrations of Co, Sb, and Ni are 
given in nanograms/gram (parts per billion). 
______________________________________ 
GXR-2: Leach Co Sb 
______________________________________ 
water 40 585 
GAOX 54 735 
ENZ 55 915 
ENZ + CAT 144 2400 
ENZ + ASC 420 3300 
______________________________________ 
GXR-5: Leach Co Ni 
______________________________________ 
water 150 630 
GAOX 195 780 
ENZ 180 705 
ENZ + CAT 510 1470 
ENZ + ASC 870 2250 
______________________________________ 
EXAMPLE VI 
Natural trace-element pollution and man-made pollution are dispersed by the 
same geochemical processes. The same processes also determine availability 
of trace elements in soils and sediments to plants and other organisms. 
Therefore, methods that are employed in geochemical mineral exploration 
are often used in studying anthropogenic pollution. Partial analysis also 
is often used by agricultural scientists to determine the availability of 
nutrient and toxic trace elements to plants. 
Water from the Argo tunnel, which was constructed to drain the mines in 
Central City, Colo., empties into Clear Creek near the town of Idaho 
Springs, Colo. Much of the bedrock outcropping in the Clear Creek drainage 
is strongly mineralized, resulting in high metal concentrations in the 
stream sediments. Numerous old mine dumps upstream from Idaho Springs also 
contribute metals to the sediments in the drainage. The effluent from the 
Argo tunnel is highly enriched in metals. An active stream sediment sample 
was collected 200 yards downstream from the point where Argo tunnel waste 
entered Clear Creek. Partial leaching offers a means of determining what 
portion of metals in a soil or sediment may have been added recently and 
what portion has resided in the sample material for a longer period of 
time. For comparison purposes, a stream sediment collected in Turkey 
Creek, 1.0 mile upstream from U.S. Highway 285, Jefferson County, Colo., 
was also analyzed. The Turkey Creek drainage does not contain highly 
mineralized bedrock although similar types of bedrock outcrop in both 
drainages. Although there has been human activity in the Turkey Creek 
drainage for over a century, anthropogenic pollution is minor compared to 
the Clear Creek basin. These samples were air dried and sieved for the 
minus-0.25-mm particle-size fraction. Both samples were subjected to the 
water leach, the GAOX leach, the ENZ leach, the ENZ+CAT leach, and the 
ENZ+ASC leach. The results are given in Table 4. 
TABLE 4 
______________________________________ 
Leachable concentrations of Co, Zn, and Mo in stream 
sediment samples collected in Clear Creek, 
downstream form the Argo Tunnel, and in Turkey 
Creek, using five partial leaches. The leachable 
concentrations of Co, Zn, and Mo are given in 
nanograms/gram (parts per billion). 
Leach/Sample Co Zn Mo 
______________________________________ 
water-Argo tunnel 
75 6,150 435 
water-Turkey Creek 
6 120 15 
GAOX-Argo tunnel 91 8,600 570 
GAOX-Turkey Creek 
17 150 27 
ENZ-Argo tunnel 165 12,450 645 
ENZ-Turkey Creek 28 180 34 
ENZ + CAT-Argo tunnel 
960 210,000 735 
ENZ + CAT-Turkey Creek 
300 3,300 41 
ENZ + ASC-Argo tunnel 
2,850 150,000 3,600 
ENZ + ASC-Turkey Creek 
750 4,200 61 
______________________________________ 
Referring to Table 4, with regard to only these two samples, the order of 
leaches shown in Table 4 is the order of progressively stronger leaching. 
In this case, the GAOX leach produces slightly weaker leaching action than 
the ENZ leach. There is substantial contrast between the Argo tunnel and 
the Turkey Creek samples with all of the partial leaching methods used. 
Thus, any of these weak partial leaching procedures is viable for use in 
mineral exploration in that region. Relatively large quantities of metals 
in the Argo tunnel sample are water soluble, loosely bound, or trapped by 
amorphous or semi-amorphous MnO.sub.2. Any metals contributed in solution 
to Clear Creek by Argo tunnel effluent will most likely be present in such 
forms. Much larger portions of these metals are held more deeply in oxide 
coatings on sediment grains and are probably derived from sources upstream 
from the Argo tunnel. Therefore, these partial leaches can be used to 
determine possible sources for both anthropogenic and natural pollution. 
EXAMPLE VII 
A very low-grade mineral prospect occurs about 2.5 miles south of Indus, 
Minn., in an area where the bedrock is typically covered by 5 to 30 feet 
of till and glacial-lake sediment. B-horizon soil samples were collected 
along a traverse across the mineralized area (sample sites listed in Table 
5). The samples were dried and sieved as described above. These samples 
were analyzed using the ENZ leach and a much stronger potassium 
iodide+ascorbic acid leach (Viets and others, 1984). Copper gives 
four-times better anomaly/background contrast with the ENZ leach than with 
the KI+ascorbic acid leach (FIG. 2A). Zinc produces no discernable 
contrast with the conventional partial leach, while an anomaly contrast of 
two-times background was found with the ENZ leach (FIG. 2B). No antimony 
(Sb) anomaly could be detected with the conventional leach, but a small 
antimony (Sb) anomaly was found with the ENZ leach (FIG. 2C). In this 
situation the highly selective nature of the enzyme leach makes it much 
better suited to detecting subtle geochemical anomalies than the 
conventional KI+ascorbic acid leach. 
