U.S. patent application number 12/573682 was filed with the patent office on 2010-04-08 for compositions and methods for controlling a honey bee parasitic mite.
This patent application is currently assigned to John I. Haas. Invention is credited to Gene Probasco.
Application Number | 20100087121 12/573682 |
Document ID | / |
Family ID | 38559799 |
Filed Date | 2010-04-08 |
United States Patent
Application |
20100087121 |
Kind Code |
A1 |
Probasco; Gene |
April 8, 2010 |
COMPOSITIONS AND METHODS FOR CONTROLLING A HONEY BEE PARASITIC
MITE
Abstract
As described below, the present invention provides methods and
compositions for controlling a honey bee parasitic mite. In
addition, the invention features compositions useful for the
treatment or prevention of a parasitic mite infestation in a honey
bee hive.
Inventors: |
Probasco; Gene; (Yakima,
WA) |
Correspondence
Address: |
EDWARDS ANGELL PALMER & DODGE LLP
P.O. BOX 55874
BOSTON
MA
02205
US
|
Assignee: |
Haas; John I.
Washington
DC
|
Family ID: |
38559799 |
Appl. No.: |
12/573682 |
Filed: |
October 5, 2009 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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11732504 |
Apr 2, 2007 |
7597912 |
|
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12573682 |
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11396360 |
Mar 31, 2006 |
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11732504 |
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Current U.S.
Class: |
449/61 ;
424/725 |
Current CPC
Class: |
A01N 65/00 20130101;
A01N 65/08 20130101; A01K 51/00 20130101; A01N 37/40 20130101; A61P
33/14 20180101 |
Class at
Publication: |
449/61 ;
424/725 |
International
Class: |
A01K 51/00 20060101
A01K051/00; A61K 36/185 20060101 A61K036/185 |
Claims
1-16. (canceled)
17. A miticide delivery device for treating or preventing a honey
bee parasitic mite infestation, the miticide delivery device
comprising an effective amount of a hop derivative in a suitable
form for delivery to a honey bee parasitic mite on a honey bee or
to a box hive.
18. The miticide delivery device of claim 17, wherein the hop
derivative is an alpha acid, beta acid, or combination of an alpha
and a beta acid.
19. The miticide delivery device of claim 18, wherein the miticide
delivery device comprises at least about 2.5, 7.5, or 15% alpha
acids
20-21. (canceled)
22. The miticide delivery device of claim 18, wherein the miticide
delivery device comprises at least about 2.5 or 5% beta acids.
23-24. (canceled)
25. The miticide delivery device of claim 17, wherein the miticide
delivery device comprises at least about 2.5% beta acid and at
least about 7.5% alpha acids.
26. The miticide delivery device of claim 17, wherein the form
suitable for delivery to a honey bee parasitic mite is selected
from the group consisting of a liquid, a powder, an oil, an
emulsion, a capsule, and a vapor.
27. The miticide delivery device of claim 17, wherein the miticide
delivery device further comprises a carrier.
28. The miticide delivery device of claim 17, wherein the miticide
delivery device provides for controlled release of a hop
derivative.
29-30. (canceled)
31. The miticide delivery device of claim 28, wherein the hop acid
maintains miticidal activity for at least about six months, nine
months, twelve months, eighteen months, twenty-four months, or
thirty-six months.
32-33. (canceled)
34. The miticide delivery device of claim 17, wherein the device is
selected from the group consisting of a porous pouch, strip,
controlled release strip, tablet, reservoir, polymer disc,
evaporation device, fiber, tube, polymeric block, membrane, pellet,
and microcapillary.
35-55. (canceled)
Description
CROSS-REFERENCE TO RELATED APPLICATION
[0001] This application is a continuation in part of U.S.
application Ser. No. 11/396,360, filed on Mar. 31, 2006, the entire
contents of which are incorporated herein by reference.
BACKGROUND OF THE INVENTION
[0002] Honey bees, Apis mellifera, are required for the effective
pollination of crops and are therefore critical to world
agriculture. Honey bees also produce economically important
products, including honey and bees wax. Honey bees are susceptible
to a number of parasites and pathogens, including the ectoparasitic
mite, Varroa destructor. Varroa mites parasitize pupae and adult
bees and reproduce in the pupal brood cells. The mites use their
mouths to puncture the exoskeleton and feed on the bee's hemolymph.
These wound sites in the exoskeleton harbor bacterial infections,
such as Melissococcus pluton, which causes European foulbrood. In
addition, to their parasitic effects, Varroa mites are suspected to
act as vectors for a number of honey bee pathogens, including
deformed wing virus (DWV), Kashmir bee virus (KBV), acute bee
paralysis virus (ABPV) and black queen cell virus (BQCV), and may
weaken the immune systems of their hosts, leaving them vulnerable
to infections. If left untreated Varroa infestations typically
result in colony-level mortality. Maintaining a supply of strong
honey bee colonies available for pollination is essential for the
sustained production of farm crops worth more than $14 billion to
U.S. agriculture. During the winter of 2004-2005, an estimated 40%
of the honey bee colonies in the U.S. were weakened or collapsed
due to Varroa infestation. Current methods of treating Varroa
infestations are proving to be ineffective as the mites develop
resistance to existing miticides. In addition, the use of such
miticides may introduce injurious chemicals into honey that is
intended for human consumption. New compositions and methods for
treating or preventing Varroa mite infestations are urgently
required. Desirably, such compositions would include only natural
ingredients that pose no risk to human health.
SUMMARY OF THE INVENTION
[0003] As described below, the present invention features methods
and compositions for controlling a honey bee parasitic mite or for
the treatment or prevention of a parasitic mite infestation in a
honey bee hive.
[0004] In general, the invention provides a method of controlling a
honey bee parasitic mite (e.g., Varroa mite, tracheal mite). The
method involves contacting the parasitic mite with an effective
amount of a composition comprising a hop derivative (e.g., alpha
acid, beta acid, or combination thereof), thereby controlling a
honey bee parasitic mite. In one embodiment, the contacting of the
mite occurs while the mite is in contact with a honey bee (e.g.,
honey bee egg, larva, or pupa).
[0005] In a related aspect, the invention provides a method of
treating or preventing a parasitic mite infestation of a honey bee
hive. The method involves contacting the hive with an effective
amount of a composition comprising a hop derivative, thereby
treating or preventing a parasitic mite infestation in a honey bee
hive.
[0006] In another aspect, the method herein further include those
wherein the hive or mite is identified as in need of the treatment
or prevention protocols delineated herein.
[0007] In another aspect, the invention provides a composition for
treating or preventing a mite infestation, the composition
comprising an effective amount of a hop derivative in a suitable
form for delivery to a mite. Suitable forms include, for example,
any one or more of a liquid, a powder, an oil, an emulsion, a
paste, a capsule, a vapor, or any other form capable of delivering
a hop derivative to a Varroa mite in contact with a honey bee or
honey bee hive. If desired, the composition further comprises a
carrier.
[0008] In yet another aspect, the invention features a composition
for treating or preventing a mite infestation, the composition
comprising an effective amount of a hop derivative in a suitable
form for delivery to a mite. In one embodiment, the hop derivative
is an alpha acid, beta acid, or combination of an alpha and a beta
acid. In another embodiment, the hop derivative is a metal alkali
salt or earth alkaline metal salt of hop acid. In other
embodiments, the composition comprises at least 2.5%, 5%, 7.5%, 15%
alpha acids, beta acids, or a combination of alpha and beta acids.
In another embodiment, the form is selected from the group
consisting of a liquid, a powder, an oil, an emulsion, a capsule,
and a vapor. In yet another embodiment, the composition further
comprises a carrier (e.g., any one or more of maltodextrin, cluster
dextrin, corn starch, corn syrup solids, glucose, cyclodextrin,
arabic gum, calaginan, inuline, rosin, partially hydrogenated
soybean oil, cellulose, hydroxymethyl cellulose, hydroxyethyl
cellulose, hydroxypropyl cellulose, and hypomellose). In yet other
embodiments, the composition is a powder having particles of a size
selected from the group consisting of 1 .mu.m, 5 .mu.m, 10 .mu.m,
25 .mu.m, 50 .mu.m, 75 .mu.m, 100 .mu.m, 150 .mu.m, 200 .mu.m, 500
.mu.m, 1 mm, 2 mm, or 5 mm, such as a powder prepared by spray
drying.
[0009] In yet another embodiment, the invention provides a miticide
composition comprising an effective amount of a hop acid alkali
metal salt or hop acid alkaline earth metal salt and a carrier in a
suitable form for delivery to a mite. In one embodiment, the hop
acid alkali metal salt is a sodium, potassium, or lithium salt. In
another embodiment, the hop acid alkaline earth metal salt is
calcium or magnesium. In yet another embodiment, the carrier is
selected from the group consisting of maltodextrin, cluster
dextrin, corn starch, corn syrup solids, glucose, cyclodextrin,
arabic gum, calaginan, inuline, rosin, partially hydrogenated
soybean oil, cellulose, hydroxymethyl cellulose, hydroxyethyl
cellulose, hydroxypropyl cellulose, and hypomellose. In still other
embodiments, the composition is spray dried to form particles of a
size selected from the group consisting of 1 .mu.m, 5 .mu.m, 10
.mu.m, 25 .mu.m, 50 .mu.m, 75 .mu.m, 100 .mu.m, 150 .mu.m, 200
.mu.m, 500 .mu.m, 1 mm, 2 mm, and 5 mm in size. In still other
embodiments, the hop acid to carrier ratio is selected from the
group consisting of 1:2, 1:5, 1:10, 1:20, 1:50, and 1:100. In other
embodiments, the hop acid maintains miticidal activity for at least
about six months, nine months, twelve months, eighteen months,
twenty-four months, or thirty-six months. In still other
embodiments, the hop acid maintains stability for at least about
six months, nine months, twelve months, eighteen months,
twenty-four months, or thirty-six months.
