Searching for Chemical Signals in Orang-utans (Pongo spp.)

2016 Orangutan SSP Husbandry Workshop: Celebrating a Decade of Care and Conservation

Tom Goodwin (goodwin@hendrix.edu)

(1 of 60)

Nashville High School

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My career in science began with a chicken……

(2 of 65)

Arkansas

Female and adolescent orang-utan in Tanjung Puting National Park, Borneo (Sir Graham Banes photo)

A flanged male orang-utan in Tanjung Puting National Park, Borneo (Sir Graham Banes photo)

IUCN RED LIST: CRITICALLY ENDANGERED!!!!!

(3 of 70)

Why do females prefer flanged

males? Are long calls amplified by flanges?

Do flanges just look sexy to the females?

Does size matter?

Are chemical signals involved?

One research project on this topic was

carried out in 1997

• Alejandra Negro. 1997. Studies of the reproductive

development of the male orang-utan and the role

of pheromones and other factors in sexual

maturation. B.S. Thesis, Bryn Mawr College.

(Courtesy of Melanie Bond)

• Dr. Lee Hagey, San Diego Zoo, Q-tip body swabs,

urine from cage floor, 6 samples analyzed (3-orang-

utans);

• Throat, chest (“sternal gland region”), armpits

• Human sweat is odorless until acted upon by

bacteria

• Do apocrine sweat glands produce chemical

signals?

a carboxylic acid: “the odor of mammalian unwashedness”

Caveats………..

I am not a biologist

I am not an expert on orang-utans

I am an organic chemist masquerading as a

chemical ecologist

Chemical ecology is the study of chemicals

involved in the interactions of living organisms.

It focuses on the production of and response to

signalling molecules (semiochemicals) and

toxins.

Pheromone vs. chemical signal (secondary

olfactory system; vomeronasal organ)

Prospecting for Mammalian Chemical Signals via

Green, Solventless Extraction Techniques

Ring-tailed lemur

(Lemur catta) Collaboration with

Dr. Christine Drea,

Duke University

Maned wolf

(Chrysocyon brachyurus) Collaboration with Dr. Nucharin

Songasen, Smithsonian’s Conservation

Biology Institute, and Little Rock Zoo

African elephant

(Loxodonta africana) Collaboration with

Dr. Bruce Schulte,

Western Kentucky University

Bowhead whale (Balaena mysticetus) Collaboration with Dr. Raphaela Stimmelmayr

On-site coordinator 2011 Northern Pinnipeds

Wildlife Veterinarian and Research Biologist NSB

Department of Wildlife Management

Barrow, Alaska

Orang-utan

(Pongo abelii) Collaboration with Dr. Graham Banes

Max Planck Institute for Evolutionary

Anthropology, Germany

Binturong

(Arctictis binturong) Collaboration with

Dr. Christine Drea, Duke University

Chapter 6

The Role of Bacteria in Chemical Signals

of Elephant Musth: Proximate Causes and

Biochemical Pathways

Thomas E. Goodwin, Innocent H. Harelimana, Laura J. MacDonald, Daniel

B. Mark, Aline Umuhire Juru, Qin Yin, James A. Engman, Randall A.

Kopper, Cheryl F. Lichti, Samuel G. Mackintosh, James D. Shoemaker,

Mark V. Sutherland, Alan J. Tackett, Bruce A. Schulte

Elephant musth: a periodic rut-like state

Elevated serum testosterone Swollen & draining temporal glands Urine dribbling Appetite suppression Increased aggression Enhanced reproductive success High concentration of urinary ketones (alkan-2-ones)

(Rasmussen & Wittemyer, Proc. R. Soc. Lond. B 2002, 269, 853-860)—chemical signals

Continued production of ketones exogenously, post-urination as the urine ages (bacterial catabolism)-a time-release chemical signal?

Goodwin et al., J. Chem. Ecol. 2012

Steps for Research on Mammalian

Secretions and Excretions

Procure and ship the samples on dry ice

Store the samples (-60-80o Celsius)

Extract the volatile compounds from the biological matrix

Separate the components of the compound mix

Identify the compounds

Behavioral bioassays of suspected chemical signals

Compared to insects, only a small number of mammalian pheromones have been verified

Solid Phase Dynamic Extraction

AUTOMATED SPDE

• Utilizes adsorption technique for concentrating headspace

analytes by repetitive dynamic flow back and forth over a polymer

coating on the inside wall of a stainless steel syringe needle*

• Rapid, efficient,

solventless,

environmentally benign

Automated SPDE Analysis

• CTC Analytics

CompiPal robot for

headspace SPDE

(EXTRACTION)

•Agilent 6890N GC and

5973N MSD for sample

analysis

Gas Chromatography (SEPARATION) Mass Spectrometry (IDENTIFICATION)

Photograph by G Dallimore of a display at the Oxford University Museum of Natural History.

The text reads: “An inordinate fondness for beetles”…reply of the British polymath J. B. S. Haldane (1892-1964) when asked what could be inferred about the mind of the Creator from a study of the works of Creation.

The Importance of Urine

“…urine necessarily conveys to the external world in its detailed composition much information concerning the internal physiological state of the animal concerned and thus provides the necessary basis for the evolution of specialized semiochemical systems.”