TABLE 5 
______________________________________ 
Latitudes and Longitudes of soil sample sites over 
mineral prospect south of Indus, MN. 
Site Number Latitude (.degree.N) 
Longitude (.degree.W) 
______________________________________ 
INL 0039 48.592165 93.862022 
INL 0040 48.592769 93.862063 
INL 0041 48.593291 93.862060 
INL 0043 48.593680 93.862128 
INL 0044 48.593939 93.862069 
INL 0045 48.594445 93.862057 
INL 0046 48.595055 93.862057 
INL 0047 48.599014 93.861813 
______________________________________ 
EXAMPLE VIII 
The Sleeper gold mine, located along the west flank of the Slumbering Hills 
about 30 miles northwest of Winnemucca, Nev., was covered by basin fill 
unrelated to the mineralized bedrock of the deposit. Undisturbed soil 
samples were collected along a traverse across the mineralized structure 
north of the mine pit, and background soil samples were collected upslope 
from that structure (FIG. 3). The overburden thickness was about 50 feet 
at sample site c13, and it dropped off to about 150 feet thick at site 
c24. All samples were collected at a depth of 10 to 18 inches to avoid 
wind-blown contamination. The samples were air dried and sieved for the 
minus-0.25-mm fraction of the soil. For comparative purposes three partial 
leaches were used to analyze the sieved samples. The oxalic acid leach 
(Alminas and Mosier, 1976) and the KI+ascorbic acid leach (Viets and 
others, 1984) were used to show the results with conventional methods of 
partial analysis, and the ENZ leach was used to detect any subtle 
anomalies that might be present. 
A large number of elements were found to be anomalous with the ENZ leach in 
an area near the mineralized structure. The ENZ-leach anomalies for 
chlorine, bromine, and iodine are shown in FIGS. 4, 5, and 6, 
respectively. More elements were found to be anomalous with the ENZ leach, 
and the anomaly to background ratio was substantially greater with the ENZ 
leach, compared to the oxalic acid and KI+ascorbic acid leaches (FIG. 7). 
Standards for elements in this figure were prepared as described above in 
a similar manner using the appropriate Spex standards. ICP/MS was used for 
ENZ leach and oxalic acid leach determinations, and ICP/AES was used for 
KI+ascorbic acid leach determinations. The Enzyme leach detected 
geochemical anomalies related to mineralized bedrock in this particular 
desert pediment situation better than either of the conventional methods 
of partial analysis tested. The KI+ascorbic acid leach did not detect Br, 
Cl, I, V, Re, W, Th, U, Co, or Ni. 
It was discovered in this test that chlorine, bromine, and iodine are key 
pathfinder elements in certain overburden situations. The ENZ leach 
produces much greater contrast for these elements than was found with the 
oxalic acid leach. It may be that these halogen elements are present in 
the samples as halide salts, such as NaBr or MnI.sub.3, or more easily 
oxidized bromine and iodine may be present as bromate or iodate salts, 
such as KBrO.sub.3 or NaIO.sub.3. These salts may be encapsulated by 
manganese oxide coatings or the various halogen ions may actually be 
trapped by amorphous MnO.sub.2. In either case, the leaching action of 
dilute H.sub.2 O.sub.2 can release these elements, while the halogen 
elements, regardless of oxidation states, in the mineral substrates 
beneath the oxide coatings are not leached. 
EXAMPLE IX 
In order to determine the effectiveness of the present process for use in 
the extraction of gold, silver, copper, and other sulfide minerals, tests 
were run in which small amounts of cyanide, iodide, bromide, or chloride 
were added to solutions that were otherwise set up like ENZ leaches. Stock 
solutions were prepared by separately dissolving each of the following 
salts in water and diluting to 250 ml in a volumetric flask; 3.68 grams 
NaCN, 10.87 grams NH.sub.4 I, 7.35 grams NH.sub.4 Br, and 4.01 grams 
NH.sub.4 Cl. This produced stock solutions that were 0.30 molar in the 
respective salt. U.S. Geological Survey exploration standard GXR-4 is a 
copper-mine millhead that contains moderate amounts of metal-rich sulfide 
minerals. USGS exploration standard GXR-6 is a composite B-horizon soil 
sample from a gold district in Davidson County, N.C. One (1.00) gram 
portions of each of these standards were weighed out for leaching with 
96-hour adaptations of the ENZ leach: the ENZ leach alone, the ENZ+cyanide 
leach, the ENZ+iodide leach, the ENZ+bromide leach, and the ENZ+chloride 
leach. Fifteen ml of 0.01 g/ml glucose solution was added to each 1.00 
gram portion of sample, 0.100 ml (119 units) glucose oxidase solution and 
0.100 ml of appropriate cyanide or halide salt solution was added to each 
tube, and the tubes were capped and thoroughly mixed. The final 
concentration of each salt in its respective leach solutions was about 
0.002 molar. Each tube was again thoroughly mixed at 24 hours, 48 hours, 
72 hours, and 96 hours. At 24 and 48 hours the caps were removed and 
resealed to allow air into the tubes, prior to mixing. After the final 
mixing, the tubes were centrifuged for 10 minutes at 2000 rpm. From that 
point on the leaches were treated as ENZ leaches, and the solutions were 
analyzed for Cu, Ag, and Au (Table 6). 