[0010] In yet another aspect, the invention provides a controlled
release composition for treating or preventing a parasitic mite
infestation, the composition comprising an effective amount of a
hop derivative in a suitable form for delivery to a honey bee
parasitic mite.
[0011] In yet another aspect, the invention provides a miticide
delivery device, the device comprising a composition of any
previous aspect. In one embodiment, the device is selected from the
group consisting of a strip (e.g., membranes, paper, plastic, or
polymer strip), controlled release strip, tablet, reservoir,
polymer disc, evaporation device, fiber, tube, polymeric block,
membrane, pellet, tray, and microcapillary. If desired, any of
these devices can be formulated in a biodegradable form.
[0012] In yet another aspect, the invention provides a hive
comprising a composition of any previous aspect.
[0013] In yet another aspect, the invention provides a honey bee
product produced in a hive of any previous aspect. Honey bee
products include, but are not limited to, honey, honey comb, and
bees wax.
[0014] In yet another aspect, the invention provides a kit for the
treatment or prevention of an parasitic mite infestation, the kit
comprising an effective amount of a hop derivative in a form
suitable for delivery to a site of infestation (e.g., a bee hive or
a bee). In one embodiment, the kit features directions for use in a
method of the invention.
[0015] In yet another aspect, the invention provides a method of
identifying a hop derivative that disrupts a biological function of
a honey bee parasitic mite. The method involves contacting the
parasitic mite with a test composition comprising a hop derivative;
and assaying a parasitic mite biological function. In one
embodiment, the test composition disrupts a parasitic mite
biological function (e.g., kills or incapacitates the parasitic
mite or reduces parasitic mite reproduction). In another
embodiment, the method further includes the steps of contacting a
honey bee with the test composition; and assaying a honey bee
biological function. In yet another embodiment, the method
identifies a test compound that does or does not disrupt a honey
bee biological function. In another embodiment, the method
identifies a test compound that kills a honey bee.
[0016] In a related aspect, the invention provides a method of
identifying a hop derivative that does not disrupt a biological
function of a honey bee. The method involves contacting the honey
bee with a test composition comprising a hop derivative; and
assaying a honey bee biological function. In one embodiment, the
method identifies a test compound that does or does not disrupt a
honey bee biological function. In another embodiment, the test
compound kills a honey bee.
[0017] In various embodiments of any previous aspect, a hop
derivative is an alpha acid or a beta acid. In other embodiments of
a previous aspect, a composition of the invention contains an alpha
acid, a beta acid, or a combination thereof, wherein the amount of
alpha or beta acid in the composition ranges between 1% and 100%,
where the bottom limit of the range is any integer between 1 and
99% and the upper limit of the range is any integer between 2% and
100%. Exemplary amounts of an alpha, a beta, or a combination
thereof include at least 1%, 2.5%, 5%, 7.5%, 10%, 12%, 15%, 20%,
25%, 35%, 40%, 50%, 60%, 75%, 85%, 90% or 95% in a composition. In
one particular embodiment, the composition comprises at least 1%,
2%, 2.5%, 3%, 5%, or 10% beta acid and at least 1%, 2%, 3%, 5%, 6%,
7.5%, 8%, 9%, 10%, 12%, or 15% alpha acids. In one embodiment of
any of the above aspects, the contacting disrupts a biological
function of a mite. Exemplary biological functions include any one
or more of respiration, neural activity, locomotion, reproduction,
or any other physiological activity required for mite survival. In
one embodiment, the contacting kills the mite.
[0018] In still other embodiments of any previous aspect, the hop
acid alkali metal salt is a sodium, potassium, or lithium salt. In
other embodiments of any previous aspect, the hop acid alkaline
earth metal salt is calcium or magnesium. In other embodiments of
any previous aspect, the carrier is selected from the group
consisting of maltodextrin, cluster dextrin, corn starch, corn
syrup solids, glucose, cyclodextrin, arabic gum, calaginan,
inuline, rosin, partially hydrogenated soybean oil, cellulose,
hydroxymethyl cellulose, hydroxyethyl cellulose, hydroxypropyl
cellulose, and hypomellose. In still other embodiments of any
previous aspect, the composition is spray dried to form particles
of a size selected from the group consisting of 1 .mu.m, 5 .mu.m,
10 .mu.m, 25 .mu.m, 50 .mu.m, 75 .mu.m, 100 .mu.m, 150 .mu.m, 200
.mu.m, 500 .mu.m, 1 mm, 2 mm, and 5 mm in size. In still other
embodiments, the hop acid to carrier ratio is selected from the
group consisting of 1:2, 1:5, 1:10, 1:20, 1:50, and 1:100. In other
embodiments, the hop acid maintains miticidal activity for at least
about six months, nine months, twelve months, eighteen months,
twenty-four months, or thirty-six months. In still other
embodiments, the hop acid maintains stability for at least about
six months, nine months, twelve months, eighteen months,
twenty-four months, or thirty-six months.
[0019] In yet other embodiments, the composition of the invention
is a controlled release composition wherein the hop derivative is
released over the course of at least one week to 12 months. For
example, the hop derivative is released over at least 5, 10, 14,
28, 36, 41, or 48 days; or is released over the course of 1, 2, 4,
6, 8, 10 or 12 weeks, or even for as long as 5, 6, 9, or 12
months.
[0020] Other features and advantages of the invention will be
apparent from the detailed description, and from the claims.
DEFINITIONS
[0021] By "acarid" is meant an arachnid of the order Acarina, which
includes mites and ticks. By "alpha acid" is meant an organic acid
derived from a hop plant (Humulus lupulus) having structural
homology to a humulone, adhumulone, cohumulone, or an analog or
derivative thereof. Humulone, adhumulone, and cohumulone are the
three most abundant alpha acid analogs. Other exemplary derivatives
of an alpha acid include, but are not limited to isoalpha acids,
rhoisoalpha acids, tetrahydroisoalpha acids, and hexahydroisoalpha
acids.
[0022] By "beta acid" is meant an organic acid derived from a hop
plant (Humulus lupulus) having structural homology to a lupulone,
adlupulone, colupulone or an analog or derivative thereof.
Lupulone, adlupulone, and colupulone are the three most abundant
beta acid analogs. Other exemplary derivatives of a beta acid
include, but are not limited to, hulupones, hexahydrobeta acids and
hexahydro hulupones.
[0023] By "biological function" is meant any physiological or
behavioral activity of an organism. Exemplary biological functions
include reproduction, respiration, neural activity, locomotion.
Honey production is a biological function that is specific to a
honey bee.
[0024] In this disclosure, "comprises," "comprising," "containing"
and "having" and the like can have the meaning ascribed to them in
U.S. patent law and can mean "includes," "including," and the like;
"consisting essentially of" or "consists essentially" likewise has
the meaning ascribed in U.S. patent law and the term is open-ended,
allowing for the presence of more than that which is recited so
long as basic or novel characteristics of that which is recited is
not changed by the presence of more than that which is recited, but
excludes prior art embodiments.
[0025] By "contacting" is meant touching, associating with, or
having proximity to a composition. For example, a hop derivative
may contact a hive either inside or outside of the hive
structure.
[0026] By "controlled release" is meant released over the course of
hours, days, weeks, or months.
[0027] By "controlling a parasitic mite" is meant inhibiting mite
survival or reducing, slowing, or stabilizing the growth of a mite
population.
[0028] By "comb" is meant sections of hexagonal bee wax cells that
are used to rear honey bee progeny ("brood") and store honey and
pollen.
[0029] By "effective amount of a miticide" is meant an amount
effective to disrupt a mite biological function.
[0030] By "hive" is meant a structure that contains a bee colony. A
modern box hive typically includes a bottom board, cover, and one
or more boxes, stacked one above the other. Inside, each box
contains a series of movable frames of comb or foundation held in a
vertical position a bee space apart.
[0031] By "honey bee" is meant a Hymenopteran insect of the genus
Apis. The term "honey bee" is not limited to the adult form of the
insect, but encompasses all honey bee developmental stages,
including but not limited to egg, larva, and pupa. Exemplary honey
bee species include Apis mellifera and Apis cerana.
[0032] By "honey bee colony" is meant a community of bees. Honey
bee colonies may occur in the wild or may be maintained by bee
keepers.
[0033] By "honey bee parasitic mite" is meant any acarid that
parasitizes a honey bee or infests a honey bee hive. Exemplary
honey bee parasitic mites include Varroa mites and tracheal
mites.
[0034] By "hop derivative" is meant any molecule that naturally
occurs in hops (Humulus lupulus) and chemical derivatives thereof.
Hop derivatives (e.g., alpha acids, beta acids) may be purified
from hops or may be chemically synthesized.
[0035] By "infestation" is meant the colonization of a site or the
parasitization of an organism by a pest.
[0036] By "miticide" is meant an agent that inhibits a biological
function of a mite.
[0037] By "miticidal activity" is meant any activity that inhibits
the growth, reproduction, or survival of a mite or other
acarid.
[0038] By "preventing a mite infestation" is meant reducing the
success that a mite infestation will be established in an Apis
colony.
[0039] By "treating a mite infestation" is meant reducing,
stabilizing, or slowing the growth of a mite population in an Apis
colony.
BRIEF DESCRIPTION OF THE DRAWINGS
[0040] FIG. 1 is a graph showing Varroa mite and honey bee
mortality at four hours and twenty-two hours respectively in
response to hop product exposure at the indicated
concentrations.
[0041] FIG. 2 is a graph showing Varroa mite and honey bee
mortality four hours and at twenty-four hours respectively in
response to hop product exposure at the indicated
concentrations.
[0042] FIG. 3 is a graph showing Varroa mite and honey bee
mortality at one hour and twenty-four hours respectively in
response to hop product exposure at the indicated
concentrations.
[0043] FIG. 4 is a graph showing Varroa mite mortality at five
hours and adult honey bee mortality at twenty-four hours in
response to hop product exposure at the indicated concentrations.