The Importance of Urine

ALBONE, E. S. 1984. Mammalian Semiochemistry. Wiley-Interscience, New York, Chapter Two, p. 165.

Orang-utan Urine Provided by…..

Hesty, Junie, Nias, Robin, & Sally: Denver Zoo

With help by: Cindy Cossaboon

Melati, Tara & Tengku: Fort Wayne Children’s Zoo

With help by: Angie Seltzer, Jen Diehl, Graham Banzz

THANK YOU!!!!

SPDE GC-MS Analysis: Orang-utan Urine Analysis:

Day of thawing from -70o Celsius: 16 compounds

(one ketone)

Aged at room temperature for 5 days: 39

compounds (ten ketones) [ Tara, female, Fort

Wayne]

Day of thawing from -70o Celsius: 7 compounds

(one ketone)

Aged at room temperature for 5 days: 29

compounds (six ketones) [ Junie, male, Denver]

Orang-utan Skin Swabs Provided by…..

Popi, Pebbles, Geri, Kiki, Tango, Louie, Bam Bam,

Radcliffe, Chuckie, Jam, Mari: Center for Great

Apes

With help by: Melanie Bond, “Billy” Graham

Banes

Jake, Hanna, Pumpkin, Peanut, Connie, Sinbad:

Jungle Island

With help by: Linda Jacobs, Graham “Cracker”

Banes

THANK YOU!!!!

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Fre

qu

ency

Compound

FMCH

uFMCH

Figure 1. Comparison of volatile compounds detected in cheek

samples from flanged male orang-utans (FMCH, n = 11) and

unflanged male orang-utans (uFMCH, n = 6) by frequency. All

samples were analyzed by SPDE/GC-MS.

It’s too early to draw any conclusions; We Need More Samples (ultra low temp freezer)

Names of Hendrix College Co-Investigators: Pratheepa Ravikumar, Aline Umuhire-Juru,

Shelly Bradley

International House of Chemistry (IHOC)

His Royal Grahamness Grahamosaurus Baron von Grahamheim Banes, James Banes 007.5

He has many aliases….. He has many affiliations……. Chinese Acad. Sci. Max Planck Institute Cambridge Univ. Univ. of Wisconsin Henry Vilas Zoo

Questions:

Do orang-utans have sternal glands? Are chemical signals in orang-utan sweat? Do flanged male orang-utans have a strong, musty smell?

Mammalian Pheromones

“Our understanding of mammalian pheromones is limited, owing to the complexity of mammalian life and the mammalian brain. Mammals integrate information received by their various senses, and this enormously complicates efforts to understand any specific signal. In addition, interpreting the reaction to a signal can be difficult. The reactions of mammals are not automatic, and their behavior is not necessarily reproducible. Sometimes a pheromone produces no obvious response in a mammal. Perhaps the animal has ignored the signal, or perhaps it has learned something for future use. In general, behavioral studies on mammalian pheromones are much more extensive than chemical research.”---William C. Agosta. 1994 Using Chemicals to Communicate. Journal of Chemical Education. 71: 242-246.

See also: William C. Agosta, Chemical Communication: The Language of Pheromones, Scientific American Library: New York, NY, 1992.

Flanged: fully mature, developed

secondary sexual characteristics

Unflanged: no secondary sexual

characteristics

Secondary sexual characteristics include:

large body size, emission of loud sounds

(“long calls”), large cheek pads (flanges)

Developmental Classes of Male

Orang-utans

Knott, C.D. and Kahlenberg S. 2007. In Primates in Perspectives, S. Beader, C.J. Campbell, A. Fuentes, K.C. and M. Panger, eds. Oxford University Press. pp. 290-305

Maggioncaldda, A.N., Sapolsky, R.M. and Czekala, N.M. 1999. Reproductive hormone profiles in captive male orangutans: Implications for understanding developmental arrest. Am. J. Phys. Anthropol. 109:1932

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1

Fre

qu

ency

Compound

FMCH

FeCH

Figure 2. Comparison of volatile compounds detected in

cheek samples from flanged male orang-utans (FMCH, n =

11) and female orang-utans (FeCH, n = 10) by frequency.

All samples were analyzed by SPDE/GC-MS.

Flanged males tend to be dominant over

unflanged males

Within any orang-utan home range, only

one flanged male becomes the dominant

male

Other flanged males become subordinate

and will move to another home range

Dominant males may suppress

development of secondary sexual

characteristics in unflanged males

Male dominance is expressed to a

considerable extent in the form of

reproductive success

Dominant Male Orang-utans

Knott and Lahlenberg, 2007

Riddles Elephant Sanctuary

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1

Fre

qu

ency

Compound

uFMCH

FeCH

Figure 3. Comparison of volatile compounds detected in cheek

samples from unflanged male orang-utans (uFMCH, n = 6) and female

orang-utans (FeCH, n = 10) by frequency. All samples were analyzed

by SPDE/GC-MS.

Table 2. Compounds detected in six sample types from captive orang-

utans using SPDE/GC-MS. Each row represents the number of samples

of each type that contained the corresponding compound. The total

number of samples of each type is indicated in parentheses.

aCheek samples from flanged males, bcontrol samples from flanged

males, ccheek samples from unflanged males, dcontrol samples from

unflanged males, echeek samples from females, fcontrol samples from

females gA dash indicates non-detectable amounts