TABLE 6 
______________________________________ 
Extractable Cu, Ag, and Au in GXR-4 and GXR-6 after 
leaching for 96 hours with the ENZ leach, the ENZ + cyanide 
leach, the ENZ + iodide leach, the ENZ + bromide leach, and 
the ENZ + chloride leach. The leachable concentrations are 
given in nanograms/gram (parts per billion). 
Standard Leach Cu Ag Au 
______________________________________ 
GXR-4: ENZ 3000 &lt;5 &lt;3 
ENZ + CN 37500 &lt;5 195 
ENZ + I 11850 9 &lt;3 
ENZ + Br 4500 40 &lt;3 
ENZ + Cl 4800 5 &lt;3 
GXR-6: ENZ -- &lt;5 &lt;3 
ENZ + CN -- 28 60 
ENZ + I -- 10 34 
ENZ + Br -- 43 6 
ENZ + Cl -- 10 4 
______________________________________ 
Copper and silver in GXR-4 are primarily contained within the sulfide 
minerals, such as chalcopyrite, while gold possibly occurs as metallic 
inclusions within pyrite. Gold and silver in GXR-6 are probably present as 
native metals within the soil. These tests show that solutions containing 
sodium or potassium cyanide, iodide, bromide, or chloride salts in 
combination with glucose and glucose oxidase can be used for leaching 
metals from sulfide minerals (GXR-4) or for leaching of precious metals 
from oxidized geological materials (GXR-6). Where extended leaching times 
do not present a problem, such as heap-leaching operations, these 
adaptions of the enzyme leach have potential applications. Maintaining a 
low hydrogen peroxide concentration in a leach solution by replenishing 
the spent H.sub.2 O.sub.2 by means of an enzyme reaction will have 
distinct metallurgical advantages in some heap leaching operations. These 
variations of the leaching process can also be used to detect precious 
metal and base metal anomalies in geological materials. 
EXAMPLE X 
A Mn solution was prepared by dissolving 21 grams of Baker analyzed reagent 
grade manganous carbonate in 100 ml deionized water and 10 ml sulfuric 
acid. Any MnO.sub.2 in the mixture was dissolved by adding 2 ml 
concentrated (30%) H.sub.2 O.sub.2 and mixing. Two hundred ml of Baker 
reagent grade phosphoric acid was added and the solution was diluted to 
one liter with distilled water, making a 0.005 grams/ml Mn solution. Two 
grams of pure quartz granules (1 mm to 2 mm in diameter) were weighed into 
a 16 mm.times.150 mm culture tube. These granules provided an inert 
surface upon which amorphous MnO.sub.2 precipitate can form. Virtually any 
nonreactive surface can be provided for this purpose, from ceramic beads 
to stainless steel shot. Next, 0.5 grams of potassium bromate (Baker 
analyzed reagent grade) and 3 ml of the Mn solution were added. The 
mixture was heated for 1 hour to ensure that all the Mn was oxidized to 
MnO.sub.2. The result was an artificial amorphous MnO.sub.2 coating on the 
quartz granules and the wall of the culture tube. The aqueous solution was 
decanted, and the solid contents were rinsed with 10 ml distilled water 
ten times, decanting the rinse water after each rinse. After the last 
rinse, the contents of the tube were warmed on a hot plate until dry. 
A solution containing 0.10 grams/ml glucose in distilled water was 
prepared. Three ml of this glucose solution was added to the culture tube 
containing the artificial MnO.sub.2 coating. Next, 0.08 grams of glucose 
oxidase (Sigma Chemical Co. product number G-6125) was added, the tube was 
sealed with a plastic stopper, and the contents were gently shaken to 
dissolve the enzyme and to uniformly mix the contents. Every ten minutes, 
the tube was gently rotated and sloshed to continue mixing the reactants. 
Within 15 minutes there was a visible difference in the thickness and 
evenness of the MnO.sub.2 coating. After 30 minutes, some of the quartz 
granules were essentially free of MnO.sub.2 coating. At 1 hour, almost all 
the MnO.sub.2 coating was gone, and after 2 hours the entire MnO.sub.2 
coating had been leached from the contents of the tube. Analysis of the 
leach solution confirmed that &gt;96% of the Mn added to the tube was 
recovered in the leach solution. 
While certain illustrative embodiments of the present invention have been 
shown in the drawings and described in detail in the specification, it 
should be understood that there is no intention to limit the invention to 
the specific form and embodiments disclosed. On the contrary, the 
intention is to cover all modifications, alternative constructions, 
equivalents and uses falling within the spirit and scope of the invention 
as expressed in the appended claims.