MgBeta denotes magnesium salts of beta acids. RIAA denotes
rhoisoalpha acids.
[0044] FIG. 5 is a graph showing Varroa mite and honey bee
mortality at twenty-four hours in response to hop product exposure
at the indicated concentrations.
DETAILED DESCRIPTION OF THE INVENTION
[0045] The present invention is directed to methods and
compositions to control acarids and other related species of the
family Varroidae. The invention is based, in part, on the discovery
that naturally occurring components of hops are useful for the
prevention or treatment of a Varroa mite infestation.
Apis
[0046] Honey bees are insects that pass through four life stages:
the egg, larva, pupa and adult.
[0047] Adult bees belong to one of three castes: queen, worker, or
drone. The queen bee is the only female in the colony that is
capable of reproduction and is responsible for all egg production.
The worker bees are non-reproductive females who gather honey and
care for the queen's progeny, or "brood." The drones are male bees
that mate with the queen. The life cycle, from egg to adult bee,
takes twenty-one days for worker bees and twenty-four days for
drones. The queen bee lays each egg in a single cell of the comb.
The egg generally hatches into a larva on the fourth day, which
continues its development within the cell. On the ninth day the
cell with the developing larva is capped with wax and the larva
undergoes pupal metamorphosis. On day twenty-one, a new adult
worker bee emerges.
Acarids
[0048] Acarids are small parasitic arachnids that act as parasites
on a variety of plants and animals, including honey bees. Parasitic
mites that prey on honey bees include Varroa mites (e.g., Varroa
destructor, Varroa jacobsoni) and tracheal mites (e.g., Acarapis
woodi) Tracheal mites are microscopic mites that inhabit the
respiratory tubes of bees. Varroa mites are ectoparasites that feed
on bee hemolymph, and infest wild and domestic honey bee colonies.
Varroa mite reproduction begins when the adult female mite enters a
brood cell shortly before it is capped. Drone brood, which is
reared in larger cells than worker brood, is preferentially
targeted for mite infestation. The female mite feeds on the larval
hemolymph prior to depositing her eggs. The Varroa eggs enclose
under the sealed cell, and the developing mites feed on the bee
pupa. The first egg laid by the female Varroa develops into a male.
Subsequent eggs develop into females that mate with their brother.
The mated female mites along with their mother are released from
the capped cell when the bee emerges. The female mites typically
attach to adult bees between the abdominal segments or between body
regions, where they feed on the bees' hemolymph. Adult bees serve
as intermediate hosts and as a means of transport to new sites of
infestation.
[0049] Desirably, miticides used in acarid control should address
the following four needs: i) should disrupt a physiological
function required for mite survival; ii) should cause no adult bee
mortality; iii) should have no adverse effects on human bee keepers
or honey intended for human consumption; and iv) should be capable
of delivery into the hive.
Mite Control
[0050] Products used to control honey bee parasitic mite
infestation reduce, stabilize, or slow the growth of a mite
population in a hive or inhibit the growth, survival, reproduction,
or other biological function of a honey bee parasitic mite.
Preferably, the miticide kills the mite. Methods for measuring
parasitic mite infestation are known in the art. A number of
parameters can be indicative of the level of infestation present in
a bee colony: the number of mites present in a sample of bees from
an infested hive can be used as one measure of the level of
infestation present in the hive; bees reared in a hive having an
active infestation are on average smaller than bees reared in a
hive without infestation; thus, bee size or weight can be used as
another measure of infestation; the amount of honey produced in an
infected hive may be less than that produced in a healthy hive;
accordingly, honey production could serve as yet another measure of
the level of infestation; and finally, severe infestations result
in complete loss of colonies. Thus, loss of colonies can be a
measure of the level of infestation present in the hive. In one
embodiment, a miticide of the invention reduces the level of
infestation in a hive by at least 10%, 25%, 50%, 75% or even by
100%. In another embodiment, a miticide of the invention induces at
least 50%, 60%, or 70% mite lethality. Preferably, the miticide
induces 75%, 80%, 90%, or even 95% or 100% mite lethality.
Screening methods are used to identify concentrations of hop
derivatives that will be lethal to a mite (e.g., induce at least
70% mite lethality) while minimizing lethal effects on adult
bees.
[0051] Alternatively, a miticide of the invention inhibits mite
reproduction. Preferably, the miticide reduces mite reproduction by
at least 25%, 50%, 75% or 100%. In another approach, the miticide
disrupts a biological function required for acarid locomotion; such
treatment allows the mite to be trapped, drowned, isolated, or
otherwise removed from an area.
Miticide Screening
[0052] Commercial products that are currently being used to control
mite infestation can be lethal to adult bees when administered at
high concentrations, can have adverse effects on human bee keepers,
and may contaminate honey intended for human consumption.
Conventional miticides include Tau-Fluvalinate (a
synthetic-pyrethroid compound used as a selective contact and
stomach poison) and Coumaphos (a systemic organic phosphate) used
on animals to control lice, ticks and mites. In contrast to
conventional miticides, compositions of the invention contain safe
natural products derived from hops. Hops have been used for
centuries to flavor beer; thus, formulations comprising hop
derivatives are generally safe. Miticidal compositions of the
invention will not adversely affect human bee keepers or honey
intended for human consumption.
[0053] Miticides of the invention contain concentrations of hop
derivatives that have few or no adverse effects on honey bees
during any of their life stages, but are effective in killing or
disrupting the biological functioning of a mite. As reported
herein, beta acids, a hop derivative, delivered at 4% concentration
killed 87% of exposed mites after four hours while causing 0%
lethality in adult bees. In one approach, mites are exposed to
varying concentrations of hop derivatives to identify those
concentrations that kill 50% to 100% of the exposed mite. Adult
honey bees are then exposed to concentrations of hop derivatives
having miticidal activity to identify those that have a minimal
effect on honey bee survival. Preferably, at least 75%, 80%, 85%,
90%, 95%, or 100% of adult bees will survive following exposure to
a miticidal composition. In a similar approach, the effect of hop
derivatives on mite and honey bee reproduction is assessed.
Screening assays are used to determine the concentration of a
miticide that reduces the number of eggs laid by the female mite,
reduces the number of eggs that hatch, or reduces the number of
mites that grow to reproductive maturity; preferably, the reduction
is by at least 25%, 50%, 75%, 85%, 95% or 100%.
Hop Derivatives
[0054] A hop derivative is a compound that occurs naturally in a
hop plant (Humulus lupulus) or is chemically derived (either
through natural biosynthetic processes (e.g., living organism
metabolism (e.g., mammal, plant, bacteria)) or by synthetic
processes using human intervention (e.g., chemical synthesis).
Compositions of the invention include one or more compounds derived
from hops. Of particular interest are the hop acids. Hops contain
two major organic acid classes, alpha acids and beta acids. Hop
acids are the bitter acid components of hops that are used in beer
making. There are three major analogs for alpha acids, humulone,
cohumulone, and adhumulone, and three major analogs for beta acids,
lupulone, colupulone, and adlupulone. The percentages of the
analogs present in the alpha acids and beta acids are
variety-dependent. Thus, hop derivatives and hop products typically
contain one or a mixture of these analogs. The percentage of analog
present is dependent on the hop variety used to produce the
derivative or product. Alpha acids and beta acids can be prepared
by purification from natural hops and also by chemical synthesis
according to traditional methods. Exemplary hop derivatives include
beta acids, hexahydrobeta acids, rhoisoalpha acids, isoalpha acids,
tetrahydroisoalpha acids, hexahydroisoalpha acids, magnesium salts
of rhoisoalpha acids and magnesium salts of beta acids.
Compositions comprising hop derivatives are also available
commercially. John I. Haas, Inc. products containing hop
derivatives include Betacide, Redihop.RTM., Isohop.RTM., Tetrahop
Gold.RTM., Hexahop Gold.RTM., MgRIAA and MgBeta. The active
ingredients in these products are beta acids, rhoisoalpha acids
(RIAA), isoalpha acids (IAA), tetrahydroisoalpha acids (THIAA),
hexahydroisoalpha acids (HHIAA), magnesium salts of rhoisoalpha
acids (MgRIAA) and magnesium salts of beta acids (MgBeta),
respectively. For convenience, the identities of these products are
also listed in Table 1. These products and/or hop derivatives are
typically diluted to a desired concentration for use in the methods
of the invention.
[0055] Plant extracts are often used for the purification of
compounds from plants (e.g., hops). An extract can be prepared by
drying and subsequently cutting or grinding the dried material. The
term "extract" refers to a concentrated preparation of the
essential constituents of a plant, such as hops. Typically, an
extract is prepared by drying and powderizing the plant.
Optionally, the plant, the dried plant or the powderized plant may
be boiled in solution. The extract may be used in liquid form, or
it may be mixed with other liquid or solid herbal extracts.
Alternatively, the extract may be obtained by further precipitating
solid extracts from the liquid form. The extraction process may
then be performed with the help of an appropriate choice of
solvent, typically ethanol/water mixture, methanol, butanol,
iso-butanol, acetone, hexane, petroleum ether or other organic
solvents by means of maceration, percolation, repercolation,
counter-current extraction, turbo-extraction, or by carbon-dioxide
supercritical (temperature/pressure) extraction. The extract may
then be further evaporated and thus concentrated to yield by means
of air drying, spray drying, vacuum oven drying, fluid-bed drying
or freeze-drying, the extract product.
[0056] Crude extracts are tested for miticidal activity as
described herein. Further fractionation of a positive lead extract
having miticidal activity is necessary to isolate chemical
constituents responsible for the observed effect. Thus, the goal of
the extraction, fractionation, and purification process is the
careful characterization and identification of a chemical entity
within the crude extract that disrupts a mite biological function.
Methods of fractionation and purification of such heterogeneous
extracts are known in the art. If desired, compounds shown to be
useful as miticides are chemically modified according to methods
known in the art.
[0057] Numerous methods are available for the chemical synthesis of
candidate compounds. Such compounds can be synthesized from readily
available starting materials using standard synthetic techniques
and methodologies known to those of ordinary skill in the art.
Synthetic chemistry transformations and protecting group
methodologies (protection and deprotection) useful in synthesizing
the compounds identified by the methods described herein are known
in the art and include, for example, those such as described in R.
Larock, Comprehensive Organic Transformations, VCH Publishers
(1989); T. W. Greene and P. G. M. Wuts, Protective Groups in
Organic Synthesis, 2nd ed., John Wiley and Sons (1991); L. Fieser
and M. Fieser, Fieser and Fieser's Reagents for Organic Synthesis,
John Wiley and Sons (1994); L. Paquette, ed., Encyclopedia of
Reagents for Organic Synthesis, John Wiley and Sons (1995); and M.
Verzele and D. De Keukeleire, Chemistry and Analysis of Hop and
Beer Bitter Acids, Elsevier: Amsterdam (1991). Chemically
synthesized alpha and beta acids can be separated from a reaction
mixture and further purified by a method such as column
chromatography, high pressure liquid chromatography, or
recrystallization. As can be appreciated by the skilled artisan,
further methods of synthesizing the compounds herein will be
evident to those of ordinary skill in the art. Additionally, the
various synthetic steps may be performed in an alternate sequence
or order to give the desired compounds.
[0058] The compounds of this invention may contain one or more
asymmetric centers and thus occur as racemates and racemic
mixtures, single enantiomers, individual diastereomers and
diastereomeric mixtures. All such isomeric forms of these compounds
are expressly included in the present invention. The compounds of
this invention may also be represented in multiple tautomeric
forms, in such instances, the invention expressly includes all
tautomeric forms of the compounds described herein. All such
isomeric forms of such compounds are expressly included in the
present invention. All crystal forms of the compounds described
herein are expressly included in the present invention. As used
herein, the compounds of this invention, including the compounds of
formulae described herein, are defined to include derivatives.
Derivatives include compounds of the invention that are modified by
appending appropriate functionalities to enhance desired
properties.
[0059] Acceptable salts of the compounds of this invention include
those derived from acceptable inorganic and organic acids and
bases. Examples of suitable acid salts include acetate, adipate,
alginate, aspartate, benzoate, benzenesulfonate, bisulfate,
butyrate, citrate, camphorate, camphorsulfonate, digluconate,
dodecylsulfate, ethanesulfonate, formate, fumarate,
glucoheptanoate, glycolate, hemisulfate, heptanoate, hexanoate,
hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate,
lactate, maleate, malonate, methanesulfonate,
2-naphthalenesulfonate, nicotinate, nitrate, palmoate, pectinate,
persulfate, 3-phenylpropionate, phosphate, picrate, pivalate,
propionate, salicylate, succinate, sulfate, tartrate, thiocyanate,
tosylate and undecanoate. Other acids, such as oxalic acid, may be
employed in the preparation of salts useful as intermediates in
obtaining the compounds of the invention and their acceptable acid
addition salts. Salts derived from appropriate bases include alkali
metal (e.g., sodium), alkaline earth metal (e.g., magnesium),
ammonium and N-(alkyl).sub.4.sup.+ salts. This invention also
envisions the quaternization of any basic nitrogen-containing
groups of the compounds disclosed herein. Water or oil-soluble or
dispersible products may be obtained by such quaternization.
[0060] In some embodiments, miticidal compositions of the invention
include water soluble hop acid alkali metal salts (e.g., sodium,
potassium, lithium salts) and water insoluble hop acid alkaline
earth metal salts (e.g., calcium, magnesium) having increased
stability. These hop acid alkali metal salt (e.g., sodium,
potassium, lithium salts) and water insoluble hop acid alkaline
earth metal salt (e.g., calcium, magnesium) compositions are
advantageously stable relative to hop acids produced by
conventional methods, which are susceptible to degradation due to
heat, light, and acid catalysis. Compositions of the invention
remain stable under conditions that induce the degradation of other
conventional hop acids. In particular, after 6 months to 1 year of
storage, the compositions of the invention are expected to retain
at least about 50%, 60%, 75%, 80%, or preferably at least about
90%, 95% or even 100% of the hop acids present at the time of
application. Surprisingly, hop B acid crystals are also resistant
to degradation and exhibit increased stability. Accordingly, hop B
acid crystals are also useful in the compositions and methods of
the invention.
[0061] Water soluble hop acid alkali metal salts (e.g., sodium,
potassium, lithium salts) and water insoluble hop acid alkaline
earth metal salts (e.g., calcium, magnesium) are typically present
in a diluent or carrier at levels ranging from about 0.1% to about
95%. The methods herein contemplate administration of an effective
amount of compound or compound composition to achieve the desired
or stated miticidal effect. Preferably, the amount of active
ingredient (e.g., hop acid alkali metal salts, hop acid alkaline
earth metal salts or combinations thereof) are combined with
carrier materials (e.g., maltodextrin, cluster dextrin, corn
starch, corn syrup solids, glucose, cyclodextrin, arabic gum,
calaginan, inuline, partially hydrogenated soybean oil, cellulose,
hydroxymethyl cellulose, hydroxyethyl cellulose, hydroxypropyl
cellulose, rosin, hypomellose) to form a powder suitable for
delivery. For some applications, miticides of the invention are
formulated as liquids using diluents (e.g., sucrose or glucose
solutions, water, juices, other aqueous solutions, water miscible
solvents (ethanol, cremophor, dimethylsulfoxide (DMSO),
dimethylformamide (DMF), isopropanol (IPA) or glycerol, and other
solvents)) to form a solution or slurry.
[0062] A typical miticidal preparation will contain from about 1%
to about 95% hop acid, where the bottom of the range is any integer
between 5 and 94 and the top of the range is any integer between 6
and 95, where the hop acids are provided in a carrier (e.g.,
maltodextrin, cluster dextrin, corn starch, corn syrup solids,
glucose, cyclodextrin, arabic gum, calaginan, inuline, rosin,
partially hydrogenated soybean oil, cellulose, hydroxymethyl
cellulose, hydroxyethyl cellulose, hydroxypropyl cellulose,
hypomellose) that is suitable for use in methods of producing a
product having miticidal activity. Where non-aqueous miticidal
compositions are desired, the miticidal of the invention are
preferably formulated with rosin or partially hydrogenated soybean
oil. Such compositions may be used for the slow release of the
active miticidal composition, for example, in an aqueous slurry. In
still other embodiments, miticidal compositions of the invention
are dispersed in cellulose powder. In each of the aforementioned
embodiments, the hop acid alkali metal (e.g., sodium, potassium,
lithium), alkaline earth metal salts (e.g., calcium, magnesium), or
other hop acid salts are dispersed or dissolved in water, ethanol,
or another diluent together with any one or more of maltodextrin,
cluster dextrin, corn starch, corn syrup solids, glucose,
cyclodextrin, arabic gum, calaginan, inuline, rosin, partially
hydrogenated soybean oil, cellulose, hydroxymethyl cellulose,
hydroxyethyl cellulose, hydroxypropyl cellulose, and hypomellose.
The composition is then spray dried to facilitate the formation of
particles less than 1 mm in size. Preferably, the conditions used
for spray drying are adjusted such that the particles are at least
about 1 .mu.m, 5 .mu.m, 10 .mu.m, 25 .mu.m, 50 .mu.m, 75 .mu.m, 100
.mu.m, 150 .mu.m, 200 .mu.m, 500 .mu.m, 1 mm, 2 mm, or 5 mm in
size. The ratio of hop acids to carrier ranges between about 1:2
and 1:100. Preferred ratios include 1:2, 1:3, 1:4, 1:5, 1:6, 1:7,
1:8, 1:9, 1:10, 1:20, 1:30, 1:50, 1:75, and 1:100. Alternatively,
compositions of the invention include at least about 1%, 10%, 20%,
30%, 50%, 60%, 75%, 80%, 90%, or 95% hop acid alkali metal (e.g.,
sodium, potassium, lithium) or hop acid alkaline earth metal salts
(e.g., calcium, magnesium) in a diluent or carrier. Not all of the
hop acids need be in the metal form. Anywhere between 5% and 100%
of the hop acids present in the composition are in the metal form
at any given time, and between 95% and 0% are present as free
acids. In various embodiments, a composition of the invention
contains hop acids where 90% are present in the metal form and 10%
are present in the acid form; 50% are present in the metal form and
50% in the acid form; and 10% are present in the metal form and 90%
in the acid form.
[0063] In preferred embodiments, the preparation includes between 1
and 95% (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 25%, 75%, 80%, 90%,
95%) hop acids in a carrier or diluent. Alternatively, such
preparations contain from about 20% to about 80% hop acids.
Compositions containing alpha or beta acids are manufactured by
ordinary methods. Hop acids suitable for addition to products can
be formulated as ordinary tablets, capsules, solids, liquids,
emulsions, slurries, fine granules or powders, which are suitable
for administration to products during their preparation, following
preparation but prior to storage, or at any time prior to their
sale to a vendor or consumer. Lower or higher amounts than those
recited above may be required. The compositions delineated herein
include the compounds of the formulae delineated herein, as well as
additional miticidal agents if present, in amounts effective for
inhibiting mite growth or survival. Miticidal compositions of the
invention may be used in virtually any application where the
inhibition of a mite is desired. For example, compositions of the
invention are used to prevent, reduce, inhibit, slow or stabilize
the growth, proliferation, or survival of a mite.
[0064] Lower or higher doses than those recited herein may be
required to effectively kill mites without adversely affecting
honey bees. Specific dosage and treatment regimens are determined
empirically as described herein. Compositions of the invention are
also useful for preventing the establishment of an acarid
infestation, for treating an established acarid infestation, and
for maintaining the health of a hive previously treated for an
acarid infestation.
Formulations
[0065] Hop derivatives can be provided to bees or bee hives in a
number of convenient formulations. In general, strategies for
dispersing a therapeutic or prophylactic agent within the hive rely
on i) providing the agent in a food source (e.g., a liquid or solid
food); ii) providing the agent in a composition that will induce
hygienic behavior designed to remove the composition from the
colony (a packet designed to be torn apart by the bees); or iii)
providing the agent in a form that the bees will distribute
throughout the colony (e.g., a tracking powder provided at an
entrance to the hive). Formulations of the invention are used to
target mites on the body of adult bees, in the brood cell, or in
the hive. Desirably, the composition of the invention is active in
the hive for at least forty-one days. This provides for the
presence of the miticide for the entirety of the mite life cycle,
which typically is completed over the course of twenty-one to
thirty days. Where activity is maintained for a shorter period
(e.g., seven, fourteen, twenty-one, or thirty days), repeated
administration of a composition of the invention may be desired or
required. Compositions that are active for longer periods (e.g.,
two, three, six, nine, or twelve months) are also envisioned. Such
compositions may be used for the long-term treatment or prevention
of a mite infestation.
Powdered Formulations
[0066] Current miticides are introduced into the beehive on plastic
non-biodegradable strips that are about 1'' wide, 9'' long and
1/4'' thick. Similar means could be used for the delivery of hop
derivatives. Other strip compositions include, but are not limited
to, membranes, paper, plastic, and polymer strips. In one
embodiment, a composition comprising a hop derivative is provided
in a powdered formulation. A substrate material is coated with a
powdered formulation of hop acids, and the coating is subsequently
encased in a layer of a substance that is attractive to bees, such
as powdered sugar. This strip is placed inside the beehive where
the adult bees chew into the powdered sugar and expose the powdered
hop acids. The powdered hop acids get onto the body of the adult
bees, thereby contacting mites present on the adult bees and
causing the mites to die. Alternatively, the hop acids are consumed
by the bees and enter their hemolymph, where they are subsequently
consumed by the mites, thereby causing the mites to die.
[0067] In another approach, the powdered mixture is delivered to
the hive within a semi-permeable pouch that resembles a "teabag".
To rid the hive of this foreign object, the bees rip up the pouch,
thereby releasing the powder. The powdered hop acids get onto the
body of the adult bees and are distributed throughout the hive,
thereby killing (or otherwise interfering with mite proliferation
or survival) mites present on the bees and inhibiting the mite
infestation.
[0068] Encapsulated Formulations
[0069] In one approach, a hop derivative is provided in an
encapsulated formulation (liquid or powder). Preferably, a hop
derivative in liquid or powder form is encapsulated in a coating
that breaks down slowly inside the beehive. The coating provides
for the long-term release of the hop derivative. Preferably, the
composition is released over the course of two to six weeks (e.g.,
two, three, four, five, six weeks). Specific materials suitable for
use in capsule materials include, but are not limited to, porous
particulates or substrates such as silica, perlite, talc, clay,
pyrophyllite, diatomaceous earth, gelatin and gels, polymers (e.g.,
polyurea, polyurethane, polyamide, polyester, etc.), polymeric
particles, or cellulose. These include, for example, hollow fibers,
hollow tubes or tubing which release a hop derivative or other
compound specified above through the walls, capillary tubing which
releases the compound out of an opening in the tubing, polymeric
blocks of different shapes, e.g., strips, blocks, tablets, discs,
which release the compound out of the polymer matrix, membrane
systems which hold the compound within an impermeable container and
release it through a measured permeable membrane, and combinations
of the foregoing. Examples of such dispensing compositions are
polymer laminates, polyvinyl chloride pellets, and
microcapillaries. Encapsulation methods suitable for use in
apiculture are described, for example, by Rieth et al., Journal of
Apiculture Research 25(2):78-84 (1986).
[0070] Encapsulation processes are typically classified as chemical
or mechanical. Examples of chemical processes for encapsulation
include, but are not limited to, complex coacervation,
polymer-polymer incompatibility, interfacial polymerization in
liquid media, in situ polymerization, in-liquid drying, thermal and
ionic gelation in liquid media, desolvation in liquid media,
starch-based chemistry processes, trapping in cyclodextrins, and
formation of liposomes. Examples of mechanical processes for
encapsulation include, but are not limited to, spray drying, spray
chilling, fluidized bed, electrostatic deposition, centrifugal
extrusion, spinning disk or rotational suspension separation,
annular-jet encapsulation, polymerization at liquid-gas or
solid-gas interface, solvent evaporation, pressure extrusion or
spraying into solvent extraction bath.
[0071] Microcapsules are also suitable for the long-term release of
miticides. Microcapsules are small particles that contain a core
material or active ingredient surrounded by a coating or shell. The
size of the microcapsule typically varies from 1 to 1000 microns
with capsules smaller than 1 micron classified as nanocapsules and
capsules larger than 1000 microns as macrocapsules. Core payload
usually varies from 0.1 to 98 weight percent. Microcapsules can
have a variety of structures (continuous core/shell, multinuclear,
or monolithic) and have irregular or geometric shapes.
[0072] In another approach, the hop derivative is provided in an
oil-based delivery system. The oil-hop derivative mix is deposited
on a solid substrate and the substrate containing the hop
derivative is placed into the hive where it subsequently contacts
and kills the mites. Oil release substrates include vegetable
and/or mineral oils. In one embodiment, the substrate also contains
a surface active agent that renders the composition readily
dispersable in water; such agents include wetting agents,
emulsifying agents, dispersing agents, and the like.
[0073] Miticides of the invention can also be provided as
emulsions. Emulsion formulations can be found as water in oil (w/o)
or oil in water (o/w). Droplet size can vary from the nanometer
scale (colloidal dispersion) to several hundred microns. A variety
of surfactants and thickeners are usually incorporated in the
formulation to modify the size of the droplets, stabilize the
emulsion, and modify the release.
[0074] Alternatively, miticides of the invention may also be
formulated in a solid tablet and comprise (and preferably consist
essentially of) an oil, a protein/carbohydrate material (preferably
vegetable based), a sweetener and an active ingredient useful in
the prevention or treatment of a parasitic infection in a honey
bee. Methods for making such compositions are known in the art and
are described, for example, in U.S. Patent Publication No.
20060008492. In one embodiment the invention provides a solid
tablet and comprises (and preferably consist essentially of) an
oil, a protein/carbohydrate material (preferably vegetable based),
a sweetener and an active ingredient (e.g., hops .alpha. and/or
.beta. acid, or combinations or derivatives thereof) useful in the
prevention or treatment of a mite infestation. Tablets typically
contain about 4-40% (e.g., 5%, 10%, 20%, 30%, 40%) by weight of an
oil (e.g., plant oil, such as corn, sunflower, peanut, olive, grape
seed, tung, turnip, soybean, cotton seed, walnut, palm, castor,
earth almond, hazelnut, avocado, sesame, croton tiglium, cacao,
linseed, rape-seed, and canola oils and their hydrogenated
derivatives; petroleum derived oils (e.g., parafins and petroleum
jelly), and other water immiscible hydrocarbons (e.g., parafins).
The tablets further contain from about 5-40% (e.g., 5%, 10%, 20%,
30%, 40%) by weight of a vegetable-based protein/carbohydrate
material. The material contains both a carbohydrate portion (e.g.,
derived from cereal grains, such as wheat, rye, barley, oat, corn,
rice, millet, sorghum, birdseed, buckwheat, alfalfa, mielga, corn
meal, soybean meal, grain flour, wheat middlings, wheat bran, corn
gluten meal, algae meal, dried yeast, beans, rice) and a protein
portion. While the relative fraction of each portion making up the
material may vary, the material should include at least a portion
of carbohydrate and protein.
[0075] The tablets also contain between about 10-75% (10, 15, 20,
25, 50, 75%) by weight of a sweetener. As used herein, the term
"sweetener" generally refers to both natural and artificial
sweeteners. Preferably, the sweetener is a sugar such as glucose,
fructose, sucrose, galactose, lactose, and reversed sugar. The
sugar is preferably selected from the group consisting of
granulated sugar (white sugar), brown sugar, confectioner's sugar,
impalpable sugar, icing sugar, and combinations thereof. Alcohols
such as glycerin and complex carbohydrates, such as starches may
also be used as the "sweetener" ingredient. The sweetener is used
primarily as an attractant for the insects, however the sweetener
also helps to impart a granular structure to the tablets,
especially when the sweetener is a sugar. As previously discussed,
this granular structure permits the tablet to crumble over time
upon the exertion of sufficient forces.
[0076] Optionally, various excipients and binders can be used in
order to assist with delivery of the active ingredient or to
provide the appropriate structure to the tablet. Preferred
excipients and binders include anhydrous lactose, microcrystalline
cellulose, corn starch, magnesium estearate, calcium estearate,
zinc estearate, sodic carboxymethylcellulose, ethyl cellulose,
hydroxypropyl methyl cellulose, and mixtures thereof.
[0077] Tablets according to the present invention are manufactured
by mixing all of the ingredients together and then compressing the
mixture into a tablet of desired shape and size for a particular
application. Preferably, the tablet is discoid in shape with a
diameter of between about 2-5 inches and a thickness of from about
0.5-2 inches. The pressing may be accomplished by a manual or
automatic pressing device. The pressure exerted on the mixture
should be sufficient so as to form the tablet into a
self-sustaining body.
[0078] Methods of delivering an active ingredient to an insect
according to the present invention comprise the steps of providing
a solid tablet containing the active ingredient as previously
described and placing the tablet in a location where the insect may
come into direct contact therewith. In treating honeybees that are
generally colonized in a manufactured bee hive, the tablet is
preferably placed inside the hive.
[0079] Over the next several weeks after the tablet is placed into
the hive, the bees chew and crumble the tablet exposing the active
ingredient to the other bees. The crumbs fall through the brood box
away from the honey supers. Preferably, the entire tablet is
disintegrated in about 30-45 days.
[0080] Miticides of the invention can also be delivered in the form
of syrups that are attractive to bees and induce feeding behavior.
The syrups for use in the invention preferably comprise sugar and
water. Particularly preferred are 50% w/v sucrose solutions. A
liquid composition is formed by dispersing hops acids in a sugar
syrup comprising 50% sucrose in water. The composition is used as a
feed supplement for the bees and can be placed at a suitable
location in or near a hive.
[0081] Miticides of the invention can also be delivered in packets
suitable for inducing hygienic behavior in bees. Such packets are
prepared by enclosing a fine powder of hops acids and sugar in a
porous material capable of being torn apart by bees. Preferably,
the porous material is made of waxed paper or filter paper.
Suitable filter papers include those comprising abaca fibers, wood
pulp and cellulose rayon fibers. If desired, the paper is coated
with polyethylene mixed with copolymers, polypropylene mixed with
copolymers or 100% polypropylene.
[0082] In other embodiments, miticides are prepared in a dusting
composition or as a powder. Dusting compositions are typically
prepared by grinding sugar to a fine powder and mixing it into the
powder hops acids. Alternatively, the dusting compositions are
prepared as described in Example 3 for maltodextrin, where the
powder is obtained by spray drying. The skilled artisan adjusts the
conditions used in the spray drying process to achieve particles or
granules of a size that facilitates delivery to the bees.
Desirably, the powder comprises fine particles that coat the bee
and all of its body parts (e.g., joints, groove, bristles). The
dusting composition can be applied directly to the top of the bee
frames, to the combs within the hive, or to the interior surfaces
of the hive, or may be applied directly to a bee cluster.
[0083] Alternatively, the miticides are prepared in a liquid spray
composition that is formed by dispersing hops acids in any suitable
liquid. Preferably, the hops acids are dispersed in water. If
desired, the spray composition also includes a surfactant that
allows the spray to be dispersed efficiently without clogging the
spraying apparatus. The composition can be used to spray the hive
interior, or the comb, or can be used to spray bee clusters
directly.
[0084] In another approach, miticides of the invention are
delivered in the form of a vapor. Methods for delivering such
vapors to a hive are described, for example, in U.S. Patent
Publication No. 20020151249.
Miticide Delivery
[0085] Devices for delivering pest control agents to bees or to a
bee hive are known in the art. Such delivery devices include
strips, controlled release strips, tablets, reservoirs, polymer
discs, trays, and evaporation devices. If desired, the delivery
device is provided in a biodegradable form. In particular, devices
suitable for delivering a composition of the invention to a
parasitic mite, to a honey bee, or to a honey bee hive are
described, for example, in U.S. Patent Publication Nos.
20070059333; 20070026765; 20060141904; 20060009122; 20060008492;
20050095954; 20050090560; 20050048093; 20040229542; 20040077291;
20030190860; 20030044443; 20030027490; 20020182977; 20020151249;
20020094756; 20010014346 and 20020151249. Dispensing means and
suitable compositions for controlled release are described in U.S.
Pat. Nos. 6,843,985; 5,750,129; 4,775,534; 5,849,317; 5,348,511;
6,037,374; 7,137,864; 6,837,770; 6,820,773; 6,702,645; 6,646,014;
6,620,025; 6,595,828; 6,585,557, 6,475,061, 6,468,129; 6,277,371;
6,221,375; 6,204,283; 6,096,350; 6,037,374; 6,010,390; 5,312,622;
5,230,894; 5,227,162; 5,135,758; 5,070,091; 5,069,651; 5,023,359;
4,876,265; 4,867,731; 4,837,216; 4,682,380; and 4,299,816, which
are incorporated herein by reference in their entirety.
Kits
[0086] The invention provides kits for the treatment or prevention
of an acarid infestation. In one embodiment, the kit includes a
composition containing an effective amount of a hop derivative in a
form suitable for delivery to a site of infestation (e.g., bee
hive). In some embodiments, the kit comprises a container which
contains a miticide; such containers can be boxes, ampules,
bottles, vials, tubes, bags, pouches, blister-packs, or other
suitable container forms known in the art. Such containers can be
made of plastic, glass, laminated paper, metal foil, or other
materials suitable for holding miticides.
[0087] If desired the miticide of the invention is provided
together with instructions for administering it to a site of
infestation. The instructions will generally include information
about the use of the composition for the treatment or prevention of
an acarid infestation. In other embodiments, the instructions
include at least one of the following: description of the miticide;
dosage schedule and administration for treatment or prevention of a
miticide infestation; precautions; warnings; description of
research studies; and/or references. The instructions may be
printed directly on the container (when present), or as a label
applied to the container, or as a separate sheet, pamphlet, card,
or folder supplied in or with the container.
EXAMPLES
Example 1
Hop Beta and Alpha Acids Used in Miticide Screening
[0088] Beta acids, alpha acids, and a combination of beta and alpha
acids were screened for efficacy as miticides. Liquid test products
containing beta acids were provided in a Betastab 10A.RTM.
formulation (10% beta acids) hereinafter called "Betacide". Liquid
test products containing alpha acids were provided in a
Redihop.RTM. formulation (30% rhoisoalpha acids), Isohop.RTM.
formulation (30% isoalpha acids), Tetrahop Gold.RTM. formulation
(9% tetrahydroisoalpha acids), Hexahop Gold.RTM. formulation (5%
hexahydroisoalpha acids and 5% tetrahydroisoalpha acids). A
combination of alpha and beta acids were prepared by mixing equal
parts Redihop.RTM. and Betacide. Powdered test products containing
beta acids were provided by a magnesium salt formulation of beta
acids. Powdered test products containing alpha acids were provided
by magnesium salt formulations of Redihop.RTM., Tetrahop Gold.RTM.
and Hexahop Gold.RTM..
[0089] Tests were carried out using the concentrations of beta,
alpha, or beta and alpha acid combinations indicated in Table
1.
[0090] Specifically, in Tests 1-4: 5% beta acids as Betacide test
solution, 15% rhoisoalpha acids as Redihop.RTM. test solution, and
a 2.5% beta acids/7.5% rhoisoalpha acids combination was used.
[0091] In Tests 5-8, 4% beta acids as Betacide test solution, 30%
rhoisoalpha acids concentration as Redihop.RTM. test solution, and
a 2% beta acids/15% rhoisoalpha acids combination were used.
[0092] In Tests 9-12, 4% beta acids as Betacide test solution, 30%
rhoisoalpha acids concentration as Redihop.RTM. test solution, and
a 2% beta acids/15% rhoisoalpha acids combination were used.
[0093] In Tests 13-15, 30% isoalpha acids as Isohop.RTM., 9%
tetrahydroisoalpha acids as Tetrahop Gold.RTM., and a combination
of 5% tetrahydroisoalpha acids and 5% hexahydroisoalpha acids from
Hexahop Gold.RTM. were used.
[0094] In Tests 16-19, 4.3% and 8.5% beta acids as a magnesium
salt, and 65.5% rhoisoalpha acids as a magnesium salt of
Redihop.RTM. were used.
[0095] In Tests 20-22, 25.3% tetrahydroisoalpha acids as a
magnesium salt of Tetrahop Gold.RTM., and a combination of 12.2%
each of tetrahydroisoalpha acids and hexahydroisoalpha acids from
magnesium salts of Hexahop Gold.RTM. were used.
Miticide Screening Assays
[0096] Tests using liquid hop products were conducted by absorbing
one milliliter of test solution onto a filter paper in a Petri
dish. Tests using the powdered hop products (magnesium salts) were
conducted by spreading 0.5 gm of test powder evenly over filter
paper in a Petri dish. Five to ten Varroa mites were then placed on
the treated filter paper and mite survival was determined at one,
four or five and twenty-four hours hour time points. Similar
methods were used to evaluate the effect of the test compounds on
adult honey bee survival. Adult honey bee survival was scored after
twenty-two hours exposure to test product. Five to ten adult honey
bees were placed in Petri dishes containing treated filter paper.
Filter paper treated with water (for liquid test solutions) or
cornstarch (for powdered test solutions) was used as a negative
control for tests with the mites and the adult honey bees. All
trials were replicated four times. Table 1 outlines the tests and
results of testing various hop products for miticidal activity.
TABLE-US-00001 TABLE 1 Hop Derivatives Efficacy Against Varroa
Mites of Honey Bees Test Figure Product Test acid % %
Mortality/Exposure Time Number Number Product Active Ingredient
(ai) Conc. % Alpha Beta Diluent Mites Hours Bees Hours 1 1
deionized water none NA NA NA none 7 4 0 22 2 1 Betacide beta acids
10 NA 5 deionized water 73 4 20 22 3 1 Redihop rhoisoalpha acids 30
15 NA deionized water 21 4 0 22 4 1 Redihop + Betacide as in test 2
+ test 3 30 + 10 7.5 2.5 deionized water 43 4 7 22 5 2 deionized
water none NA NA NA none 7 4 0 24 6 2 Betacide beta acids 10 NA 4
deionized water 87 4 0 24 7 2 Redihop rhoisoalpha acids 30 30 NA
deionized water 68 4 0 24 8 2 Redihop + Betacide as in test 2 +
test 3 30 + 10 15 2 deionized water 80 4 0 24 9 3 deionized water
none NA NA NA none 0 1 0 24 10 3 Betacide beta acids 10 NA 4
deionized water 20 1 ND 24 11 3 Redihop rhoisoalpha acids 30 30 NA
deionized water 13 1 ND 24 12 3 Redihop + Betacide as in test 2 +
test 3 30 + 10 15 2 deionized water 13 1 ND 24 13 3 Isohop isoalpha
acids 30 30 NA deionized water 70 1 33 24 14 3 Tetrahop Gold
tetrahydroisoalpha acids 9 9 NA deionized water 81 1 0 24 15 3
Hexahop Gold hexahydroisoalpha acids plus 5 5 NA deionized water
100 1 7 24 tetrahydroisoalpha acids 5 5 NA 16 4 corn starch none NA
NA NA none 13 5 ND ND 17 4 MgBeta magnesium salt of beta acids 59.5
NA 4.3 corn starch 38 5 ND ND 18 4 MgBeta magnesium salt of beta
acids 59.5 NA 8.5 corn starch 67 5 0 24 19 4 MgRIAA magnesium salt
of rhoisoalpha 65.5 65.5 NA corn starch 7 5 ND ND acids 20 5 corn
starch none NA NA NA none 17 24 0 24 21 5 MgTetrahop Gold mg salt
of tetrahydroisoalpha 75.8 25.3 NA corn starch 50 24 0 24 acids 22
5 MgHexahop Gold mg salt of hexahydroisoalpha 36.7 12.2 NA corn
starch 50 24 0 24 acids plus mg salt of 36.7 12.2 NA
tetrahydroisoalpha acids Notes: NA means Not Applicable, ND means
No Data
[0097] Results for the tests described in Table 1 are shown in
FIGS. 1-5.
[0098] In Tests 1-4 after five hours exposure, 5% beta acids killed
73% of Varroa mites; 15% rhoisoalpha acids killed 21% of Varroa
mites; and a combination of 2.5% beta acids/7.5% rhoisoalpha acids
produced 43% mortality of mites. Under control conditions only 7%
mite mortality was observed. The majority of adult bees survived
exposure to these same concentrations of alpha and beta acids.
Specifically, 100% adult bees survived exposure to rhoisoalpha
acids; 80% of adult bees survived exposure to 5% beta acids; and
93% of adult bees survived exposure to the combination of 2.5% beta
acids/7.5% alpha acids. These results are presented in Table 1 and
FIG. 1.
[0099] In Tests 5-8 following four hours of exposure, 4% beta acids
killed 87% of Varroa mites; 30% rhoisoalpha acids killed 68% of
mites; and the combination of 15% rhoisoalpha acids and 2% beta
acids killed 80% of mites. 7% mite mortality was observed under
control conditions. Adult bees exposed to these same product
concentrations for 24 hours showed 100% survival. These results are
presented in Table 1 and FIG. 2.
[0100] In Tests 9-15 after one hour of exposure, 4% beta acids
killed 20% of Varroa mites; 30% rhoisoalpha acids killed 13% of
mites; the combination of 15% rhoisoalpha acids and 2% beta acids
killed 13% of mites; 9% tetrahydroisoalpha acids killed 81% of
mites; the combination of 5% tetrahydroisoalpha acids and 5%
hexahydroisoalpha acids killed 100% of mites. No mite mortality was
observed under control conditions. Adult bees exposed to these
product concentrations for 24 hours showed 67% survival after
exposure to isoalpha acids; 93% survival after exposure to the
combination of 5% tetrahydroisoalpha acids and 5% hexahydroisoalpha
acids; 100% survival after exposure to 9% tetrahydroisoalpha acids;
and 100% survival after exposure to control conditions. These
results are presented in Table 1 and FIG. 3.
[0101] In Tests 16-19, after five hours of exposure, 8.5% beta
acids in the form of a magnesium salt killed 67% of Varroa mites;
65.45% rhoisoalpha acids in the form of a magnesium salt killed 7%
of Varroa mites. 13% of mites died under control conditions. 100%
of bees survived after 24 hours exposure to 8.5% beta acids as a
magnesium salt. These results are presented in Table 1 and FIG.
4.
[0102] In Tests 20-22 after 24 hours of exposure, 25.27%
tetrahydroisoalpha acids in the form of the magnesium salt killed
50% of Varroa mites; and a combination of 12.23% tetrahydroisoalpha
acids and 12.23% hexahydroisoalpha acids both in the form of
magnesium salt killed 50% of Varroa mites. 17% of mites died under
control conditions. 100% of adult honey bees survived for 24 hours
under the same conditions. These results are presented in Table 1
and FIG. 5.
Example 2
Miticidal Effect of Hop Acids in Simulated Apiary Assays
[0103] In the apiary, a bee brush was used to gently sweep bees
into a 1/2 cup measuring cup. To prevent them from flying, the side
of the bee brush was used as a lid over the cup. The 1/2 cup of
bees was transferred to a 1/2 pint or pint Mason jar with a
screened lid (i.e., a sugar shake jar). One jar containing a 1/2
cup of bees was used per container of treatment.
[0104] The containers were prepared by inserting a screen at the
opening of the container and placing a cut piece of sticky board in
the lid of the container. The lids were attached to the containers
and the containers were turned upside down so that from bottom to
top there was a lid, sticky board, and screen. A #5 rubber cork was
placed in one of the two feeding holes. One jar of bees was placed
briefly into the -20 freezer. The bees remained in the freezer
until bee movement slowed considerably (3 to 5 minutes). At this
point the bees were ready for painting using Bee
paint--Testors.RTM. or another similar brand. Bees were removed
from the freezer and dumped onto a metal tray. A small dot of paint
was deposited on the thorax of each bee using a small paint brush.
The painted bees were placed into the container by dropping them
through the feeding hole until all of the bees in the jar were
painted.
[0105] The desired amount of treatment was prepared and
administered by quickly placing the bees in the Mason jar together
with a mixture containing 2 grams of a 1:1 mixture of 52% magnesium
salt of hop beta acids (Mgbeta) and cornstarch or a corn starch
control. The bees in the jar were then rolled in the powder until
they were completely covered in powder. Bees were then transferred
to a new container where miticidal activity was assayed. During the
assay, the bees were fed a 1:1 solution of sucrose with hot water
and were maintained in an incubator at 32.5.degree. C. and 60%
humidity. Bee mortality was monitored daily. Feeders were
replenished by removing/refilling the vials and temporarily
replacing the rubber corks.
[0106] The treated bees were shaken in sugar shake to determine how
many mites remained on the bees after one week. This method
involves mixing 1/2 cup of bees with a few tablespoons of powdered
sugar and shaking the bees vigorously. The loose sugar and mites
are collected, and the mites are counted. The number of mites
present after the sugar shake is combined with the number of dead
mites that were found on the sticky boards to determine the Total
Number of mites per container (T1-T4). The mortality percentages
were calculated by dividing the number of dead mites on the sticky
boards by the total number of mites.
TABLE-US-00002 Treatment Group Mite Mortality T1 92.86% mite
mortality T2 83.33% T3 90.9% T4 (control) 42.86% mite mortality
The total number of mites in each treatment group, which included
approximately 80 bees, ranged from 7 to 14.
Example 3
Sodium and Magnesium Salts of Hop Acids Kill Mites
[0107] Tests using the powdered hop products (magnesium salts) were
conducted by spreading test powder evenly over filter paper in a
Petri dish. The MgBeta test powder contained a 1:1 mixture of 0.25
g of magnesium salt of beta acids (52% magnesium beta salts of hop
acids) and corn starch; thus, 26% magnesium salt of beta acids was
used in the assay. The NaBeta test powder contained a 1:1 mixture
of 0.25 g of sodium salt of beta acids (6.4% sodium salt of beta
acids); thus, 3.2% magnesium salt of beta acids was used. Varroa
mites were placed on the treated filter paper and mite survival was
determined at one, two, three, four or five hours following
exposure. Results are shown in Table 2.
TABLE-US-00003 TABLE 2 MITE BIOASSAY - 0.25 g Treatment Date #
Hours Exposure Treatment % Mortality Mar. 28, 2007 1 MgBeta 4.8
NaBeta 15.1 Cornstarch 0 Control 0 2 MgBeta 9.5 NaBeta 45.2
Cornstarch 0 Control 11.1 3 MgBeta 14.3 NaBeta 45.2 Cornstarch 0
Control 11.1 4 MgBeta 28.6 NaBeta 75.4 Cornstarch 0 Control 16.7 5
MgBeta 57.1 NaBeta 89.7 Cornstarch 0 Control 39.2
Example 4
Preparation of Water-Soluble Beta Acid Sodium Salts
[0108] Step 1: Commercially available hop CO.sub.2 extract (55%:
Alpha acids, 30% Beta acids, 10% uncharacterized residue) (10 kg)
is placed into Tank 1. CO.sub.2 extracts are produced by natural
carbon dioxide extraction of hops. Carbon dioxide is a natural
solvent that eliminates residual solvents that typically present in
hop extracts produced using hexane or ethylene chloride solvents.
Food grade KOH (100 g) is dissolved in deionized water (20 L). The
KOH solution is added into the Tank 1 and the mixture is stirred at
55-65.degree. C. for 1 hour and then agitation is stopped to form
two layers.
[0109] Step 2: The lower aqueous layer (15 L) is transferred into
Tank 2. The crude beta acid potassium salts are cooled down to room
temperature for two hours and then Celite.RTM.(diatomaceous earth)
is added (0.5% wt/wt mix) for 20-30 minutes. The resulting mixture
is filtered through a Buchner type filtration apparatus under
vacuum.
[0110] Step 3: The filtrate (10 L) is transferred to Tank 3 and
heated to 70.degree. C. with agitation and then acidified with 30%
of aqueous H.sub.2SO.sub.4 until the mixture reaches pH 2-3. The
agitation is stopped, and the mixture is allowed to form two
layers. The upper layer (5 L), which is retained, contains about
70% beta acids.
[0111] Step 4: Aqueous NaOH solution (about 9 L) is added to the
upper layer (3 L) and the pH is adjusted to pH 10-10.5 at
65.degree. C. with agitation then active charcoal (Norit A.RTM. 200
mesh) (2% wt/wt mix) is added to the solution, which is gently
stirred for thirty minutes. The mixture is incubated overnight and
then filtered. The filtrate is diluted with deionized water to
achieve 10% beta acid sodium salts in an aqueous composition.
Alternatively, the mixtures is passed over a column containing 60
mesh active charcoal.
Example 5
Preparation of Powder of Hop Beta Acid Sodium Salts
[0112] Hop beta acids are prepared as described in Example 4 with
the following modification. In step 4 of Example 4, aqueous
maltodextrin solution was prepared at pH 10 by mixing an aqueous
beta acid sodium salts solution with maltodextrin, such that the
hop acids to maltodextrin ration is 5:1 to 10:1 ratio after the
filtration. The solution is dried by spray drying to obtain a pale
yellow powder containing 5-10% beta acid sodium salts.
Example 6
Preparation of Hop Beta Acid Sodium Salts in 67% EtOH Solution
[0113] Beta acids are prepared as described in Example 4 with the
following modification. In step 4 of Example 4, 500 ml of the
aqueous solution, which contains about 30% beta acid sodium salts
is mixed with 1000 ml of 100% EtOH with stirring to form 67% pale
yellow ethanol solution containing 10% beta acid sodium salts.
Example 7
Preparation of Beta Acid Sodium Salts in 90% EtOH Solution
[0114] 500 ml of an aqueous solution containing about 30% beta acid
sodium salts is neutralized with 0.1 N H.sub.2SO.sub.4. The beta
acid-rich fraction is precipitated out at pH 7-9. The solid is
separated and washed with water three times. The solid (200 g) is
dissolved into 1700 ml of 100% EtOH under stiffing. 100 ml of
aqueous NaOH solution (16 g of NaOH and 84 ml of water) is added to
the EtOH solution under stirring to form a pale yellow clear 90%
ethanol solution that contains 10% beta acid sodium salt.
Example 8
Stability Study of Hop Acids
[0115] The following samples containing hop acids were incubated
under aerobic conditions at 75.degree. C. for 0-6 days. Liquid
samples were dissolved in a volume of 0.1 ml
[0116] Samples
[0117] 1. Beta acids rich hop extract (10%) in water (pH=5.0)
[0118] 2. Powder of 10% Beta acids and 90% Maltodextrin
[0119] 3. Powder of 5% Beta acid Na salts and 95% Maltodextrin
[0120] 4. Powder of 10% Beta acid Na salts and 90% Maltodextrin
[0121] 5. 10% beta acid Na salts in water (pH=10)
[0122] 6. Hop extract with 10% hop beta acid Mg salts
Following this incubation, the presence of hop acids was assayed.
After incubation of 20 mg of each sample at 75.degree. C. under
aerobic conditions, the sample was dissolved into 1 ml of 70%
aqueous EtOH. The solution was diluted 50 times with methanol and
then 20 uL of the diluted sample was injected into a high pressure
liquid chromatography (HPLC) for analysis. The HPLC conditions used
were: [0123] Temperature: 35.degree. C. [0124] Eluent A: 10 mM
Triethylammonium acetate/water [0125] Eluent B: 10 mM
Triethylammonium acetate/acetonitrile [0126] Gradient: Eluent
B=from 30% to 90% in 20 minutes then keep B=90% for 5 minutes
[0127] Detection: 370 nm for beta acids, 254 nm for other
degradation peaks [0128] Determination: Area under the curve of
three peaks (15-17 min) at 370 nm [0129] Authentic sample:
International Calibration Extract 2 from American Society of
Brewing Chemists [0130] HPLC Type: Agilent HP1100 series with diode
array detector.
[0131] Samples referred to in Table 3 are described above as
Samples 1-6. As shown in Table 3, alkali salts of beta acids
(sample 3, 4, and 5) were more stable than the neutral form of beta
acids in neutral or acidic conditions (1, 2 and 6). The results of
these studies are summarized in Table 3.
TABLE-US-00004 TABLE 3 Stability study of hop beta acids under
force conditions Beta acids remained (%) Period (Day) Sample 1 3 6
1 52 20 0 2 40 15 0 3 100 100 98 4 100 100 100 5 100 90 90 6 60 25
0
This method provides for the rapid assessment of the chemical
stability of hop acid salts relative to the degradation observed in
hop acids. The degradation observed after six days at 75.degree. C.
is equivalent to the degradation that would be expected if the hop
acids and salts were stored for 6 months at room temperature.
Example 9
Preparation of Rhoisoalpha Acid Salts
[0132] An inorganic salt of rhoisoalpha acids is produced using any
standard method known in the art. In one embodiment, a rhoisoalpha
acid is produced according to the following method.
[0133] An empty drum was placed on a scale and tared. To the drum
was added 80 kg of a mixture of rhoisoalpha acids (30%) in
deionized water (75 L) at room temperature. The mixture was
subjected to gentle agitation to form an aqueous slurry. MgSO.sub.4
(45 kg) was added to the slurry at one time and the agitation was
continued for 5-10 minutes until the MgSO.sub.4 was homogeneously
distributed. After 10 minutes, a small sample was removed to
determine whether the reaction had reached completion. This was
determined using an HPLC to assay the presence of rhoisoalpha acids
magnesium salt. When the reaction was complete, the mixture was
removed and deionized water was added to adjust the concentration
of rhoisoalpha acids magnesium salt to 15-17% having 83-85% water
content. The mixture was then dried using standard methods. When
the drying was completed, the flaky products were packed in
aluminum coated polyethelene bags, heat sealed and stored at room
temperature prior to analysis.
Example 10
Preparation of Rhoisoalpha Acids Calcium Salts
[0134] To prepare the calcium salt of rhoisoalpha acid, 300 grams
of an aqueous 30% rhoisoalpha acid solution having a pH of 11 was
mixed with 37 grams of CaCl.sub.2-2H.sub.2O, which had been mixed
previously with 200 mL deionized water. This slurry was mixed until
homogeneous. The slurry was then poured directly onto a drying tray
and dried.
Example 11
Preparation of Rhoisoalpha Acids Lithium Salts
[0135] To prepare the lithium salt of rhoisoalpha acid, 300 grams
of an aqueous 30% rhoisoalpha acid solution having a pH of 9 was
mixed with 21 grams of LiOH--H.sub.2O, which had been mixed
previously with 300 mL deionized water. This slurry was mixed until
homogeneous. The slurry was then filtered through a Buchner funnel
to remove excess water and placed onto a drying tray and dried.
Example 12
Preparation of Rhoisoalpha Acids Calcium Salts
[0136] To prepare the calcium salt of rhoisoalpha acid, 300 grams
of an aqueous 30% rhoisoalpha acid solution having a pH of 11 was
mixed with 37 grams of CaCl.sub.2-2H.sub.2O, which had been mixed
previously with 200 mL deionized water. This slurry was mixed until
homogeneous. The slurry was then poured directly onto a drying tray
and dried.
Example 13
Preparation of Rhoisoalpha Acids Potassium Salts
[0137] To prepare the potassium salt of rhoisoalpha acid, 300 grams
of an aqueous 30% rhoisoalpha acid solution having a pH of 10 was
mixed with 35 grams of K.sub.2CO.sub.3 which had been mixed
previously with 300 mL deionized water. This slurry was mixed until
homogeneous. The slurry was then poured directly onto a drying tray
and dried.
Example 14
Preparation of Tetrahydroisoalpha Acid Calcium Salts
[0138] To prepare the calcium salt of tetrahydroisoalpha acid, 1000
grams of an aqueous 9% tetrahydroisoalpha acid solution having a pH
of 10.5 was mixed with 42 grams of CaCl.sub.2-2H.sub.2O, which had
been mixed previously with 100 mL deionized water. This slurry was
mixed until homogeneous. The slurry was then filtered through a
Buchner funnel to remove excess water and placed onto a drying tray
and dried.
[0139] Virtually any hop acid alkali salt (e.g., sodium, potassium,
lithium), hop acid alkaline earth metal salt (e.g., magnesium,
calcium salts), or other hop acid salts may be used in the process
set forth above. As set forth in the above examples, the invention
provides processes for producing water soluble alkali salts or
water insoluble alkaline earth metal salts of alpha acids or beta
acids. Virtually any isoalpha acid, rhoisoalpha acid,
tetrahydroisoalpha acid, hexahydroisoalpha acid, beta acid,
hexahydrobeta acid, tetrahydrobeta acid, lupulone, colupulone,
adlupulone, or derivatives or combinations thereof may be used in
the processes of the invention. In one embodiment, the
concentration of hop acids present in the aqueous solution ranges
between 5% and 50%, inclusive. In other embodiments, the
concentration ranges between 5-45% (e.g., 9%, 10%, 15%, 20%, 25%,
30%, 35%, 40%, and 45%), inclusive. In yet other embodiments, the
lower end of the range is any number between 9 and 49%; and the
upper end of the range is any number between 10 and 50%. The hop
acids of step 4 may be dried to obtain salts any standard method or
combination of methods, including but not limited to, spray drying,
vacuum drying, drum drying, pan drying, window drying and freeze
drying. Preferably, spray drying is used.
[0140] Compounds of the invention are prepared in a manner
essentially as described above and in the general schemes. The
recitation of a listing of chemical groups in any definition of a
variable herein includes definitions of that variable as any single
group or combination of listed groups. The recitation of an
embodiment for a variable herein includes that embodiment as any
single embodiment or in combination with any other embodiments or
portions thereof. Another embodiment is a compound of any of the
formulae herein made by a process delineated herein, including the
processes exemplified in the schemes and examples herein. Another
aspect of the invention is a compound of any of the formulae herein
for use in as a miticide as delineated herein.
Other Embodiments
[0141] From the foregoing description, it will be apparent that
variations and modifications may be made to the invention described
herein to adopt it to various usages and conditions. Such
embodiments are also within the scope of the following claims.
[0142] The recitation of a listing of elements in any definition of
a variable herein includes definitions of that variable as any
single element or combination (or sub combination) of listed
elements. The recitation of an embodiment herein includes that
embodiment as any single embodiment or in combination with any
other embodiments or portions thereof.
[0143] All patents and publications mentioned in this specification
are herein incorporated by reference to the same extent as if each
independent patent and publication was specifically and
individually indicated to be incorporated by reference.
* * * * *